zfx  (Cell Signaling Technology Inc)


Bioz Verified Symbol Cell Signaling Technology Inc is a verified supplier
Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94

    Structured Review

    Cell Signaling Technology Inc zfx
    Zfx, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/zfx/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    zfx - by Bioz Stars, 2023-01
    94/100 stars

    Images

    zfx  (Cell Signaling Technology Inc)


    Bioz Verified Symbol Cell Signaling Technology Inc is a verified supplier
    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94

    Structured Review

    Cell Signaling Technology Inc zfx
    Zfx, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/zfx/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    zfx - by Bioz Stars, 2023-01
    94/100 stars

    Images

    zfx  (Cell Signaling Technology Inc)


    Bioz Verified Symbol Cell Signaling Technology Inc is a verified supplier
    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94

    Structured Review

    Cell Signaling Technology Inc zfx
    Zfx, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/zfx/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    zfx - by Bioz Stars, 2023-01
    94/100 stars

    Images

    antibodies against zfx  (Cell Signaling Technology Inc)


    Bioz Verified Symbol Cell Signaling Technology Inc is a verified supplier
    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94

    Structured Review

    Cell Signaling Technology Inc antibodies against zfx
    Antibodies Against Zfx, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/antibodies against zfx/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    antibodies against zfx - by Bioz Stars, 2023-01
    94/100 stars

    Images

    zfx  (Cell Signaling Technology Inc)


    Bioz Verified Symbol Cell Signaling Technology Inc is a verified supplier
    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94

    Structured Review

    Cell Signaling Technology Inc zfx
    The <t>ZFX</t> gene family. Shown are gene structure schematics for ZFX, ZFY <t>and</t> <t>ZNF711.</t> Dashed lines indicate zinc fingers conserved between ZFX and the other two family members. NLS: nuclear localization sequence.
    Zfx, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/zfx/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    zfx - by Bioz Stars, 2023-01
    94/100 stars

    Images

    1) Product Images from "Characterization of the ZFX family of transcription factors that bind downstream of the start site of CpG island promoters"

    Article Title: Characterization of the ZFX family of transcription factors that bind downstream of the start site of CpG island promoters

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkaa384

    The ZFX gene family. Shown are gene structure schematics for ZFX, ZFY and ZNF711. Dashed lines indicate zinc fingers conserved between ZFX and the other two family members. NLS: nuclear localization sequence.
    Figure Legend Snippet: The ZFX gene family. Shown are gene structure schematics for ZFX, ZFY and ZNF711. Dashed lines indicate zinc fingers conserved between ZFX and the other two family members. NLS: nuclear localization sequence.

    Techniques Used: Zinc-Fingers, Sequencing

    Loss of ZFX and ZNF711 in HEK293T cells inhibits cell proliferation. ( A ) Expression levels of ZFX/ZFY/ZNF711 in wt HEK293T cells. ( B ) Locations of gRNAs used to create CRISPR/Cas9-mediated ZFX and/or ZNF711 knockouts. The deletion of ZFX in ZFX KO clone1 and clone2 and the DKO clones were generated using ZFX gRNA1 and gRNA2. The deletion of ZNF711 in ZNF711 KO clone1 and the DKO clones was generated using ZNF711 gRNA1 and gRNA2; the deletion of ZNF711 KO clone2 was generated using ZNF711 gRNA2 and gRNA3. ( C ) Western blots showing the protein levels of ZFX and ZNF711 in wt HEK293T, ZFX KO clones, ZNF711 KO clones, and DKO clones; also shown is the level of p62 as a loading control. ( D ) Proliferation assays using wt HEK293T, two different ZFX and two different ZNF711 KO clones, and two DKO clones; data points are the mean of three biological replicates.
    Figure Legend Snippet: Loss of ZFX and ZNF711 in HEK293T cells inhibits cell proliferation. ( A ) Expression levels of ZFX/ZFY/ZNF711 in wt HEK293T cells. ( B ) Locations of gRNAs used to create CRISPR/Cas9-mediated ZFX and/or ZNF711 knockouts. The deletion of ZFX in ZFX KO clone1 and clone2 and the DKO clones were generated using ZFX gRNA1 and gRNA2. The deletion of ZNF711 in ZNF711 KO clone1 and the DKO clones was generated using ZNF711 gRNA1 and gRNA2; the deletion of ZNF711 KO clone2 was generated using ZNF711 gRNA2 and gRNA3. ( C ) Western blots showing the protein levels of ZFX and ZNF711 in wt HEK293T, ZFX KO clones, ZNF711 KO clones, and DKO clones; also shown is the level of p62 as a loading control. ( D ) Proliferation assays using wt HEK293T, two different ZFX and two different ZNF711 KO clones, and two DKO clones; data points are the mean of three biological replicates.

    Techniques Used: Expressing, CRISPR, Clone Assay, Generated, Western Blot

    Reduction in ZFX and ZNF711 levels causes large effects on the transcriptome. ( A ) Volcano plots showing the differentially expressed genes (DEGs) identified via RNA-seq in comparisons of wt HEK293T versus ZFX KO clone1, KO clone2, ZNF711 KO clone1, KO clone2, DKO clone1, DKO clone2, or DKO clone3. ( B ) Comparison of DEGs commonly downregulated or upregulated in all three DKO clones. ( C ) Gene ontology analysis of the 1166 commonly downregulated and 2124 commonly upregulated genes in all three DKO clones.
    Figure Legend Snippet: Reduction in ZFX and ZNF711 levels causes large effects on the transcriptome. ( A ) Volcano plots showing the differentially expressed genes (DEGs) identified via RNA-seq in comparisons of wt HEK293T versus ZFX KO clone1, KO clone2, ZNF711 KO clone1, KO clone2, DKO clone1, DKO clone2, or DKO clone3. ( B ) Comparison of DEGs commonly downregulated or upregulated in all three DKO clones. ( C ) Gene ontology analysis of the 1166 commonly downregulated and 2124 commonly upregulated genes in all three DKO clones.

    Techniques Used: RNA Sequencing Assay, Clone Assay

    ZFX family members have essentially identical binding patterns at CpG island promoters. ( A ) Browser tracks showing ZFX family member binding profiles in female HEK293T kidney cells and male 22Rv1 prostate cells. Also shown is a zoom in on a single peak located in the DOCK7 promoter region. ( B ) Shown is a heatmap illustrating the genome-wide correlation of ZFX family member binding patterns in 22Rv1 cells. ( C ) Bar graph of genomic distributions of ZFX family member binding sites in 22Rv1 cells in promoter and non-promoter regions (left) and bar graph showing the relative distribution of binding sites in CpG island (CGI) promoters and non-CpG island promoters (right). ( D ) Venn diagram comparing the sets of CpG island promoters bound by ZFX, ZFY and ZNF711 in 22Rv1 cells.
    Figure Legend Snippet: ZFX family members have essentially identical binding patterns at CpG island promoters. ( A ) Browser tracks showing ZFX family member binding profiles in female HEK293T kidney cells and male 22Rv1 prostate cells. Also shown is a zoom in on a single peak located in the DOCK7 promoter region. ( B ) Shown is a heatmap illustrating the genome-wide correlation of ZFX family member binding patterns in 22Rv1 cells. ( C ) Bar graph of genomic distributions of ZFX family member binding sites in 22Rv1 cells in promoter and non-promoter regions (left) and bar graph showing the relative distribution of binding sites in CpG island (CGI) promoters and non-CpG island promoters (right). ( D ) Venn diagram comparing the sets of CpG island promoters bound by ZFX, ZFY and ZNF711 in 22Rv1 cells.

    Techniques Used: Binding Assay, Genome Wide

    ZFX and ZNF711 have properties of a transcription activator when bound downstream of the TSS. ( A ) ZFX and ZNF711 peak sets from HEK293T cells were clustered using K-means clustering, identifying four sets of peaks with distinct binding sites (left); cluster 4 (combination peaks) was subsequently re-clustered, identifying 4 subsets (right). Tag density plots for each of the 4 different clusters are presented on top of the heatmaps. ( B ) Average signals of ZFX and ZNF711 ChIP-seq reads in wt HEK293T at all promoters bound by each TF (top), promoters of genes with decreased expression in all three DKO clones (middle), and promoters of genes with increased expressions in all three DKO clones (bottom). Also shown, for both the downregulated and the upregulated gene categories, is the percentage of genes whose promoters are bound by ZFX or ZNF711 in peak categories 1–4, or not bound by ZFX or ZNF711.
    Figure Legend Snippet: ZFX and ZNF711 have properties of a transcription activator when bound downstream of the TSS. ( A ) ZFX and ZNF711 peak sets from HEK293T cells were clustered using K-means clustering, identifying four sets of peaks with distinct binding sites (left); cluster 4 (combination peaks) was subsequently re-clustered, identifying 4 subsets (right). Tag density plots for each of the 4 different clusters are presented on top of the heatmaps. ( B ) Average signals of ZFX and ZNF711 ChIP-seq reads in wt HEK293T at all promoters bound by each TF (top), promoters of genes with decreased expression in all three DKO clones (middle), and promoters of genes with increased expressions in all three DKO clones (bottom). Also shown, for both the downregulated and the upregulated gene categories, is the percentage of genes whose promoters are bound by ZFX or ZNF711 in peak categories 1–4, or not bound by ZFX or ZNF711.

    Techniques Used: Binding Assay, ChIP-sequencing, Expressing, Clone Assay

    Characterization of ZFX and ZNF711 binding sites. ( A ) Classification of binding sites based on genomic locations of all ZFX peaks located downstream of the TSS and ZFX downstream peaks at promoters of genes down-regulated in all 3 DKO clones. ( B ) Classification of ZNF711 downstream ChIP-seq peaks, downstream peaks identified using read2 of the ChIP-exo dataset, and downstream peaks identified by the ChexMix program (+/-10 nt from the nt identified as the binding site by the program). ( C ) Tag density plots of all ZNF711 peaks from standard ChIP-seq and from ChIP-exo. ( D ) Motif analysis using the top 5000 peaks identified from standard ZNF711 ChIP-seq (average width 1800 nt), ChIP-exo read2 (average width 300 nt), ChIP-exo by the ChexMix program (20 nt), and randomized CpG island promoter regions (width 2 kb, 200 bp, and 20 nt). The peaks were searched for the known ZNF711 motif and the 5 nt motif identified by ChIP-exo. ( E) Zoom-in comparison of peaks from ZNF711 standard ChIP-seq replicates and ChIP-exo replicates.
    Figure Legend Snippet: Characterization of ZFX and ZNF711 binding sites. ( A ) Classification of binding sites based on genomic locations of all ZFX peaks located downstream of the TSS and ZFX downstream peaks at promoters of genes down-regulated in all 3 DKO clones. ( B ) Classification of ZNF711 downstream ChIP-seq peaks, downstream peaks identified using read2 of the ChIP-exo dataset, and downstream peaks identified by the ChexMix program (+/-10 nt from the nt identified as the binding site by the program). ( C ) Tag density plots of all ZNF711 peaks from standard ChIP-seq and from ChIP-exo. ( D ) Motif analysis using the top 5000 peaks identified from standard ZNF711 ChIP-seq (average width 1800 nt), ChIP-exo read2 (average width 300 nt), ChIP-exo by the ChexMix program (20 nt), and randomized CpG island promoter regions (width 2 kb, 200 bp, and 20 nt). The peaks were searched for the known ZNF711 motif and the 5 nt motif identified by ChIP-exo. ( E) Zoom-in comparison of peaks from ZNF711 standard ChIP-seq replicates and ChIP-exo replicates.

    Techniques Used: Binding Assay, Clone Assay, ChIP-sequencing

    Functional analysis of the ZFX protein. ( A ) Schematic of FLAG-tagged ZFX zinc finger (ZF) mutant constructs. ( B ) Browser tracks showing genomic binding profiles of endogenous ZFX and FLAG-tagged wt ZFX and ZFX mutants in HEK293T cells. ( C ) Tag density plots of ChIP-seq peaks comparing endogenous ZFX and ZFX ZF9-13 peak locations in HEK293T cells. ( D ) Heatmaps showing ChIP-seq data from FLAG-tagged wt ZFX and ZFX ZF9-13 centered on the genomic locations of the endogenous ZFX peaks. ( E ) Expression levels following transfection with different ZFX constructs (as analyzed by RT-qPCR) of two genes (LONRF2 and CAPN2) whose promoters are bound by both ZFX and ZNF711 in wt HEK293T cells and which show a reduction in gene expression in all three DKO clones, of one gene (FOS) that is upregulated in all three DKO cells (a putative indirect target gene), and of one gene (HOXC4) that shows no expression changes in the DKO cells. Expression data were normalized to the control (cells transfected with an unrelated plasmid). Three independent experiments were performed using two different clonal populations of DKO cells; data points represent results from triplicate wells and duplicate PCR readings. Error bars indicate the pooled standard deviations of the means for the constructs and for the normalizing control.
    Figure Legend Snippet: Functional analysis of the ZFX protein. ( A ) Schematic of FLAG-tagged ZFX zinc finger (ZF) mutant constructs. ( B ) Browser tracks showing genomic binding profiles of endogenous ZFX and FLAG-tagged wt ZFX and ZFX mutants in HEK293T cells. ( C ) Tag density plots of ChIP-seq peaks comparing endogenous ZFX and ZFX ZF9-13 peak locations in HEK293T cells. ( D ) Heatmaps showing ChIP-seq data from FLAG-tagged wt ZFX and ZFX ZF9-13 centered on the genomic locations of the endogenous ZFX peaks. ( E ) Expression levels following transfection with different ZFX constructs (as analyzed by RT-qPCR) of two genes (LONRF2 and CAPN2) whose promoters are bound by both ZFX and ZNF711 in wt HEK293T cells and which show a reduction in gene expression in all three DKO clones, of one gene (FOS) that is upregulated in all three DKO cells (a putative indirect target gene), and of one gene (HOXC4) that shows no expression changes in the DKO cells. Expression data were normalized to the control (cells transfected with an unrelated plasmid). Three independent experiments were performed using two different clonal populations of DKO cells; data points represent results from triplicate wells and duplicate PCR readings. Error bars indicate the pooled standard deviations of the means for the constructs and for the normalizing control.

    Techniques Used: Functional Assay, Mutagenesis, Construct, Binding Assay, ChIP-sequencing, Expressing, Transfection, Quantitative RT-PCR, Clone Assay, Plasmid Preparation

    ZFX ZF11-13 has very similar transcriptional activities as wt ZFX. ( A ) Volcano plots showing the differentially expressed genes (DEGs) identified via RNA-seq in comparisons of DKO cells 24 hr after transfection with FLAG-tagged wt ZFX vs. a control plasmid, FLAG-tagged ZFX ZF11-13 vs. a control plasmid, and FLAG-tagged wt ZFX vs. FLAG-tagged ZFX ZF11-13. ( B ) Shown is a Venn diagram comparing the 846 genes that are bound by ZFX and ZNF711 in wt HEK293T cells and show a decrease in mRNA levels in all three DKO clones and the 2275 genes that show increased levels in DKO cells transfected with either FLAG-tagged wt ZFX or ZFX ZF11-13. ( C ) Shown is a tag density plot of ChIP-seq data for endogenous ZFX, FLAG-tagged wt ZFX, ZFX ZF9-13, or ZFX ZF1-8 at the set of 277 responding promoters identified in panel B. ( D ) Motif coverage analysis of the 277 responding promoters.
    Figure Legend Snippet: ZFX ZF11-13 has very similar transcriptional activities as wt ZFX. ( A ) Volcano plots showing the differentially expressed genes (DEGs) identified via RNA-seq in comparisons of DKO cells 24 hr after transfection with FLAG-tagged wt ZFX vs. a control plasmid, FLAG-tagged ZFX ZF11-13 vs. a control plasmid, and FLAG-tagged wt ZFX vs. FLAG-tagged ZFX ZF11-13. ( B ) Shown is a Venn diagram comparing the 846 genes that are bound by ZFX and ZNF711 in wt HEK293T cells and show a decrease in mRNA levels in all three DKO clones and the 2275 genes that show increased levels in DKO cells transfected with either FLAG-tagged wt ZFX or ZFX ZF11-13. ( C ) Shown is a tag density plot of ChIP-seq data for endogenous ZFX, FLAG-tagged wt ZFX, ZFX ZF9-13, or ZFX ZF1-8 at the set of 277 responding promoters identified in panel B. ( D ) Motif coverage analysis of the 277 responding promoters.

    Techniques Used: RNA Sequencing Assay, Transfection, Plasmid Preparation, Clone Assay, ChIP-sequencing

    zfx antibody  (Cell Signaling Technology Inc)


    Bioz Verified Symbol Cell Signaling Technology Inc is a verified supplier
    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94

    Structured Review

    Cell Signaling Technology Inc zfx antibody
    The <t>ZFX</t> gene family. Shown are gene structure schematics for ZFX, ZFY <t>and</t> <t>ZNF711.</t> Dashed lines indicate zinc fingers conserved between ZFX and the other two family members. NLS: nuclear localization sequence.
    Zfx Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/zfx antibody/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    zfx antibody - by Bioz Stars, 2023-01
    94/100 stars

    Images

    1) Product Images from "Characterization of the ZFX family of transcription factors that bind downstream of the start site of CpG island promoters"

    Article Title: Characterization of the ZFX family of transcription factors that bind downstream of the start site of CpG island promoters

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkaa384

    The ZFX gene family. Shown are gene structure schematics for ZFX, ZFY and ZNF711. Dashed lines indicate zinc fingers conserved between ZFX and the other two family members. NLS: nuclear localization sequence.
    Figure Legend Snippet: The ZFX gene family. Shown are gene structure schematics for ZFX, ZFY and ZNF711. Dashed lines indicate zinc fingers conserved between ZFX and the other two family members. NLS: nuclear localization sequence.

    Techniques Used: Zinc-Fingers, Sequencing

    Loss of ZFX and ZNF711 in HEK293T cells inhibits cell proliferation. ( A ) Expression levels of ZFX/ZFY/ZNF711 in wt HEK293T cells. ( B ) Locations of gRNAs used to create CRISPR/Cas9-mediated ZFX and/or ZNF711 knockouts. The deletion of ZFX in ZFX KO clone1 and clone2 and the DKO clones were generated using ZFX gRNA1 and gRNA2. The deletion of ZNF711 in ZNF711 KO clone1 and the DKO clones was generated using ZNF711 gRNA1 and gRNA2; the deletion of ZNF711 KO clone2 was generated using ZNF711 gRNA2 and gRNA3. ( C ) Western blots showing the protein levels of ZFX and ZNF711 in wt HEK293T, ZFX KO clones, ZNF711 KO clones, and DKO clones; also shown is the level of p62 as a loading control. ( D ) Proliferation assays using wt HEK293T, two different ZFX and two different ZNF711 KO clones, and two DKO clones; data points are the mean of three biological replicates.
    Figure Legend Snippet: Loss of ZFX and ZNF711 in HEK293T cells inhibits cell proliferation. ( A ) Expression levels of ZFX/ZFY/ZNF711 in wt HEK293T cells. ( B ) Locations of gRNAs used to create CRISPR/Cas9-mediated ZFX and/or ZNF711 knockouts. The deletion of ZFX in ZFX KO clone1 and clone2 and the DKO clones were generated using ZFX gRNA1 and gRNA2. The deletion of ZNF711 in ZNF711 KO clone1 and the DKO clones was generated using ZNF711 gRNA1 and gRNA2; the deletion of ZNF711 KO clone2 was generated using ZNF711 gRNA2 and gRNA3. ( C ) Western blots showing the protein levels of ZFX and ZNF711 in wt HEK293T, ZFX KO clones, ZNF711 KO clones, and DKO clones; also shown is the level of p62 as a loading control. ( D ) Proliferation assays using wt HEK293T, two different ZFX and two different ZNF711 KO clones, and two DKO clones; data points are the mean of three biological replicates.

    Techniques Used: Expressing, CRISPR, Clone Assay, Generated, Western Blot

    Reduction in ZFX and ZNF711 levels causes large effects on the transcriptome. ( A ) Volcano plots showing the differentially expressed genes (DEGs) identified via RNA-seq in comparisons of wt HEK293T versus ZFX KO clone1, KO clone2, ZNF711 KO clone1, KO clone2, DKO clone1, DKO clone2, or DKO clone3. ( B ) Comparison of DEGs commonly downregulated or upregulated in all three DKO clones. ( C ) Gene ontology analysis of the 1166 commonly downregulated and 2124 commonly upregulated genes in all three DKO clones.
    Figure Legend Snippet: Reduction in ZFX and ZNF711 levels causes large effects on the transcriptome. ( A ) Volcano plots showing the differentially expressed genes (DEGs) identified via RNA-seq in comparisons of wt HEK293T versus ZFX KO clone1, KO clone2, ZNF711 KO clone1, KO clone2, DKO clone1, DKO clone2, or DKO clone3. ( B ) Comparison of DEGs commonly downregulated or upregulated in all three DKO clones. ( C ) Gene ontology analysis of the 1166 commonly downregulated and 2124 commonly upregulated genes in all three DKO clones.

    Techniques Used: RNA Sequencing Assay, Clone Assay

    ZFX family members have essentially identical binding patterns at CpG island promoters. ( A ) Browser tracks showing ZFX family member binding profiles in female HEK293T kidney cells and male 22Rv1 prostate cells. Also shown is a zoom in on a single peak located in the DOCK7 promoter region. ( B ) Shown is a heatmap illustrating the genome-wide correlation of ZFX family member binding patterns in 22Rv1 cells. ( C ) Bar graph of genomic distributions of ZFX family member binding sites in 22Rv1 cells in promoter and non-promoter regions (left) and bar graph showing the relative distribution of binding sites in CpG island (CGI) promoters and non-CpG island promoters (right). ( D ) Venn diagram comparing the sets of CpG island promoters bound by ZFX, ZFY and ZNF711 in 22Rv1 cells.
    Figure Legend Snippet: ZFX family members have essentially identical binding patterns at CpG island promoters. ( A ) Browser tracks showing ZFX family member binding profiles in female HEK293T kidney cells and male 22Rv1 prostate cells. Also shown is a zoom in on a single peak located in the DOCK7 promoter region. ( B ) Shown is a heatmap illustrating the genome-wide correlation of ZFX family member binding patterns in 22Rv1 cells. ( C ) Bar graph of genomic distributions of ZFX family member binding sites in 22Rv1 cells in promoter and non-promoter regions (left) and bar graph showing the relative distribution of binding sites in CpG island (CGI) promoters and non-CpG island promoters (right). ( D ) Venn diagram comparing the sets of CpG island promoters bound by ZFX, ZFY and ZNF711 in 22Rv1 cells.

    Techniques Used: Binding Assay, Genome Wide

    ZFX and ZNF711 have properties of a transcription activator when bound downstream of the TSS. ( A ) ZFX and ZNF711 peak sets from HEK293T cells were clustered using K-means clustering, identifying four sets of peaks with distinct binding sites (left); cluster 4 (combination peaks) was subsequently re-clustered, identifying 4 subsets (right). Tag density plots for each of the 4 different clusters are presented on top of the heatmaps. ( B ) Average signals of ZFX and ZNF711 ChIP-seq reads in wt HEK293T at all promoters bound by each TF (top), promoters of genes with decreased expression in all three DKO clones (middle), and promoters of genes with increased expressions in all three DKO clones (bottom). Also shown, for both the downregulated and the upregulated gene categories, is the percentage of genes whose promoters are bound by ZFX or ZNF711 in peak categories 1–4, or not bound by ZFX or ZNF711.
    Figure Legend Snippet: ZFX and ZNF711 have properties of a transcription activator when bound downstream of the TSS. ( A ) ZFX and ZNF711 peak sets from HEK293T cells were clustered using K-means clustering, identifying four sets of peaks with distinct binding sites (left); cluster 4 (combination peaks) was subsequently re-clustered, identifying 4 subsets (right). Tag density plots for each of the 4 different clusters are presented on top of the heatmaps. ( B ) Average signals of ZFX and ZNF711 ChIP-seq reads in wt HEK293T at all promoters bound by each TF (top), promoters of genes with decreased expression in all three DKO clones (middle), and promoters of genes with increased expressions in all three DKO clones (bottom). Also shown, for both the downregulated and the upregulated gene categories, is the percentage of genes whose promoters are bound by ZFX or ZNF711 in peak categories 1–4, or not bound by ZFX or ZNF711.

    Techniques Used: Binding Assay, ChIP-sequencing, Expressing, Clone Assay

    Characterization of ZFX and ZNF711 binding sites. ( A ) Classification of binding sites based on genomic locations of all ZFX peaks located downstream of the TSS and ZFX downstream peaks at promoters of genes down-regulated in all 3 DKO clones. ( B ) Classification of ZNF711 downstream ChIP-seq peaks, downstream peaks identified using read2 of the ChIP-exo dataset, and downstream peaks identified by the ChexMix program (+/-10 nt from the nt identified as the binding site by the program). ( C ) Tag density plots of all ZNF711 peaks from standard ChIP-seq and from ChIP-exo. ( D ) Motif analysis using the top 5000 peaks identified from standard ZNF711 ChIP-seq (average width 1800 nt), ChIP-exo read2 (average width 300 nt), ChIP-exo by the ChexMix program (20 nt), and randomized CpG island promoter regions (width 2 kb, 200 bp, and 20 nt). The peaks were searched for the known ZNF711 motif and the 5 nt motif identified by ChIP-exo. ( E) Zoom-in comparison of peaks from ZNF711 standard ChIP-seq replicates and ChIP-exo replicates.
    Figure Legend Snippet: Characterization of ZFX and ZNF711 binding sites. ( A ) Classification of binding sites based on genomic locations of all ZFX peaks located downstream of the TSS and ZFX downstream peaks at promoters of genes down-regulated in all 3 DKO clones. ( B ) Classification of ZNF711 downstream ChIP-seq peaks, downstream peaks identified using read2 of the ChIP-exo dataset, and downstream peaks identified by the ChexMix program (+/-10 nt from the nt identified as the binding site by the program). ( C ) Tag density plots of all ZNF711 peaks from standard ChIP-seq and from ChIP-exo. ( D ) Motif analysis using the top 5000 peaks identified from standard ZNF711 ChIP-seq (average width 1800 nt), ChIP-exo read2 (average width 300 nt), ChIP-exo by the ChexMix program (20 nt), and randomized CpG island promoter regions (width 2 kb, 200 bp, and 20 nt). The peaks were searched for the known ZNF711 motif and the 5 nt motif identified by ChIP-exo. ( E) Zoom-in comparison of peaks from ZNF711 standard ChIP-seq replicates and ChIP-exo replicates.

    Techniques Used: Binding Assay, Clone Assay, ChIP-sequencing

    Functional analysis of the ZFX protein. ( A ) Schematic of FLAG-tagged ZFX zinc finger (ZF) mutant constructs. ( B ) Browser tracks showing genomic binding profiles of endogenous ZFX and FLAG-tagged wt ZFX and ZFX mutants in HEK293T cells. ( C ) Tag density plots of ChIP-seq peaks comparing endogenous ZFX and ZFX ZF9-13 peak locations in HEK293T cells. ( D ) Heatmaps showing ChIP-seq data from FLAG-tagged wt ZFX and ZFX ZF9-13 centered on the genomic locations of the endogenous ZFX peaks. ( E ) Expression levels following transfection with different ZFX constructs (as analyzed by RT-qPCR) of two genes (LONRF2 and CAPN2) whose promoters are bound by both ZFX and ZNF711 in wt HEK293T cells and which show a reduction in gene expression in all three DKO clones, of one gene (FOS) that is upregulated in all three DKO cells (a putative indirect target gene), and of one gene (HOXC4) that shows no expression changes in the DKO cells. Expression data were normalized to the control (cells transfected with an unrelated plasmid). Three independent experiments were performed using two different clonal populations of DKO cells; data points represent results from triplicate wells and duplicate PCR readings. Error bars indicate the pooled standard deviations of the means for the constructs and for the normalizing control.
    Figure Legend Snippet: Functional analysis of the ZFX protein. ( A ) Schematic of FLAG-tagged ZFX zinc finger (ZF) mutant constructs. ( B ) Browser tracks showing genomic binding profiles of endogenous ZFX and FLAG-tagged wt ZFX and ZFX mutants in HEK293T cells. ( C ) Tag density plots of ChIP-seq peaks comparing endogenous ZFX and ZFX ZF9-13 peak locations in HEK293T cells. ( D ) Heatmaps showing ChIP-seq data from FLAG-tagged wt ZFX and ZFX ZF9-13 centered on the genomic locations of the endogenous ZFX peaks. ( E ) Expression levels following transfection with different ZFX constructs (as analyzed by RT-qPCR) of two genes (LONRF2 and CAPN2) whose promoters are bound by both ZFX and ZNF711 in wt HEK293T cells and which show a reduction in gene expression in all three DKO clones, of one gene (FOS) that is upregulated in all three DKO cells (a putative indirect target gene), and of one gene (HOXC4) that shows no expression changes in the DKO cells. Expression data were normalized to the control (cells transfected with an unrelated plasmid). Three independent experiments were performed using two different clonal populations of DKO cells; data points represent results from triplicate wells and duplicate PCR readings. Error bars indicate the pooled standard deviations of the means for the constructs and for the normalizing control.

    Techniques Used: Functional Assay, Mutagenesis, Construct, Binding Assay, ChIP-sequencing, Expressing, Transfection, Quantitative RT-PCR, Clone Assay, Plasmid Preparation

    ZFX ZF11-13 has very similar transcriptional activities as wt ZFX. ( A ) Volcano plots showing the differentially expressed genes (DEGs) identified via RNA-seq in comparisons of DKO cells 24 hr after transfection with FLAG-tagged wt ZFX vs. a control plasmid, FLAG-tagged ZFX ZF11-13 vs. a control plasmid, and FLAG-tagged wt ZFX vs. FLAG-tagged ZFX ZF11-13. ( B ) Shown is a Venn diagram comparing the 846 genes that are bound by ZFX and ZNF711 in wt HEK293T cells and show a decrease in mRNA levels in all three DKO clones and the 2275 genes that show increased levels in DKO cells transfected with either FLAG-tagged wt ZFX or ZFX ZF11-13. ( C ) Shown is a tag density plot of ChIP-seq data for endogenous ZFX, FLAG-tagged wt ZFX, ZFX ZF9-13, or ZFX ZF1-8 at the set of 277 responding promoters identified in panel B. ( D ) Motif coverage analysis of the 277 responding promoters.
    Figure Legend Snippet: ZFX ZF11-13 has very similar transcriptional activities as wt ZFX. ( A ) Volcano plots showing the differentially expressed genes (DEGs) identified via RNA-seq in comparisons of DKO cells 24 hr after transfection with FLAG-tagged wt ZFX vs. a control plasmid, FLAG-tagged ZFX ZF11-13 vs. a control plasmid, and FLAG-tagged wt ZFX vs. FLAG-tagged ZFX ZF11-13. ( B ) Shown is a Venn diagram comparing the 846 genes that are bound by ZFX and ZNF711 in wt HEK293T cells and show a decrease in mRNA levels in all three DKO clones and the 2275 genes that show increased levels in DKO cells transfected with either FLAG-tagged wt ZFX or ZFX ZF11-13. ( C ) Shown is a tag density plot of ChIP-seq data for endogenous ZFX, FLAG-tagged wt ZFX, ZFX ZF9-13, or ZFX ZF1-8 at the set of 277 responding promoters identified in panel B. ( D ) Motif coverage analysis of the 277 responding promoters.

    Techniques Used: RNA Sequencing Assay, Transfection, Plasmid Preparation, Clone Assay, ChIP-sequencing

    anti tubulin dm1a mouse mab cell signaling tech  (Cell Signaling Technology Inc)


    Bioz Verified Symbol Cell Signaling Technology Inc is a verified supplier
    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94

    Structured Review

    Cell Signaling Technology Inc anti tubulin dm1a mouse mab cell signaling tech
    KEY RESOURCES TABLE
    Anti Tubulin Dm1a Mouse Mab Cell Signaling Tech, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti tubulin dm1a mouse mab cell signaling tech/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    anti tubulin dm1a mouse mab cell signaling tech - by Bioz Stars, 2023-01
    94/100 stars

    Images

    1) Product Images from "ZFX mediates non-canonical oncogenic functions of the androgen receptor splice variant 7 (AR-V7) in castrate-resistant prostate cancer"

    Article Title: ZFX mediates non-canonical oncogenic functions of the androgen receptor splice variant 7 (AR-V7) in castrate-resistant prostate cancer

    Journal: Molecular cell

    doi: 10.1016/j.molcel.2018.08.029

    KEY RESOURCES TABLE
    Figure Legend Snippet: KEY RESOURCES TABLE

    Techniques Used: Immunoprecipitation, Recombinant, Electron Microscopy, Transfection, Protease Inhibitor, SYBR Green Assay, Purification, Multiplex Assay, Proliferation Assay, Mutagenesis, Magnetic Beads, Western Blot, Stripping, shRNA, Construct, Plasmid Preparation, Software

    zfx  (Cell Signaling Technology Inc)


    Bioz Verified Symbol Cell Signaling Technology Inc is a verified supplier
    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94

    Structured Review

    Cell Signaling Technology Inc zfx
    KEY RESOURCES TABLE
    Zfx, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/zfx/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    zfx - by Bioz Stars, 2023-01
    94/100 stars

    Images

    1) Product Images from "ZFX mediates non-canonical oncogenic functions of the androgen receptor splice variant 7 (AR-V7) in castrate-resistant prostate cancer"

    Article Title: ZFX mediates non-canonical oncogenic functions of the androgen receptor splice variant 7 (AR-V7) in castrate-resistant prostate cancer

    Journal: Molecular cell

    doi: 10.1016/j.molcel.2018.08.029

    KEY RESOURCES TABLE
    Figure Legend Snippet: KEY RESOURCES TABLE

    Techniques Used: Immunoprecipitation, Recombinant, Electron Microscopy, Transfection, Protease Inhibitor, SYBR Green Assay, Purification, Multiplex Assay, Proliferation Assay, Mutagenesis, Magnetic Beads, Western Blot, Stripping, shRNA, Construct, Plasmid Preparation, Software

    mouse anti zfx  (Cell Signaling Technology Inc)


    Bioz Verified Symbol Cell Signaling Technology Inc is a verified supplier
    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94

    Structured Review

    Cell Signaling Technology Inc mouse anti zfx
    KEY RESOURCES TABLE
    Mouse Anti Zfx, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse anti zfx/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mouse anti zfx - by Bioz Stars, 2023-01
    94/100 stars

    Images

    1) Product Images from "ZFX mediates non-canonical oncogenic functions of the androgen receptor splice variant 7 (AR-V7) in castrate-resistant prostate cancer"

    Article Title: ZFX mediates non-canonical oncogenic functions of the androgen receptor splice variant 7 (AR-V7) in castrate-resistant prostate cancer

    Journal: Molecular cell

    doi: 10.1016/j.molcel.2018.08.029

    KEY RESOURCES TABLE
    Figure Legend Snippet: KEY RESOURCES TABLE

    Techniques Used: Immunoprecipitation, Recombinant, Electron Microscopy, Transfection, Protease Inhibitor, SYBR Green Assay, Purification, Multiplex Assay, Proliferation Assay, Mutagenesis, Magnetic Beads, Western Blot, Stripping, shRNA, Construct, Plasmid Preparation, Software

    2008 na mouse anti zfx cell signaling mab  (Cell Signaling Technology Inc)


    Bioz Verified Symbol Cell Signaling Technology Inc is a verified supplier
    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94

    Structured Review

    Cell Signaling Technology Inc 2008 na mouse anti zfx cell signaling mab
    KEY RESOURCES TABLE
    2008 Na Mouse Anti Zfx Cell Signaling Mab, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/2008 na mouse anti zfx cell signaling mab/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    2008 na mouse anti zfx cell signaling mab - by Bioz Stars, 2023-01
    94/100 stars

    Images

    1) Product Images from "ZFX mediates non-canonical oncogenic functions of the androgen receptor splice variant 7 (AR-V7) in castrate-resistant prostate cancer"

    Article Title: ZFX mediates non-canonical oncogenic functions of the androgen receptor splice variant 7 (AR-V7) in castrate-resistant prostate cancer

    Journal: Molecular cell

    doi: 10.1016/j.molcel.2018.08.029

    KEY RESOURCES TABLE
    Figure Legend Snippet: KEY RESOURCES TABLE

    Techniques Used: Immunoprecipitation, Recombinant, Electron Microscopy, Transfection, Protease Inhibitor, SYBR Green Assay, Purification, Multiplex Assay, Proliferation Assay, Mutagenesis, Magnetic Beads, Western Blot, Stripping, shRNA, Construct, Plasmid Preparation, Software

    dusp4  (Cell Signaling Technology Inc)


    Bioz Verified Symbol Cell Signaling Technology Inc is a verified supplier
    Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94

    Structured Review

    Cell Signaling Technology Inc dusp4
    A) Cancer Cell Line Encyclopedia data SNP copy number analysis for the <t>DUSP4</t> locus across 964 cancer cell lines from various tissue sources (49). Red bars show the median log2 copy number for each tissue origin. B) Left panel; frequency histogram of TCGA (14) SNP copy number analysis for the DUSP4 locus for 443 breast cancers normalized to patient matched control tissue (log2 ratio). Right panel; DUSP4 log2 copy number ratio plotted by molecular subtype as determined by gene expression using the PAM50 centroids. P-value represents the ANOVA result. C) CD44:CD24 mRNA ratio for the ICBP50 panel of breast cancer cell lines, as determined by microarray data, plotted vs. the MEK activation score of Pratilas et al. (20). D) Unsupervised clustering of RNA from 15 mammosphere cultures vs. 11 primary tumor profiles, representing 16 patients including 10 patient pairs (7) using the genes defined in the MEK signature (20). E) Box and whisker plot of the data in (D), according to the total sum score of the MEK signature. P-value represents the result of a two-tailed t-test. F) CD44:CD24 mRNA ratio for the ICBP50 panel of breast cancer cell lines, as determined by microarray data plotted vs. DUSP4 mRNA expression. Points and text in black represent the correlation for all cell lines, and points and text in red represent the correlation of only the cell lines with a high MEK activation score (>median of all cell lines). G) Immunoblot analysis of a panel of breast cancer cell lines. TNBC: triple negative breast cancer; L: luminal, A: Basal A; B: Basal B.
    Dusp4, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/dusp4/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    dusp4 - by Bioz Stars, 2023-01
    94/100 stars

    Images

    1) Product Images from "Activation of MAPK pathways due to DUSP4 loss promotes cancer stem cell-like phenotypes in basal-like breast cancer"

    Article Title: Activation of MAPK pathways due to DUSP4 loss promotes cancer stem cell-like phenotypes in basal-like breast cancer

    Journal: Cancer research

    doi: 10.1158/0008-5472.CAN-13-1385

    A) Cancer Cell Line Encyclopedia data SNP copy number analysis for the DUSP4 locus across 964 cancer cell lines from various tissue sources (49). Red bars show the median log2 copy number for each tissue origin. B) Left panel; frequency histogram of TCGA (14) SNP copy number analysis for the DUSP4 locus for 443 breast cancers normalized to patient matched control tissue (log2 ratio). Right panel; DUSP4 log2 copy number ratio plotted by molecular subtype as determined by gene expression using the PAM50 centroids. P-value represents the ANOVA result. C) CD44:CD24 mRNA ratio for the ICBP50 panel of breast cancer cell lines, as determined by microarray data, plotted vs. the MEK activation score of Pratilas et al. (20). D) Unsupervised clustering of RNA from 15 mammosphere cultures vs. 11 primary tumor profiles, representing 16 patients including 10 patient pairs (7) using the genes defined in the MEK signature (20). E) Box and whisker plot of the data in (D), according to the total sum score of the MEK signature. P-value represents the result of a two-tailed t-test. F) CD44:CD24 mRNA ratio for the ICBP50 panel of breast cancer cell lines, as determined by microarray data plotted vs. DUSP4 mRNA expression. Points and text in black represent the correlation for all cell lines, and points and text in red represent the correlation of only the cell lines with a high MEK activation score (>median of all cell lines). G) Immunoblot analysis of a panel of breast cancer cell lines. TNBC: triple negative breast cancer; L: luminal, A: Basal A; B: Basal B.
    Figure Legend Snippet: A) Cancer Cell Line Encyclopedia data SNP copy number analysis for the DUSP4 locus across 964 cancer cell lines from various tissue sources (49). Red bars show the median log2 copy number for each tissue origin. B) Left panel; frequency histogram of TCGA (14) SNP copy number analysis for the DUSP4 locus for 443 breast cancers normalized to patient matched control tissue (log2 ratio). Right panel; DUSP4 log2 copy number ratio plotted by molecular subtype as determined by gene expression using the PAM50 centroids. P-value represents the ANOVA result. C) CD44:CD24 mRNA ratio for the ICBP50 panel of breast cancer cell lines, as determined by microarray data, plotted vs. the MEK activation score of Pratilas et al. (20). D) Unsupervised clustering of RNA from 15 mammosphere cultures vs. 11 primary tumor profiles, representing 16 patients including 10 patient pairs (7) using the genes defined in the MEK signature (20). E) Box and whisker plot of the data in (D), according to the total sum score of the MEK signature. P-value represents the result of a two-tailed t-test. F) CD44:CD24 mRNA ratio for the ICBP50 panel of breast cancer cell lines, as determined by microarray data plotted vs. DUSP4 mRNA expression. Points and text in black represent the correlation for all cell lines, and points and text in red represent the correlation of only the cell lines with a high MEK activation score (>median of all cell lines). G) Immunoblot analysis of a panel of breast cancer cell lines. TNBC: triple negative breast cancer; L: luminal, A: Basal A; B: Basal B.

    Techniques Used: Expressing, Microarray, Activation Assay, Whisker Assay, Two Tailed Test, Western Blot

    A) Immunoblot of MDA-231 cells 96 hrs after siCONTROL or siDUSP4 transfection in 10% FBS containing medium or after 24 hr of serum starvation in 0.1% FBS containing medium B) Mammosphere growth assay in MDA-231 cells, quantitated by GelCount software 6 days after siRNA transfection (5 days after plating to mammosphere culture conditions; ***p<0.001 for a two-tailed t-test). C) Immunoblot analysis performed on lysates from MDA-231 cells grown under non-adherent (mammosphere) conditions. D) Serum-free media was collected 72-96 hrs after siRNA transfection of MDA-231 cells and normalized to cell number. Conditioned medium was added to SUM159PT cells cultured in a mammosphere assay for 5 days and quantitated by GelCount software (*p=<0.05, two-tailed t-test E) qRT-PCR analysis of DUSP4 and IL6 mRNA expression in MDA-231 cells 72 hr after siRNA transfection. F) IL6 ELISA in serum-free medium conditioned by siRNA-transfected MDA-231 cells G) qRT-PCR analysis of IL6 mRNA expression in MDA-231 cells 96 hr after siRNA transfection and 4 hr after treatment with 1 μM selumetinib (MEKi) or 10 μM SP600125 (JNKi). H) Immunoblot analysis of MDA-231 cells after treatment for 24 hrs with 1 μM selumetinib (MEKi) or 10 μM SP600125 (JNKi). I. MDA-231 mammosphere formation quantitated by GelCount software 7 days after siRNA transfection. Where indicated, selumetinib (MEKi) or SP600125 (JNK1) or the combination was added to the mammosphere cultures. All bars represent the mean of 3 replicates ± SD.
    Figure Legend Snippet: A) Immunoblot of MDA-231 cells 96 hrs after siCONTROL or siDUSP4 transfection in 10% FBS containing medium or after 24 hr of serum starvation in 0.1% FBS containing medium B) Mammosphere growth assay in MDA-231 cells, quantitated by GelCount software 6 days after siRNA transfection (5 days after plating to mammosphere culture conditions; ***p<0.001 for a two-tailed t-test). C) Immunoblot analysis performed on lysates from MDA-231 cells grown under non-adherent (mammosphere) conditions. D) Serum-free media was collected 72-96 hrs after siRNA transfection of MDA-231 cells and normalized to cell number. Conditioned medium was added to SUM159PT cells cultured in a mammosphere assay for 5 days and quantitated by GelCount software (*p=<0.05, two-tailed t-test E) qRT-PCR analysis of DUSP4 and IL6 mRNA expression in MDA-231 cells 72 hr after siRNA transfection. F) IL6 ELISA in serum-free medium conditioned by siRNA-transfected MDA-231 cells G) qRT-PCR analysis of IL6 mRNA expression in MDA-231 cells 96 hr after siRNA transfection and 4 hr after treatment with 1 μM selumetinib (MEKi) or 10 μM SP600125 (JNKi). H) Immunoblot analysis of MDA-231 cells after treatment for 24 hrs with 1 μM selumetinib (MEKi) or 10 μM SP600125 (JNKi). I. MDA-231 mammosphere formation quantitated by GelCount software 7 days after siRNA transfection. Where indicated, selumetinib (MEKi) or SP600125 (JNK1) or the combination was added to the mammosphere cultures. All bars represent the mean of 3 replicates ± SD.

    Techniques Used: Western Blot, Transfection, Growth Assay, Software, Two Tailed Test, Cell Culture, Quantitative RT-PCR, Expressing, Enzyme-linked Immunosorbent Assay

    A) Correlation of IL6 mRNA and IL8 mRNA with DUSP4 mRNA in 2 external microarray datasets of primary breast cancer. B) qRT-PCR analysis of BT549 and SUM159PT cells treated for 16 hr with 10 μM SP600125 (JNKi), 1 μM selumetinib (MEKi) or the combination. C) IL6 and IL8 ELISA analyses of conditioned media from BT549 and SUM159PT cells collected after 24-48 hr of treatment with the indicated inhibitors. D) Immunoblot analysis of lysates from cells shown in (B) and (C). E) Immunoblot analysis of BT549 and SUM159PT cells after 16 hr of adenovirus transduction with AdDUSP4 or AdGFP control. F) qRT-PCR analysis of cells from (E). G) Immunoblot analysis of SUM159PT harvested 96 hr after transfection with siCONTROL, siETS-1, siCJUN or the combination. H) qRT-PCR analysis of IL6 and IL8 mRNAs in cells from (G).
    Figure Legend Snippet: A) Correlation of IL6 mRNA and IL8 mRNA with DUSP4 mRNA in 2 external microarray datasets of primary breast cancer. B) qRT-PCR analysis of BT549 and SUM159PT cells treated for 16 hr with 10 μM SP600125 (JNKi), 1 μM selumetinib (MEKi) or the combination. C) IL6 and IL8 ELISA analyses of conditioned media from BT549 and SUM159PT cells collected after 24-48 hr of treatment with the indicated inhibitors. D) Immunoblot analysis of lysates from cells shown in (B) and (C). E) Immunoblot analysis of BT549 and SUM159PT cells after 16 hr of adenovirus transduction with AdDUSP4 or AdGFP control. F) qRT-PCR analysis of cells from (E). G) Immunoblot analysis of SUM159PT harvested 96 hr after transfection with siCONTROL, siETS-1, siCJUN or the combination. H) qRT-PCR analysis of IL6 and IL8 mRNAs in cells from (G).

    Techniques Used: Microarray, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Western Blot, Transduction, Transfection

    Microarray analysis was performed on RNA derived from MDA-231, BT549 and SUM159PT cells 96 hr after transfection with siCONTROL or siDUSP4 and after 4 hr or 24 hr of selumetinib treatment. A) Heatmap of significantly altered genes for MDA-231 cells (ANOVA FDR-adjusted p<0.01). B) DUSP4 loss gene expression score derived from MDA-231 cells transfected with siDUSP4 was calculated across all samples. C) Expression of selected chemotherapy-resistance genes which were altered in MDA-231 cells transfected with siDUSP4. D) DUSP4-loss gene expression score derived from MDA-231 cells transfected with siDUSP4 was calculated across 444 tumors in the TCGA breast data. Molecular subtype was determined using the PAM50 centroids (50) and the genefu package in R. E) Association of the claudin-low nine-cell line predictive genes with MDA-231 cells following siDUSP4 or selumetinib treatment (5).
    Figure Legend Snippet: Microarray analysis was performed on RNA derived from MDA-231, BT549 and SUM159PT cells 96 hr after transfection with siCONTROL or siDUSP4 and after 4 hr or 24 hr of selumetinib treatment. A) Heatmap of significantly altered genes for MDA-231 cells (ANOVA FDR-adjusted p<0.01). B) DUSP4 loss gene expression score derived from MDA-231 cells transfected with siDUSP4 was calculated across all samples. C) Expression of selected chemotherapy-resistance genes which were altered in MDA-231 cells transfected with siDUSP4. D) DUSP4-loss gene expression score derived from MDA-231 cells transfected with siDUSP4 was calculated across 444 tumors in the TCGA breast data. Molecular subtype was determined using the PAM50 centroids (50) and the genefu package in R. E) Association of the claudin-low nine-cell line predictive genes with MDA-231 cells following siDUSP4 or selumetinib treatment (5).

    Techniques Used: Microarray, Derivative Assay, Transfection, Expressing

    SUM159PT cells transduced with DOX-inducible DUSP4-HA were generated. A) Flow cytometry analysis of CD44/CD24 expression in cells treated for 1-10 days with DOX (2 ng/mL). B) SUM159PT/pINDUCER-LACZ cells or SUM159PT/pINDUCER-DUSP4 pre-treated for 4 days ± DOX prior to plating in a mammosphere assay in the presence or absence of DOX as in the pretreatment. C) Immunoblot of parental SUM159PT/pINDUCER-LACZ cells or SUM159PT/pINDUCER-DUSP4 treated for 4 days with DOX. D) Tumor formation of cells from (C) 60 days after injection the mammary fatpad of athymic mice. Red bars indicate palpable tumors and blue bars indicate tumors identified on histological examination of the injection site. E) Schematic of proposed model based on the presented data: The tumor suppressor DUSP4 negatively regulates ERK and JNK. Upon loss of DUSP4, derepressed ERK and JNK activity stimulates ETS1 and cJUN-mediated transcription of IL6 and IL8, cytokines that expand the cancer stem-like cell population. De-repressed ERK transcription following DUSP4 loss also suppresses CD24 expression thus increasing the CD44HI CD24LO compartment, a marker of the CSC population.
    Figure Legend Snippet: SUM159PT cells transduced with DOX-inducible DUSP4-HA were generated. A) Flow cytometry analysis of CD44/CD24 expression in cells treated for 1-10 days with DOX (2 ng/mL). B) SUM159PT/pINDUCER-LACZ cells or SUM159PT/pINDUCER-DUSP4 pre-treated for 4 days ± DOX prior to plating in a mammosphere assay in the presence or absence of DOX as in the pretreatment. C) Immunoblot of parental SUM159PT/pINDUCER-LACZ cells or SUM159PT/pINDUCER-DUSP4 treated for 4 days with DOX. D) Tumor formation of cells from (C) 60 days after injection the mammary fatpad of athymic mice. Red bars indicate palpable tumors and blue bars indicate tumors identified on histological examination of the injection site. E) Schematic of proposed model based on the presented data: The tumor suppressor DUSP4 negatively regulates ERK and JNK. Upon loss of DUSP4, derepressed ERK and JNK activity stimulates ETS1 and cJUN-mediated transcription of IL6 and IL8, cytokines that expand the cancer stem-like cell population. De-repressed ERK transcription following DUSP4 loss also suppresses CD24 expression thus increasing the CD44HI CD24LO compartment, a marker of the CSC population.

    Techniques Used: Transduction, Generated, Flow Cytometry, Expressing, Western Blot, Injection, Activity Assay, Marker

    Similar Products

  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94
    Cell Signaling Technology Inc zfx
    Zfx, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/zfx/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    zfx - by Bioz Stars, 2023-01
    94/100 stars
      Buy from Supplier

    94
    Cell Signaling Technology Inc antibodies against zfx
    Antibodies Against Zfx, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/antibodies against zfx/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    antibodies against zfx - by Bioz Stars, 2023-01
    94/100 stars
      Buy from Supplier

    94
    Cell Signaling Technology Inc zfx antibody
    The <t>ZFX</t> gene family. Shown are gene structure schematics for ZFX, ZFY <t>and</t> <t>ZNF711.</t> Dashed lines indicate zinc fingers conserved between ZFX and the other two family members. NLS: nuclear localization sequence.
    Zfx Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/zfx antibody/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    zfx antibody - by Bioz Stars, 2023-01
    94/100 stars
      Buy from Supplier

    94
    Cell Signaling Technology Inc anti tubulin dm1a mouse mab cell signaling tech
    KEY RESOURCES TABLE
    Anti Tubulin Dm1a Mouse Mab Cell Signaling Tech, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti tubulin dm1a mouse mab cell signaling tech/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    anti tubulin dm1a mouse mab cell signaling tech - by Bioz Stars, 2023-01
    94/100 stars
      Buy from Supplier

    94
    Cell Signaling Technology Inc mouse anti zfx
    KEY RESOURCES TABLE
    Mouse Anti Zfx, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mouse anti zfx/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    mouse anti zfx - by Bioz Stars, 2023-01
    94/100 stars
      Buy from Supplier

    94
    Cell Signaling Technology Inc 2008 na mouse anti zfx cell signaling mab
    KEY RESOURCES TABLE
    2008 Na Mouse Anti Zfx Cell Signaling Mab, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/2008 na mouse anti zfx cell signaling mab/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    2008 na mouse anti zfx cell signaling mab - by Bioz Stars, 2023-01
    94/100 stars
      Buy from Supplier

    94
    Cell Signaling Technology Inc dusp4
    A) Cancer Cell Line Encyclopedia data SNP copy number analysis for the <t>DUSP4</t> locus across 964 cancer cell lines from various tissue sources (49). Red bars show the median log2 copy number for each tissue origin. B) Left panel; frequency histogram of TCGA (14) SNP copy number analysis for the DUSP4 locus for 443 breast cancers normalized to patient matched control tissue (log2 ratio). Right panel; DUSP4 log2 copy number ratio plotted by molecular subtype as determined by gene expression using the PAM50 centroids. P-value represents the ANOVA result. C) CD44:CD24 mRNA ratio for the ICBP50 panel of breast cancer cell lines, as determined by microarray data, plotted vs. the MEK activation score of Pratilas et al. (20). D) Unsupervised clustering of RNA from 15 mammosphere cultures vs. 11 primary tumor profiles, representing 16 patients including 10 patient pairs (7) using the genes defined in the MEK signature (20). E) Box and whisker plot of the data in (D), according to the total sum score of the MEK signature. P-value represents the result of a two-tailed t-test. F) CD44:CD24 mRNA ratio for the ICBP50 panel of breast cancer cell lines, as determined by microarray data plotted vs. DUSP4 mRNA expression. Points and text in black represent the correlation for all cell lines, and points and text in red represent the correlation of only the cell lines with a high MEK activation score (>median of all cell lines). G) Immunoblot analysis of a panel of breast cancer cell lines. TNBC: triple negative breast cancer; L: luminal, A: Basal A; B: Basal B.
    Dusp4, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/dusp4/product/Cell Signaling Technology Inc
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    dusp4 - by Bioz Stars, 2023-01
    94/100 stars
      Buy from Supplier

    Image Search Results


    The ZFX gene family. Shown are gene structure schematics for ZFX, ZFY and ZNF711. Dashed lines indicate zinc fingers conserved between ZFX and the other two family members. NLS: nuclear localization sequence.

    Journal: Nucleic Acids Research

    Article Title: Characterization of the ZFX family of transcription factors that bind downstream of the start site of CpG island promoters

    doi: 10.1093/nar/gkaa384

    Figure Lengend Snippet: The ZFX gene family. Shown are gene structure schematics for ZFX, ZFY and ZNF711. Dashed lines indicate zinc fingers conserved between ZFX and the other two family members. NLS: nuclear localization sequence.

    Article Snippet: Crosslinked cells, ZFX antibody (Cell Signaling Technology # 5419S), and ZNF711 antibody (Thermo Fisher #PA5-31815) were sent to Peconic, where the ChIP-exo assay was performed ( http://www.peconicgenomics.com/services.html ).

    Techniques: Zinc-Fingers, Sequencing

    Loss of ZFX and ZNF711 in HEK293T cells inhibits cell proliferation. ( A ) Expression levels of ZFX/ZFY/ZNF711 in wt HEK293T cells. ( B ) Locations of gRNAs used to create CRISPR/Cas9-mediated ZFX and/or ZNF711 knockouts. The deletion of ZFX in ZFX KO clone1 and clone2 and the DKO clones were generated using ZFX gRNA1 and gRNA2. The deletion of ZNF711 in ZNF711 KO clone1 and the DKO clones was generated using ZNF711 gRNA1 and gRNA2; the deletion of ZNF711 KO clone2 was generated using ZNF711 gRNA2 and gRNA3. ( C ) Western blots showing the protein levels of ZFX and ZNF711 in wt HEK293T, ZFX KO clones, ZNF711 KO clones, and DKO clones; also shown is the level of p62 as a loading control. ( D ) Proliferation assays using wt HEK293T, two different ZFX and two different ZNF711 KO clones, and two DKO clones; data points are the mean of three biological replicates.

    Journal: Nucleic Acids Research

    Article Title: Characterization of the ZFX family of transcription factors that bind downstream of the start site of CpG island promoters

    doi: 10.1093/nar/gkaa384

    Figure Lengend Snippet: Loss of ZFX and ZNF711 in HEK293T cells inhibits cell proliferation. ( A ) Expression levels of ZFX/ZFY/ZNF711 in wt HEK293T cells. ( B ) Locations of gRNAs used to create CRISPR/Cas9-mediated ZFX and/or ZNF711 knockouts. The deletion of ZFX in ZFX KO clone1 and clone2 and the DKO clones were generated using ZFX gRNA1 and gRNA2. The deletion of ZNF711 in ZNF711 KO clone1 and the DKO clones was generated using ZNF711 gRNA1 and gRNA2; the deletion of ZNF711 KO clone2 was generated using ZNF711 gRNA2 and gRNA3. ( C ) Western blots showing the protein levels of ZFX and ZNF711 in wt HEK293T, ZFX KO clones, ZNF711 KO clones, and DKO clones; also shown is the level of p62 as a loading control. ( D ) Proliferation assays using wt HEK293T, two different ZFX and two different ZNF711 KO clones, and two DKO clones; data points are the mean of three biological replicates.

    Article Snippet: Crosslinked cells, ZFX antibody (Cell Signaling Technology # 5419S), and ZNF711 antibody (Thermo Fisher #PA5-31815) were sent to Peconic, where the ChIP-exo assay was performed ( http://www.peconicgenomics.com/services.html ).

    Techniques: Expressing, CRISPR, Clone Assay, Generated, Western Blot

    Reduction in ZFX and ZNF711 levels causes large effects on the transcriptome. ( A ) Volcano plots showing the differentially expressed genes (DEGs) identified via RNA-seq in comparisons of wt HEK293T versus ZFX KO clone1, KO clone2, ZNF711 KO clone1, KO clone2, DKO clone1, DKO clone2, or DKO clone3. ( B ) Comparison of DEGs commonly downregulated or upregulated in all three DKO clones. ( C ) Gene ontology analysis of the 1166 commonly downregulated and 2124 commonly upregulated genes in all three DKO clones.

    Journal: Nucleic Acids Research

    Article Title: Characterization of the ZFX family of transcription factors that bind downstream of the start site of CpG island promoters

    doi: 10.1093/nar/gkaa384

    Figure Lengend Snippet: Reduction in ZFX and ZNF711 levels causes large effects on the transcriptome. ( A ) Volcano plots showing the differentially expressed genes (DEGs) identified via RNA-seq in comparisons of wt HEK293T versus ZFX KO clone1, KO clone2, ZNF711 KO clone1, KO clone2, DKO clone1, DKO clone2, or DKO clone3. ( B ) Comparison of DEGs commonly downregulated or upregulated in all three DKO clones. ( C ) Gene ontology analysis of the 1166 commonly downregulated and 2124 commonly upregulated genes in all three DKO clones.

    Article Snippet: Crosslinked cells, ZFX antibody (Cell Signaling Technology # 5419S), and ZNF711 antibody (Thermo Fisher #PA5-31815) were sent to Peconic, where the ChIP-exo assay was performed ( http://www.peconicgenomics.com/services.html ).

    Techniques: RNA Sequencing Assay, Clone Assay

    ZFX family members have essentially identical binding patterns at CpG island promoters. ( A ) Browser tracks showing ZFX family member binding profiles in female HEK293T kidney cells and male 22Rv1 prostate cells. Also shown is a zoom in on a single peak located in the DOCK7 promoter region. ( B ) Shown is a heatmap illustrating the genome-wide correlation of ZFX family member binding patterns in 22Rv1 cells. ( C ) Bar graph of genomic distributions of ZFX family member binding sites in 22Rv1 cells in promoter and non-promoter regions (left) and bar graph showing the relative distribution of binding sites in CpG island (CGI) promoters and non-CpG island promoters (right). ( D ) Venn diagram comparing the sets of CpG island promoters bound by ZFX, ZFY and ZNF711 in 22Rv1 cells.

    Journal: Nucleic Acids Research

    Article Title: Characterization of the ZFX family of transcription factors that bind downstream of the start site of CpG island promoters

    doi: 10.1093/nar/gkaa384

    Figure Lengend Snippet: ZFX family members have essentially identical binding patterns at CpG island promoters. ( A ) Browser tracks showing ZFX family member binding profiles in female HEK293T kidney cells and male 22Rv1 prostate cells. Also shown is a zoom in on a single peak located in the DOCK7 promoter region. ( B ) Shown is a heatmap illustrating the genome-wide correlation of ZFX family member binding patterns in 22Rv1 cells. ( C ) Bar graph of genomic distributions of ZFX family member binding sites in 22Rv1 cells in promoter and non-promoter regions (left) and bar graph showing the relative distribution of binding sites in CpG island (CGI) promoters and non-CpG island promoters (right). ( D ) Venn diagram comparing the sets of CpG island promoters bound by ZFX, ZFY and ZNF711 in 22Rv1 cells.

    Article Snippet: Crosslinked cells, ZFX antibody (Cell Signaling Technology # 5419S), and ZNF711 antibody (Thermo Fisher #PA5-31815) were sent to Peconic, where the ChIP-exo assay was performed ( http://www.peconicgenomics.com/services.html ).

    Techniques: Binding Assay, Genome Wide

    ZFX and ZNF711 have properties of a transcription activator when bound downstream of the TSS. ( A ) ZFX and ZNF711 peak sets from HEK293T cells were clustered using K-means clustering, identifying four sets of peaks with distinct binding sites (left); cluster 4 (combination peaks) was subsequently re-clustered, identifying 4 subsets (right). Tag density plots for each of the 4 different clusters are presented on top of the heatmaps. ( B ) Average signals of ZFX and ZNF711 ChIP-seq reads in wt HEK293T at all promoters bound by each TF (top), promoters of genes with decreased expression in all three DKO clones (middle), and promoters of genes with increased expressions in all three DKO clones (bottom). Also shown, for both the downregulated and the upregulated gene categories, is the percentage of genes whose promoters are bound by ZFX or ZNF711 in peak categories 1–4, or not bound by ZFX or ZNF711.

    Journal: Nucleic Acids Research

    Article Title: Characterization of the ZFX family of transcription factors that bind downstream of the start site of CpG island promoters

    doi: 10.1093/nar/gkaa384

    Figure Lengend Snippet: ZFX and ZNF711 have properties of a transcription activator when bound downstream of the TSS. ( A ) ZFX and ZNF711 peak sets from HEK293T cells were clustered using K-means clustering, identifying four sets of peaks with distinct binding sites (left); cluster 4 (combination peaks) was subsequently re-clustered, identifying 4 subsets (right). Tag density plots for each of the 4 different clusters are presented on top of the heatmaps. ( B ) Average signals of ZFX and ZNF711 ChIP-seq reads in wt HEK293T at all promoters bound by each TF (top), promoters of genes with decreased expression in all three DKO clones (middle), and promoters of genes with increased expressions in all three DKO clones (bottom). Also shown, for both the downregulated and the upregulated gene categories, is the percentage of genes whose promoters are bound by ZFX or ZNF711 in peak categories 1–4, or not bound by ZFX or ZNF711.

    Article Snippet: Crosslinked cells, ZFX antibody (Cell Signaling Technology # 5419S), and ZNF711 antibody (Thermo Fisher #PA5-31815) were sent to Peconic, where the ChIP-exo assay was performed ( http://www.peconicgenomics.com/services.html ).

    Techniques: Binding Assay, ChIP-sequencing, Expressing, Clone Assay

    Characterization of ZFX and ZNF711 binding sites. ( A ) Classification of binding sites based on genomic locations of all ZFX peaks located downstream of the TSS and ZFX downstream peaks at promoters of genes down-regulated in all 3 DKO clones. ( B ) Classification of ZNF711 downstream ChIP-seq peaks, downstream peaks identified using read2 of the ChIP-exo dataset, and downstream peaks identified by the ChexMix program (+/-10 nt from the nt identified as the binding site by the program). ( C ) Tag density plots of all ZNF711 peaks from standard ChIP-seq and from ChIP-exo. ( D ) Motif analysis using the top 5000 peaks identified from standard ZNF711 ChIP-seq (average width 1800 nt), ChIP-exo read2 (average width 300 nt), ChIP-exo by the ChexMix program (20 nt), and randomized CpG island promoter regions (width 2 kb, 200 bp, and 20 nt). The peaks were searched for the known ZNF711 motif and the 5 nt motif identified by ChIP-exo. ( E) Zoom-in comparison of peaks from ZNF711 standard ChIP-seq replicates and ChIP-exo replicates.

    Journal: Nucleic Acids Research

    Article Title: Characterization of the ZFX family of transcription factors that bind downstream of the start site of CpG island promoters

    doi: 10.1093/nar/gkaa384

    Figure Lengend Snippet: Characterization of ZFX and ZNF711 binding sites. ( A ) Classification of binding sites based on genomic locations of all ZFX peaks located downstream of the TSS and ZFX downstream peaks at promoters of genes down-regulated in all 3 DKO clones. ( B ) Classification of ZNF711 downstream ChIP-seq peaks, downstream peaks identified using read2 of the ChIP-exo dataset, and downstream peaks identified by the ChexMix program (+/-10 nt from the nt identified as the binding site by the program). ( C ) Tag density plots of all ZNF711 peaks from standard ChIP-seq and from ChIP-exo. ( D ) Motif analysis using the top 5000 peaks identified from standard ZNF711 ChIP-seq (average width 1800 nt), ChIP-exo read2 (average width 300 nt), ChIP-exo by the ChexMix program (20 nt), and randomized CpG island promoter regions (width 2 kb, 200 bp, and 20 nt). The peaks were searched for the known ZNF711 motif and the 5 nt motif identified by ChIP-exo. ( E) Zoom-in comparison of peaks from ZNF711 standard ChIP-seq replicates and ChIP-exo replicates.

    Article Snippet: Crosslinked cells, ZFX antibody (Cell Signaling Technology # 5419S), and ZNF711 antibody (Thermo Fisher #PA5-31815) were sent to Peconic, where the ChIP-exo assay was performed ( http://www.peconicgenomics.com/services.html ).

    Techniques: Binding Assay, Clone Assay, ChIP-sequencing

    Functional analysis of the ZFX protein. ( A ) Schematic of FLAG-tagged ZFX zinc finger (ZF) mutant constructs. ( B ) Browser tracks showing genomic binding profiles of endogenous ZFX and FLAG-tagged wt ZFX and ZFX mutants in HEK293T cells. ( C ) Tag density plots of ChIP-seq peaks comparing endogenous ZFX and ZFX ZF9-13 peak locations in HEK293T cells. ( D ) Heatmaps showing ChIP-seq data from FLAG-tagged wt ZFX and ZFX ZF9-13 centered on the genomic locations of the endogenous ZFX peaks. ( E ) Expression levels following transfection with different ZFX constructs (as analyzed by RT-qPCR) of two genes (LONRF2 and CAPN2) whose promoters are bound by both ZFX and ZNF711 in wt HEK293T cells and which show a reduction in gene expression in all three DKO clones, of one gene (FOS) that is upregulated in all three DKO cells (a putative indirect target gene), and of one gene (HOXC4) that shows no expression changes in the DKO cells. Expression data were normalized to the control (cells transfected with an unrelated plasmid). Three independent experiments were performed using two different clonal populations of DKO cells; data points represent results from triplicate wells and duplicate PCR readings. Error bars indicate the pooled standard deviations of the means for the constructs and for the normalizing control.

    Journal: Nucleic Acids Research

    Article Title: Characterization of the ZFX family of transcription factors that bind downstream of the start site of CpG island promoters

    doi: 10.1093/nar/gkaa384

    Figure Lengend Snippet: Functional analysis of the ZFX protein. ( A ) Schematic of FLAG-tagged ZFX zinc finger (ZF) mutant constructs. ( B ) Browser tracks showing genomic binding profiles of endogenous ZFX and FLAG-tagged wt ZFX and ZFX mutants in HEK293T cells. ( C ) Tag density plots of ChIP-seq peaks comparing endogenous ZFX and ZFX ZF9-13 peak locations in HEK293T cells. ( D ) Heatmaps showing ChIP-seq data from FLAG-tagged wt ZFX and ZFX ZF9-13 centered on the genomic locations of the endogenous ZFX peaks. ( E ) Expression levels following transfection with different ZFX constructs (as analyzed by RT-qPCR) of two genes (LONRF2 and CAPN2) whose promoters are bound by both ZFX and ZNF711 in wt HEK293T cells and which show a reduction in gene expression in all three DKO clones, of one gene (FOS) that is upregulated in all three DKO cells (a putative indirect target gene), and of one gene (HOXC4) that shows no expression changes in the DKO cells. Expression data were normalized to the control (cells transfected with an unrelated plasmid). Three independent experiments were performed using two different clonal populations of DKO cells; data points represent results from triplicate wells and duplicate PCR readings. Error bars indicate the pooled standard deviations of the means for the constructs and for the normalizing control.

    Article Snippet: Crosslinked cells, ZFX antibody (Cell Signaling Technology # 5419S), and ZNF711 antibody (Thermo Fisher #PA5-31815) were sent to Peconic, where the ChIP-exo assay was performed ( http://www.peconicgenomics.com/services.html ).

    Techniques: Functional Assay, Mutagenesis, Construct, Binding Assay, ChIP-sequencing, Expressing, Transfection, Quantitative RT-PCR, Clone Assay, Plasmid Preparation

    ZFX ZF11-13 has very similar transcriptional activities as wt ZFX. ( A ) Volcano plots showing the differentially expressed genes (DEGs) identified via RNA-seq in comparisons of DKO cells 24 hr after transfection with FLAG-tagged wt ZFX vs. a control plasmid, FLAG-tagged ZFX ZF11-13 vs. a control plasmid, and FLAG-tagged wt ZFX vs. FLAG-tagged ZFX ZF11-13. ( B ) Shown is a Venn diagram comparing the 846 genes that are bound by ZFX and ZNF711 in wt HEK293T cells and show a decrease in mRNA levels in all three DKO clones and the 2275 genes that show increased levels in DKO cells transfected with either FLAG-tagged wt ZFX or ZFX ZF11-13. ( C ) Shown is a tag density plot of ChIP-seq data for endogenous ZFX, FLAG-tagged wt ZFX, ZFX ZF9-13, or ZFX ZF1-8 at the set of 277 responding promoters identified in panel B. ( D ) Motif coverage analysis of the 277 responding promoters.

    Journal: Nucleic Acids Research

    Article Title: Characterization of the ZFX family of transcription factors that bind downstream of the start site of CpG island promoters

    doi: 10.1093/nar/gkaa384

    Figure Lengend Snippet: ZFX ZF11-13 has very similar transcriptional activities as wt ZFX. ( A ) Volcano plots showing the differentially expressed genes (DEGs) identified via RNA-seq in comparisons of DKO cells 24 hr after transfection with FLAG-tagged wt ZFX vs. a control plasmid, FLAG-tagged ZFX ZF11-13 vs. a control plasmid, and FLAG-tagged wt ZFX vs. FLAG-tagged ZFX ZF11-13. ( B ) Shown is a Venn diagram comparing the 846 genes that are bound by ZFX and ZNF711 in wt HEK293T cells and show a decrease in mRNA levels in all three DKO clones and the 2275 genes that show increased levels in DKO cells transfected with either FLAG-tagged wt ZFX or ZFX ZF11-13. ( C ) Shown is a tag density plot of ChIP-seq data for endogenous ZFX, FLAG-tagged wt ZFX, ZFX ZF9-13, or ZFX ZF1-8 at the set of 277 responding promoters identified in panel B. ( D ) Motif coverage analysis of the 277 responding promoters.

    Article Snippet: Crosslinked cells, ZFX antibody (Cell Signaling Technology # 5419S), and ZNF711 antibody (Thermo Fisher #PA5-31815) were sent to Peconic, where the ChIP-exo assay was performed ( http://www.peconicgenomics.com/services.html ).

    Techniques: RNA Sequencing Assay, Transfection, Plasmid Preparation, Clone Assay, ChIP-sequencing

    KEY RESOURCES TABLE

    Journal: Molecular cell

    Article Title: ZFX mediates non-canonical oncogenic functions of the androgen receptor splice variant 7 (AR-V7) in castrate-resistant prostate cancer

    doi: 10.1016/j.molcel.2018.08.029

    Figure Lengend Snippet: KEY RESOURCES TABLE

    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies anti-full-length AR (AR-FL) C-terminus (C19) Santa Cruz Biotechnology catalog # sc-815X anti-AR-V7 C-terminus specific Precision Antibody AG10008 anti-pan-AR N-terminus (N20) Santa Cruz Biotechnology sc-816 anti-BRD4 ChIP Grade Bethyl A301–985A100 Mouse anti-Flag tag (M2) Sigma F1804 Mouse Anti-HA tag antibody - ChIP Grade Abcam ab9110 Anti-HA HA.11 (16B12) Covance MMS-101 P-200 anti-ZFX Thermo Fisher PA5–34376 anti-ZFX Chen et al., 2008 NA Mouse anti-ZFX Cell Signaling mAb #5419 anti-Tubulin (DM1A) Mouse mAb Cell signaling tech.

    Techniques: Immunoprecipitation, Recombinant, Electron Microscopy, Transfection, Protease Inhibitor, SYBR Green Assay, Purification, Multiplex Assay, Proliferation Assay, Mutagenesis, Magnetic Beads, Western Blot, Stripping, shRNA, Construct, Plasmid Preparation, Software

    KEY RESOURCES TABLE

    Journal: Molecular cell

    Article Title: ZFX mediates non-canonical oncogenic functions of the androgen receptor splice variant 7 (AR-V7) in castrate-resistant prostate cancer

    doi: 10.1016/j.molcel.2018.08.029

    Figure Lengend Snippet: KEY RESOURCES TABLE

    Article Snippet: Mouse anti-ZFX , Cell Signaling , , mAb #5419 , .

    Techniques: Immunoprecipitation, Recombinant, Electron Microscopy, Transfection, Protease Inhibitor, SYBR Green Assay, Purification, Multiplex Assay, Proliferation Assay, Mutagenesis, Magnetic Beads, Western Blot, Stripping, shRNA, Construct, Plasmid Preparation, Software

    KEY RESOURCES TABLE

    Journal: Molecular cell

    Article Title: ZFX mediates non-canonical oncogenic functions of the androgen receptor splice variant 7 (AR-V7) in castrate-resistant prostate cancer

    doi: 10.1016/j.molcel.2018.08.029

    Figure Lengend Snippet: KEY RESOURCES TABLE

    Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies anti-full-length AR (AR-FL) C-terminus (C19) Santa Cruz Biotechnology catalog # sc-815X anti-AR-V7 C-terminus specific Precision Antibody AG10008 anti-pan-AR N-terminus (N20) Santa Cruz Biotechnology sc-816 anti-BRD4 ChIP Grade Bethyl A301–985A100 Mouse anti-Flag tag (M2) Sigma F1804 Mouse Anti-HA tag antibody - ChIP Grade Abcam ab9110 Anti-HA HA.11 (16B12) Covance MMS-101 P-200 anti-ZFX Thermo Fisher PA5–34376 anti-ZFX Chen et al., 2008 NA Mouse anti-ZFX Cell Signaling mAb #5419 anti-Tubulin (DM1A) Mouse mAb Cell signaling tech.

    Techniques: Immunoprecipitation, Recombinant, Electron Microscopy, Transfection, Protease Inhibitor, SYBR Green Assay, Purification, Multiplex Assay, Proliferation Assay, Mutagenesis, Magnetic Beads, Western Blot, Stripping, shRNA, Construct, Plasmid Preparation, Software

    A) Cancer Cell Line Encyclopedia data SNP copy number analysis for the DUSP4 locus across 964 cancer cell lines from various tissue sources (49). Red bars show the median log2 copy number for each tissue origin. B) Left panel; frequency histogram of TCGA (14) SNP copy number analysis for the DUSP4 locus for 443 breast cancers normalized to patient matched control tissue (log2 ratio). Right panel; DUSP4 log2 copy number ratio plotted by molecular subtype as determined by gene expression using the PAM50 centroids. P-value represents the ANOVA result. C) CD44:CD24 mRNA ratio for the ICBP50 panel of breast cancer cell lines, as determined by microarray data, plotted vs. the MEK activation score of Pratilas et al. (20). D) Unsupervised clustering of RNA from 15 mammosphere cultures vs. 11 primary tumor profiles, representing 16 patients including 10 patient pairs (7) using the genes defined in the MEK signature (20). E) Box and whisker plot of the data in (D), according to the total sum score of the MEK signature. P-value represents the result of a two-tailed t-test. F) CD44:CD24 mRNA ratio for the ICBP50 panel of breast cancer cell lines, as determined by microarray data plotted vs. DUSP4 mRNA expression. Points and text in black represent the correlation for all cell lines, and points and text in red represent the correlation of only the cell lines with a high MEK activation score (>median of all cell lines). G) Immunoblot analysis of a panel of breast cancer cell lines. TNBC: triple negative breast cancer; L: luminal, A: Basal A; B: Basal B.

    Journal: Cancer research

    Article Title: Activation of MAPK pathways due to DUSP4 loss promotes cancer stem cell-like phenotypes in basal-like breast cancer

    doi: 10.1158/0008-5472.CAN-13-1385

    Figure Lengend Snippet: A) Cancer Cell Line Encyclopedia data SNP copy number analysis for the DUSP4 locus across 964 cancer cell lines from various tissue sources (49). Red bars show the median log2 copy number for each tissue origin. B) Left panel; frequency histogram of TCGA (14) SNP copy number analysis for the DUSP4 locus for 443 breast cancers normalized to patient matched control tissue (log2 ratio). Right panel; DUSP4 log2 copy number ratio plotted by molecular subtype as determined by gene expression using the PAM50 centroids. P-value represents the ANOVA result. C) CD44:CD24 mRNA ratio for the ICBP50 panel of breast cancer cell lines, as determined by microarray data, plotted vs. the MEK activation score of Pratilas et al. (20). D) Unsupervised clustering of RNA from 15 mammosphere cultures vs. 11 primary tumor profiles, representing 16 patients including 10 patient pairs (7) using the genes defined in the MEK signature (20). E) Box and whisker plot of the data in (D), according to the total sum score of the MEK signature. P-value represents the result of a two-tailed t-test. F) CD44:CD24 mRNA ratio for the ICBP50 panel of breast cancer cell lines, as determined by microarray data plotted vs. DUSP4 mRNA expression. Points and text in black represent the correlation for all cell lines, and points and text in red represent the correlation of only the cell lines with a high MEK activation score (>median of all cell lines). G) Immunoblot analysis of a panel of breast cancer cell lines. TNBC: triple negative breast cancer; L: luminal, A: Basal A; B: Basal B.

    Article Snippet: Antibodies used for immunoblotting were: p-ERK1/2 (p-T202/Y204; #9101), calnexin (#2433), p-cJUN (#2361), cJUN (#9165), p-JNK1/2 (#4668), JNK1/2 (#9252) (all from Cell Signaling), p-ETS-1(p-T38, Invitrogen 44-1104G), ETS-1 (Santa Cruz sc-350), DUSP4 (Cell Signaling #5419), Anti-HA tag (Santa Cruz sc-805) and actin (Sigma, A2066).

    Techniques: Expressing, Microarray, Activation Assay, Whisker Assay, Two Tailed Test, Western Blot

    A) Immunoblot of MDA-231 cells 96 hrs after siCONTROL or siDUSP4 transfection in 10% FBS containing medium or after 24 hr of serum starvation in 0.1% FBS containing medium B) Mammosphere growth assay in MDA-231 cells, quantitated by GelCount software 6 days after siRNA transfection (5 days after plating to mammosphere culture conditions; ***p<0.001 for a two-tailed t-test). C) Immunoblot analysis performed on lysates from MDA-231 cells grown under non-adherent (mammosphere) conditions. D) Serum-free media was collected 72-96 hrs after siRNA transfection of MDA-231 cells and normalized to cell number. Conditioned medium was added to SUM159PT cells cultured in a mammosphere assay for 5 days and quantitated by GelCount software (*p=<0.05, two-tailed t-test E) qRT-PCR analysis of DUSP4 and IL6 mRNA expression in MDA-231 cells 72 hr after siRNA transfection. F) IL6 ELISA in serum-free medium conditioned by siRNA-transfected MDA-231 cells G) qRT-PCR analysis of IL6 mRNA expression in MDA-231 cells 96 hr after siRNA transfection and 4 hr after treatment with 1 μM selumetinib (MEKi) or 10 μM SP600125 (JNKi). H) Immunoblot analysis of MDA-231 cells after treatment for 24 hrs with 1 μM selumetinib (MEKi) or 10 μM SP600125 (JNKi). I. MDA-231 mammosphere formation quantitated by GelCount software 7 days after siRNA transfection. Where indicated, selumetinib (MEKi) or SP600125 (JNK1) or the combination was added to the mammosphere cultures. All bars represent the mean of 3 replicates ± SD.

    Journal: Cancer research

    Article Title: Activation of MAPK pathways due to DUSP4 loss promotes cancer stem cell-like phenotypes in basal-like breast cancer

    doi: 10.1158/0008-5472.CAN-13-1385

    Figure Lengend Snippet: A) Immunoblot of MDA-231 cells 96 hrs after siCONTROL or siDUSP4 transfection in 10% FBS containing medium or after 24 hr of serum starvation in 0.1% FBS containing medium B) Mammosphere growth assay in MDA-231 cells, quantitated by GelCount software 6 days after siRNA transfection (5 days after plating to mammosphere culture conditions; ***p<0.001 for a two-tailed t-test). C) Immunoblot analysis performed on lysates from MDA-231 cells grown under non-adherent (mammosphere) conditions. D) Serum-free media was collected 72-96 hrs after siRNA transfection of MDA-231 cells and normalized to cell number. Conditioned medium was added to SUM159PT cells cultured in a mammosphere assay for 5 days and quantitated by GelCount software (*p=<0.05, two-tailed t-test E) qRT-PCR analysis of DUSP4 and IL6 mRNA expression in MDA-231 cells 72 hr after siRNA transfection. F) IL6 ELISA in serum-free medium conditioned by siRNA-transfected MDA-231 cells G) qRT-PCR analysis of IL6 mRNA expression in MDA-231 cells 96 hr after siRNA transfection and 4 hr after treatment with 1 μM selumetinib (MEKi) or 10 μM SP600125 (JNKi). H) Immunoblot analysis of MDA-231 cells after treatment for 24 hrs with 1 μM selumetinib (MEKi) or 10 μM SP600125 (JNKi). I. MDA-231 mammosphere formation quantitated by GelCount software 7 days after siRNA transfection. Where indicated, selumetinib (MEKi) or SP600125 (JNK1) or the combination was added to the mammosphere cultures. All bars represent the mean of 3 replicates ± SD.

    Article Snippet: Antibodies used for immunoblotting were: p-ERK1/2 (p-T202/Y204; #9101), calnexin (#2433), p-cJUN (#2361), cJUN (#9165), p-JNK1/2 (#4668), JNK1/2 (#9252) (all from Cell Signaling), p-ETS-1(p-T38, Invitrogen 44-1104G), ETS-1 (Santa Cruz sc-350), DUSP4 (Cell Signaling #5419), Anti-HA tag (Santa Cruz sc-805) and actin (Sigma, A2066).

    Techniques: Western Blot, Transfection, Growth Assay, Software, Two Tailed Test, Cell Culture, Quantitative RT-PCR, Expressing, Enzyme-linked Immunosorbent Assay

    A) Correlation of IL6 mRNA and IL8 mRNA with DUSP4 mRNA in 2 external microarray datasets of primary breast cancer. B) qRT-PCR analysis of BT549 and SUM159PT cells treated for 16 hr with 10 μM SP600125 (JNKi), 1 μM selumetinib (MEKi) or the combination. C) IL6 and IL8 ELISA analyses of conditioned media from BT549 and SUM159PT cells collected after 24-48 hr of treatment with the indicated inhibitors. D) Immunoblot analysis of lysates from cells shown in (B) and (C). E) Immunoblot analysis of BT549 and SUM159PT cells after 16 hr of adenovirus transduction with AdDUSP4 or AdGFP control. F) qRT-PCR analysis of cells from (E). G) Immunoblot analysis of SUM159PT harvested 96 hr after transfection with siCONTROL, siETS-1, siCJUN or the combination. H) qRT-PCR analysis of IL6 and IL8 mRNAs in cells from (G).

    Journal: Cancer research

    Article Title: Activation of MAPK pathways due to DUSP4 loss promotes cancer stem cell-like phenotypes in basal-like breast cancer

    doi: 10.1158/0008-5472.CAN-13-1385

    Figure Lengend Snippet: A) Correlation of IL6 mRNA and IL8 mRNA with DUSP4 mRNA in 2 external microarray datasets of primary breast cancer. B) qRT-PCR analysis of BT549 and SUM159PT cells treated for 16 hr with 10 μM SP600125 (JNKi), 1 μM selumetinib (MEKi) or the combination. C) IL6 and IL8 ELISA analyses of conditioned media from BT549 and SUM159PT cells collected after 24-48 hr of treatment with the indicated inhibitors. D) Immunoblot analysis of lysates from cells shown in (B) and (C). E) Immunoblot analysis of BT549 and SUM159PT cells after 16 hr of adenovirus transduction with AdDUSP4 or AdGFP control. F) qRT-PCR analysis of cells from (E). G) Immunoblot analysis of SUM159PT harvested 96 hr after transfection with siCONTROL, siETS-1, siCJUN or the combination. H) qRT-PCR analysis of IL6 and IL8 mRNAs in cells from (G).

    Article Snippet: Antibodies used for immunoblotting were: p-ERK1/2 (p-T202/Y204; #9101), calnexin (#2433), p-cJUN (#2361), cJUN (#9165), p-JNK1/2 (#4668), JNK1/2 (#9252) (all from Cell Signaling), p-ETS-1(p-T38, Invitrogen 44-1104G), ETS-1 (Santa Cruz sc-350), DUSP4 (Cell Signaling #5419), Anti-HA tag (Santa Cruz sc-805) and actin (Sigma, A2066).

    Techniques: Microarray, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Western Blot, Transduction, Transfection

    Microarray analysis was performed on RNA derived from MDA-231, BT549 and SUM159PT cells 96 hr after transfection with siCONTROL or siDUSP4 and after 4 hr or 24 hr of selumetinib treatment. A) Heatmap of significantly altered genes for MDA-231 cells (ANOVA FDR-adjusted p<0.01). B) DUSP4 loss gene expression score derived from MDA-231 cells transfected with siDUSP4 was calculated across all samples. C) Expression of selected chemotherapy-resistance genes which were altered in MDA-231 cells transfected with siDUSP4. D) DUSP4-loss gene expression score derived from MDA-231 cells transfected with siDUSP4 was calculated across 444 tumors in the TCGA breast data. Molecular subtype was determined using the PAM50 centroids (50) and the genefu package in R. E) Association of the claudin-low nine-cell line predictive genes with MDA-231 cells following siDUSP4 or selumetinib treatment (5).

    Journal: Cancer research

    Article Title: Activation of MAPK pathways due to DUSP4 loss promotes cancer stem cell-like phenotypes in basal-like breast cancer

    doi: 10.1158/0008-5472.CAN-13-1385

    Figure Lengend Snippet: Microarray analysis was performed on RNA derived from MDA-231, BT549 and SUM159PT cells 96 hr after transfection with siCONTROL or siDUSP4 and after 4 hr or 24 hr of selumetinib treatment. A) Heatmap of significantly altered genes for MDA-231 cells (ANOVA FDR-adjusted p<0.01). B) DUSP4 loss gene expression score derived from MDA-231 cells transfected with siDUSP4 was calculated across all samples. C) Expression of selected chemotherapy-resistance genes which were altered in MDA-231 cells transfected with siDUSP4. D) DUSP4-loss gene expression score derived from MDA-231 cells transfected with siDUSP4 was calculated across 444 tumors in the TCGA breast data. Molecular subtype was determined using the PAM50 centroids (50) and the genefu package in R. E) Association of the claudin-low nine-cell line predictive genes with MDA-231 cells following siDUSP4 or selumetinib treatment (5).

    Article Snippet: Antibodies used for immunoblotting were: p-ERK1/2 (p-T202/Y204; #9101), calnexin (#2433), p-cJUN (#2361), cJUN (#9165), p-JNK1/2 (#4668), JNK1/2 (#9252) (all from Cell Signaling), p-ETS-1(p-T38, Invitrogen 44-1104G), ETS-1 (Santa Cruz sc-350), DUSP4 (Cell Signaling #5419), Anti-HA tag (Santa Cruz sc-805) and actin (Sigma, A2066).

    Techniques: Microarray, Derivative Assay, Transfection, Expressing

    SUM159PT cells transduced with DOX-inducible DUSP4-HA were generated. A) Flow cytometry analysis of CD44/CD24 expression in cells treated for 1-10 days with DOX (2 ng/mL). B) SUM159PT/pINDUCER-LACZ cells or SUM159PT/pINDUCER-DUSP4 pre-treated for 4 days ± DOX prior to plating in a mammosphere assay in the presence or absence of DOX as in the pretreatment. C) Immunoblot of parental SUM159PT/pINDUCER-LACZ cells or SUM159PT/pINDUCER-DUSP4 treated for 4 days with DOX. D) Tumor formation of cells from (C) 60 days after injection the mammary fatpad of athymic mice. Red bars indicate palpable tumors and blue bars indicate tumors identified on histological examination of the injection site. E) Schematic of proposed model based on the presented data: The tumor suppressor DUSP4 negatively regulates ERK and JNK. Upon loss of DUSP4, derepressed ERK and JNK activity stimulates ETS1 and cJUN-mediated transcription of IL6 and IL8, cytokines that expand the cancer stem-like cell population. De-repressed ERK transcription following DUSP4 loss also suppresses CD24 expression thus increasing the CD44HI CD24LO compartment, a marker of the CSC population.

    Journal: Cancer research

    Article Title: Activation of MAPK pathways due to DUSP4 loss promotes cancer stem cell-like phenotypes in basal-like breast cancer

    doi: 10.1158/0008-5472.CAN-13-1385

    Figure Lengend Snippet: SUM159PT cells transduced with DOX-inducible DUSP4-HA were generated. A) Flow cytometry analysis of CD44/CD24 expression in cells treated for 1-10 days with DOX (2 ng/mL). B) SUM159PT/pINDUCER-LACZ cells or SUM159PT/pINDUCER-DUSP4 pre-treated for 4 days ± DOX prior to plating in a mammosphere assay in the presence or absence of DOX as in the pretreatment. C) Immunoblot of parental SUM159PT/pINDUCER-LACZ cells or SUM159PT/pINDUCER-DUSP4 treated for 4 days with DOX. D) Tumor formation of cells from (C) 60 days after injection the mammary fatpad of athymic mice. Red bars indicate palpable tumors and blue bars indicate tumors identified on histological examination of the injection site. E) Schematic of proposed model based on the presented data: The tumor suppressor DUSP4 negatively regulates ERK and JNK. Upon loss of DUSP4, derepressed ERK and JNK activity stimulates ETS1 and cJUN-mediated transcription of IL6 and IL8, cytokines that expand the cancer stem-like cell population. De-repressed ERK transcription following DUSP4 loss also suppresses CD24 expression thus increasing the CD44HI CD24LO compartment, a marker of the CSC population.

    Article Snippet: Antibodies used for immunoblotting were: p-ERK1/2 (p-T202/Y204; #9101), calnexin (#2433), p-cJUN (#2361), cJUN (#9165), p-JNK1/2 (#4668), JNK1/2 (#9252) (all from Cell Signaling), p-ETS-1(p-T38, Invitrogen 44-1104G), ETS-1 (Santa Cruz sc-350), DUSP4 (Cell Signaling #5419), Anti-HA tag (Santa Cruz sc-805) and actin (Sigma, A2066).

    Techniques: Transduction, Generated, Flow Cytometry, Expressing, Western Blot, Injection, Activity Assay, Marker