rabbit polyclonal anti cdk1 t161p  (Cell Signaling Technology Inc)


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Bioz Manufacturer Symbol Cell Signaling Technology Inc manufactures this product  
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    Structured Review

    Cell Signaling Technology Inc rabbit polyclonal anti cdk1 t161p
    RNAi duplex sequences
    Rabbit Polyclonal Anti Cdk1 T161p, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal anti cdk1 t161p/product/Cell Signaling Technology Inc
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal anti cdk1 t161p - by Bioz Stars, 2023-09
    95/100 stars

    Images

    1) Product Images from "Phosphorylation of islet-1 serine 269 by CDK1 increases its transcriptional activity and promotes cell proliferation in gastric cancer"

    Article Title: Phosphorylation of islet-1 serine 269 by CDK1 increases its transcriptional activity and promotes cell proliferation in gastric cancer

    Journal: Molecular Medicine

    doi: 10.1186/s10020-021-00302-6

    RNAi duplex sequences
    Figure Legend Snippet: RNAi duplex sequences

    Techniques Used:

    ISL1 interacts with CDK1. a–c Co-IP assays were performed to measure the ISL1/CDK1/cyclinB1 complex in MGC803 cells. Lysate was immunoprecipitated with antibodies against ISL1, CDK1, or cyclinB1 and analyzed by immunoblotting. IgG: immunoglobulin G. d Full-length or truncated ISL1 was used to construct GST-fusion proteins (Top) for pull-down assays with CDK1 protein (Bottom)
    Figure Legend Snippet: ISL1 interacts with CDK1. a–c Co-IP assays were performed to measure the ISL1/CDK1/cyclinB1 complex in MGC803 cells. Lysate was immunoprecipitated with antibodies against ISL1, CDK1, or cyclinB1 and analyzed by immunoblotting. IgG: immunoglobulin G. d Full-length or truncated ISL1 was used to construct GST-fusion proteins (Top) for pull-down assays with CDK1 protein (Bottom)

    Techniques Used: Co-Immunoprecipitation Assay, Immunoprecipitation, Western Blot, Construct

    ISL1 is serine-phosphorylated. a MGC803 cells lysates were immunoprecipitated (IP) using anti‑ISL1 and analyzed by immunoblotting using antibody against phosphorylated serine and ISL1 (HC: heavy chain, LC: light chain). b Alignments of the cluster of ISL1; S269 fit the CDK1 phosphorylation consensus, and (S/T)PX(R/K) sequences (red shade) are shown
    Figure Legend Snippet: ISL1 is serine-phosphorylated. a MGC803 cells lysates were immunoprecipitated (IP) using anti‑ISL1 and analyzed by immunoblotting using antibody against phosphorylated serine and ISL1 (HC: heavy chain, LC: light chain). b Alignments of the cluster of ISL1; S269 fit the CDK1 phosphorylation consensus, and (S/T)PX(R/K) sequences (red shade) are shown

    Techniques Used: Immunoprecipitation, Western Blot

    CDK1 mediates ISL1 phosphorylation at serine 269. a MGC803 cells transfected with plasmid encoding CDK1 or control vector (left), negative control or CDK1 si-RNA (right). Lysate was analyzed by immunoblotting with antibodies against the indicated proteins. b MGC803 cells were transfected with plasmids encoding dominant‑negative mutant CDK1 (DN‑CDK1) or CDK1, and lysate was analyzed by immunoblotting with antibodies against the indicated proteins. c MKN28 cells expressing wild-type ISL1 or ISL1-S269A were transfected with plasmid encoding CDK1 or control vector. Cell lysates were immunoblotted with antibodies against the indicated proteins. d MKN28 cells were transfected with plasmids encoding wild-type ISL1 or ISL1-S269A and treated with CDK1 inhibitor RO-3306 or DMSO. Lysates were analyzed by immunoblotting with antibodies against the indicated proteins. GAPDH served as the internal control. e Representative IHC staining of ISL1-S269-p expression in GC (right, n = 60) and paired adjacent tissues (left, n = 60). f Relationship between expression of ISL1-S629-p and overall survival in gastric cancer
    Figure Legend Snippet: CDK1 mediates ISL1 phosphorylation at serine 269. a MGC803 cells transfected with plasmid encoding CDK1 or control vector (left), negative control or CDK1 si-RNA (right). Lysate was analyzed by immunoblotting with antibodies against the indicated proteins. b MGC803 cells were transfected with plasmids encoding dominant‑negative mutant CDK1 (DN‑CDK1) or CDK1, and lysate was analyzed by immunoblotting with antibodies against the indicated proteins. c MKN28 cells expressing wild-type ISL1 or ISL1-S269A were transfected with plasmid encoding CDK1 or control vector. Cell lysates were immunoblotted with antibodies against the indicated proteins. d MKN28 cells were transfected with plasmids encoding wild-type ISL1 or ISL1-S269A and treated with CDK1 inhibitor RO-3306 or DMSO. Lysates were analyzed by immunoblotting with antibodies against the indicated proteins. GAPDH served as the internal control. e Representative IHC staining of ISL1-S269-p expression in GC (right, n = 60) and paired adjacent tissues (left, n = 60). f Relationship between expression of ISL1-S629-p and overall survival in gastric cancer

    Techniques Used: Transfection, Plasmid Preparation, Negative Control, Western Blot, Dominant Negative Mutation, Expressing, Immunohistochemistry

    Phosphorylated ISL1 showed improved localization on cyclinB1 and cyclinB2 promoters. a ChIP assay analysis and PCR gel electrophoresis analysis of ISL-1 recruitment to the cyclinB1 and cyclinB2 promoters in MGC803 cells. b ChIP-re-IP assay was performed with anti-ISL-1 or rabbit IgG antibodies and then with anti-CDK1 or IgG antibodies for immunoprecipitation using chromatin harvested from MGC803 cells. c , d Quantitative chromatin immunoprecipitation analysis of cyclinB1 and cyclinB2 promoters in MKN28 cell line transfected with control vector or expressing wild-type ISL1 or ISL1-S269A plasmids. Data are expressed as mean ± s. d. from three individual experiments. (* p < 0.05, WT vs. control, # p < 0.05, S269A vs. control, $ p < 0.05, WT vs. S269A)
    Figure Legend Snippet: Phosphorylated ISL1 showed improved localization on cyclinB1 and cyclinB2 promoters. a ChIP assay analysis and PCR gel electrophoresis analysis of ISL-1 recruitment to the cyclinB1 and cyclinB2 promoters in MGC803 cells. b ChIP-re-IP assay was performed with anti-ISL-1 or rabbit IgG antibodies and then with anti-CDK1 or IgG antibodies for immunoprecipitation using chromatin harvested from MGC803 cells. c , d Quantitative chromatin immunoprecipitation analysis of cyclinB1 and cyclinB2 promoters in MKN28 cell line transfected with control vector or expressing wild-type ISL1 or ISL1-S269A plasmids. Data are expressed as mean ± s. d. from three individual experiments. (* p < 0.05, WT vs. control, # p < 0.05, S269A vs. control, $ p < 0.05, WT vs. S269A)

    Techniques Used: Nucleic Acid Electrophoresis, Immunoprecipitation, Chromatin Immunoprecipitation, Transfection, Plasmid Preparation, Expressing

    Phosphorylation Ser 269 of ISL1 increases transactivation potency. a , b Effect of CDK1 activity on the transcriptional activity of wild-type ISL1 or S269A ISL1. Transcriptional activity of wild-type ISL1 and S269A ISL1 on cyclinB1-luc and cyclinB2-luc determined by luciferase reporter assay in MKN28 cells. c Cell proliferation was studied by CCK-8 analysis of MKN28 cells transfected with plasmids encoding control vector and wild-type ISL1 or ISL1-S269A. d Cell cycle distributions were analyzed by flow cytometry in MKN28 transfected with plasmids encoding control vector and wild-type ISL1 or ISL1-S269A. The data represent 3 independent experiments, each performed in triplicate. Each bar represents mean ± SD. (* p < 0.05, WT vs. control, # p < 0.05, S269A vs. control, $ p < 0.05, WT vs. S269A)
    Figure Legend Snippet: Phosphorylation Ser 269 of ISL1 increases transactivation potency. a , b Effect of CDK1 activity on the transcriptional activity of wild-type ISL1 or S269A ISL1. Transcriptional activity of wild-type ISL1 and S269A ISL1 on cyclinB1-luc and cyclinB2-luc determined by luciferase reporter assay in MKN28 cells. c Cell proliferation was studied by CCK-8 analysis of MKN28 cells transfected with plasmids encoding control vector and wild-type ISL1 or ISL1-S269A. d Cell cycle distributions were analyzed by flow cytometry in MKN28 transfected with plasmids encoding control vector and wild-type ISL1 or ISL1-S269A. The data represent 3 independent experiments, each performed in triplicate. Each bar represents mean ± SD. (* p < 0.05, WT vs. control, # p < 0.05, S269A vs. control, $ p < 0.05, WT vs. S269A)

    Techniques Used: Activity Assay, Luciferase, Reporter Assay, CCK-8 Assay, Transfection, Plasmid Preparation, Flow Cytometry

    Phosphorylation of ISL1 increases self-stability but does not increase its interaction with CDK1. a NIH3T3 cells were transfected with wild-type ISL1 or ISL1-S269A plasmids. After 48 h, cells were treated with CHX (100 µg/ml) for 12 h. Whole cell extracts were harvested for western blotting. b Co-IP assays were performed in NIH3T3 cells to detect the interaction between ISL1 (wild-type or mutants) and CDK1. GAPDH served as a loading and negative control. c Schematic representation of positive feedback regulation by the ISL1-CDK1/cyclin B1/2 complex in GC. Full arrow: confirmed; broken arrow: to be verified
    Figure Legend Snippet: Phosphorylation of ISL1 increases self-stability but does not increase its interaction with CDK1. a NIH3T3 cells were transfected with wild-type ISL1 or ISL1-S269A plasmids. After 48 h, cells were treated with CHX (100 µg/ml) for 12 h. Whole cell extracts were harvested for western blotting. b Co-IP assays were performed in NIH3T3 cells to detect the interaction between ISL1 (wild-type or mutants) and CDK1. GAPDH served as a loading and negative control. c Schematic representation of positive feedback regulation by the ISL1-CDK1/cyclin B1/2 complex in GC. Full arrow: confirmed; broken arrow: to be verified

    Techniques Used: Transfection, Western Blot, Co-Immunoprecipitation Assay, Negative Control

    rabbit polyclonal anti cdk1 t161p  (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  
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  • 95

    Structured Review

    Cell Signaling Technology Inc rabbit polyclonal anti cdk1 t161p
    RNAi duplex sequences
    Rabbit Polyclonal Anti Cdk1 T161p, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal anti cdk1 t161p/product/Cell Signaling Technology Inc
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal anti cdk1 t161p - by Bioz Stars, 2023-09
    95/100 stars

    Images

    1) Product Images from "Phosphorylation of islet-1 serine 269 by CDK1 increases its transcriptional activity and promotes cell proliferation in gastric cancer"

    Article Title: Phosphorylation of islet-1 serine 269 by CDK1 increases its transcriptional activity and promotes cell proliferation in gastric cancer

    Journal: Molecular Medicine

    doi: 10.1186/s10020-021-00302-6

    RNAi duplex sequences
    Figure Legend Snippet: RNAi duplex sequences

    Techniques Used:

    ISL1 interacts with CDK1. a–c Co-IP assays were performed to measure the ISL1/CDK1/cyclinB1 complex in MGC803 cells. Lysate was immunoprecipitated with antibodies against ISL1, CDK1, or cyclinB1 and analyzed by immunoblotting. IgG: immunoglobulin G. d Full-length or truncated ISL1 was used to construct GST-fusion proteins (Top) for pull-down assays with CDK1 protein (Bottom)
    Figure Legend Snippet: ISL1 interacts with CDK1. a–c Co-IP assays were performed to measure the ISL1/CDK1/cyclinB1 complex in MGC803 cells. Lysate was immunoprecipitated with antibodies against ISL1, CDK1, or cyclinB1 and analyzed by immunoblotting. IgG: immunoglobulin G. d Full-length or truncated ISL1 was used to construct GST-fusion proteins (Top) for pull-down assays with CDK1 protein (Bottom)

    Techniques Used: Co-Immunoprecipitation Assay, Immunoprecipitation, Western Blot, Construct

    ISL1 is serine-phosphorylated. a MGC803 cells lysates were immunoprecipitated (IP) using anti‑ISL1 and analyzed by immunoblotting using antibody against phosphorylated serine and ISL1 (HC: heavy chain, LC: light chain). b Alignments of the cluster of ISL1; S269 fit the CDK1 phosphorylation consensus, and (S/T)PX(R/K) sequences (red shade) are shown
    Figure Legend Snippet: ISL1 is serine-phosphorylated. a MGC803 cells lysates were immunoprecipitated (IP) using anti‑ISL1 and analyzed by immunoblotting using antibody against phosphorylated serine and ISL1 (HC: heavy chain, LC: light chain). b Alignments of the cluster of ISL1; S269 fit the CDK1 phosphorylation consensus, and (S/T)PX(R/K) sequences (red shade) are shown

    Techniques Used: Immunoprecipitation, Western Blot

    CDK1 mediates ISL1 phosphorylation at serine 269. a MGC803 cells transfected with plasmid encoding CDK1 or control vector (left), negative control or CDK1 si-RNA (right). Lysate was analyzed by immunoblotting with antibodies against the indicated proteins. b MGC803 cells were transfected with plasmids encoding dominant‑negative mutant CDK1 (DN‑CDK1) or CDK1, and lysate was analyzed by immunoblotting with antibodies against the indicated proteins. c MKN28 cells expressing wild-type ISL1 or ISL1-S269A were transfected with plasmid encoding CDK1 or control vector. Cell lysates were immunoblotted with antibodies against the indicated proteins. d MKN28 cells were transfected with plasmids encoding wild-type ISL1 or ISL1-S269A and treated with CDK1 inhibitor RO-3306 or DMSO. Lysates were analyzed by immunoblotting with antibodies against the indicated proteins. GAPDH served as the internal control. e Representative IHC staining of ISL1-S269-p expression in GC (right, n = 60) and paired adjacent tissues (left, n = 60). f Relationship between expression of ISL1-S629-p and overall survival in gastric cancer
    Figure Legend Snippet: CDK1 mediates ISL1 phosphorylation at serine 269. a MGC803 cells transfected with plasmid encoding CDK1 or control vector (left), negative control or CDK1 si-RNA (right). Lysate was analyzed by immunoblotting with antibodies against the indicated proteins. b MGC803 cells were transfected with plasmids encoding dominant‑negative mutant CDK1 (DN‑CDK1) or CDK1, and lysate was analyzed by immunoblotting with antibodies against the indicated proteins. c MKN28 cells expressing wild-type ISL1 or ISL1-S269A were transfected with plasmid encoding CDK1 or control vector. Cell lysates were immunoblotted with antibodies against the indicated proteins. d MKN28 cells were transfected with plasmids encoding wild-type ISL1 or ISL1-S269A and treated with CDK1 inhibitor RO-3306 or DMSO. Lysates were analyzed by immunoblotting with antibodies against the indicated proteins. GAPDH served as the internal control. e Representative IHC staining of ISL1-S269-p expression in GC (right, n = 60) and paired adjacent tissues (left, n = 60). f Relationship between expression of ISL1-S629-p and overall survival in gastric cancer

    Techniques Used: Transfection, Plasmid Preparation, Negative Control, Western Blot, Dominant Negative Mutation, Expressing, Immunohistochemistry

    Phosphorylated ISL1 showed improved localization on cyclinB1 and cyclinB2 promoters. a ChIP assay analysis and PCR gel electrophoresis analysis of ISL-1 recruitment to the cyclinB1 and cyclinB2 promoters in MGC803 cells. b ChIP-re-IP assay was performed with anti-ISL-1 or rabbit IgG antibodies and then with anti-CDK1 or IgG antibodies for immunoprecipitation using chromatin harvested from MGC803 cells. c , d Quantitative chromatin immunoprecipitation analysis of cyclinB1 and cyclinB2 promoters in MKN28 cell line transfected with control vector or expressing wild-type ISL1 or ISL1-S269A plasmids. Data are expressed as mean ± s. d. from three individual experiments. (* p < 0.05, WT vs. control, # p < 0.05, S269A vs. control, $ p < 0.05, WT vs. S269A)
    Figure Legend Snippet: Phosphorylated ISL1 showed improved localization on cyclinB1 and cyclinB2 promoters. a ChIP assay analysis and PCR gel electrophoresis analysis of ISL-1 recruitment to the cyclinB1 and cyclinB2 promoters in MGC803 cells. b ChIP-re-IP assay was performed with anti-ISL-1 or rabbit IgG antibodies and then with anti-CDK1 or IgG antibodies for immunoprecipitation using chromatin harvested from MGC803 cells. c , d Quantitative chromatin immunoprecipitation analysis of cyclinB1 and cyclinB2 promoters in MKN28 cell line transfected with control vector or expressing wild-type ISL1 or ISL1-S269A plasmids. Data are expressed as mean ± s. d. from three individual experiments. (* p < 0.05, WT vs. control, # p < 0.05, S269A vs. control, $ p < 0.05, WT vs. S269A)

    Techniques Used: Nucleic Acid Electrophoresis, Immunoprecipitation, Chromatin Immunoprecipitation, Transfection, Plasmid Preparation, Expressing

    Phosphorylation Ser 269 of ISL1 increases transactivation potency. a , b Effect of CDK1 activity on the transcriptional activity of wild-type ISL1 or S269A ISL1. Transcriptional activity of wild-type ISL1 and S269A ISL1 on cyclinB1-luc and cyclinB2-luc determined by luciferase reporter assay in MKN28 cells. c Cell proliferation was studied by CCK-8 analysis of MKN28 cells transfected with plasmids encoding control vector and wild-type ISL1 or ISL1-S269A. d Cell cycle distributions were analyzed by flow cytometry in MKN28 transfected with plasmids encoding control vector and wild-type ISL1 or ISL1-S269A. The data represent 3 independent experiments, each performed in triplicate. Each bar represents mean ± SD. (* p < 0.05, WT vs. control, # p < 0.05, S269A vs. control, $ p < 0.05, WT vs. S269A)
    Figure Legend Snippet: Phosphorylation Ser 269 of ISL1 increases transactivation potency. a , b Effect of CDK1 activity on the transcriptional activity of wild-type ISL1 or S269A ISL1. Transcriptional activity of wild-type ISL1 and S269A ISL1 on cyclinB1-luc and cyclinB2-luc determined by luciferase reporter assay in MKN28 cells. c Cell proliferation was studied by CCK-8 analysis of MKN28 cells transfected with plasmids encoding control vector and wild-type ISL1 or ISL1-S269A. d Cell cycle distributions were analyzed by flow cytometry in MKN28 transfected with plasmids encoding control vector and wild-type ISL1 or ISL1-S269A. The data represent 3 independent experiments, each performed in triplicate. Each bar represents mean ± SD. (* p < 0.05, WT vs. control, # p < 0.05, S269A vs. control, $ p < 0.05, WT vs. S269A)

    Techniques Used: Activity Assay, Luciferase, Reporter Assay, CCK-8 Assay, Transfection, Plasmid Preparation, Flow Cytometry

    Phosphorylation of ISL1 increases self-stability but does not increase its interaction with CDK1. a NIH3T3 cells were transfected with wild-type ISL1 or ISL1-S269A plasmids. After 48 h, cells were treated with CHX (100 µg/ml) for 12 h. Whole cell extracts were harvested for western blotting. b Co-IP assays were performed in NIH3T3 cells to detect the interaction between ISL1 (wild-type or mutants) and CDK1. GAPDH served as a loading and negative control. c Schematic representation of positive feedback regulation by the ISL1-CDK1/cyclin B1/2 complex in GC. Full arrow: confirmed; broken arrow: to be verified
    Figure Legend Snippet: Phosphorylation of ISL1 increases self-stability but does not increase its interaction with CDK1. a NIH3T3 cells were transfected with wild-type ISL1 or ISL1-S269A plasmids. After 48 h, cells were treated with CHX (100 µg/ml) for 12 h. Whole cell extracts were harvested for western blotting. b Co-IP assays were performed in NIH3T3 cells to detect the interaction between ISL1 (wild-type or mutants) and CDK1. GAPDH served as a loading and negative control. c Schematic representation of positive feedback regulation by the ISL1-CDK1/cyclin B1/2 complex in GC. Full arrow: confirmed; broken arrow: to be verified

    Techniques Used: Transfection, Western Blot, Co-Immunoprecipitation Assay, Negative Control

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    Cell Signaling Technology Inc rabbit polyclonal anti cdk1 t161p
    RNAi duplex sequences
    Rabbit Polyclonal Anti Cdk1 T161p, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit polyclonal anti cdk1 t161p/product/Cell Signaling Technology Inc
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit polyclonal anti cdk1 t161p - by Bioz Stars, 2023-09
    95/100 stars
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    RNAi duplex sequences

    Journal: Molecular Medicine

    Article Title: Phosphorylation of islet-1 serine 269 by CDK1 increases its transcriptional activity and promotes cell proliferation in gastric cancer

    doi: 10.1186/s10020-021-00302-6

    Figure Lengend Snippet: RNAi duplex sequences

    Article Snippet: Monoclonal anti-CDK1 (#9116), GAPDH (#2118), rabbit polyclonal anti-CDK1-T161P (#9114), and mouse monoclonal anti-cyclin B1 (#4135) were purchased from Cell Signaling Technology (Danvers, MA, USA).

    Techniques:

    ISL1 interacts with CDK1. a–c Co-IP assays were performed to measure the ISL1/CDK1/cyclinB1 complex in MGC803 cells. Lysate was immunoprecipitated with antibodies against ISL1, CDK1, or cyclinB1 and analyzed by immunoblotting. IgG: immunoglobulin G. d Full-length or truncated ISL1 was used to construct GST-fusion proteins (Top) for pull-down assays with CDK1 protein (Bottom)

    Journal: Molecular Medicine

    Article Title: Phosphorylation of islet-1 serine 269 by CDK1 increases its transcriptional activity and promotes cell proliferation in gastric cancer

    doi: 10.1186/s10020-021-00302-6

    Figure Lengend Snippet: ISL1 interacts with CDK1. a–c Co-IP assays were performed to measure the ISL1/CDK1/cyclinB1 complex in MGC803 cells. Lysate was immunoprecipitated with antibodies against ISL1, CDK1, or cyclinB1 and analyzed by immunoblotting. IgG: immunoglobulin G. d Full-length or truncated ISL1 was used to construct GST-fusion proteins (Top) for pull-down assays with CDK1 protein (Bottom)

    Article Snippet: Monoclonal anti-CDK1 (#9116), GAPDH (#2118), rabbit polyclonal anti-CDK1-T161P (#9114), and mouse monoclonal anti-cyclin B1 (#4135) were purchased from Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Co-Immunoprecipitation Assay, Immunoprecipitation, Western Blot, Construct

    ISL1 is serine-phosphorylated. a MGC803 cells lysates were immunoprecipitated (IP) using anti‑ISL1 and analyzed by immunoblotting using antibody against phosphorylated serine and ISL1 (HC: heavy chain, LC: light chain). b Alignments of the cluster of ISL1; S269 fit the CDK1 phosphorylation consensus, and (S/T)PX(R/K) sequences (red shade) are shown

    Journal: Molecular Medicine

    Article Title: Phosphorylation of islet-1 serine 269 by CDK1 increases its transcriptional activity and promotes cell proliferation in gastric cancer

    doi: 10.1186/s10020-021-00302-6

    Figure Lengend Snippet: ISL1 is serine-phosphorylated. a MGC803 cells lysates were immunoprecipitated (IP) using anti‑ISL1 and analyzed by immunoblotting using antibody against phosphorylated serine and ISL1 (HC: heavy chain, LC: light chain). b Alignments of the cluster of ISL1; S269 fit the CDK1 phosphorylation consensus, and (S/T)PX(R/K) sequences (red shade) are shown

    Article Snippet: Monoclonal anti-CDK1 (#9116), GAPDH (#2118), rabbit polyclonal anti-CDK1-T161P (#9114), and mouse monoclonal anti-cyclin B1 (#4135) were purchased from Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Immunoprecipitation, Western Blot

    CDK1 mediates ISL1 phosphorylation at serine 269. a MGC803 cells transfected with plasmid encoding CDK1 or control vector (left), negative control or CDK1 si-RNA (right). Lysate was analyzed by immunoblotting with antibodies against the indicated proteins. b MGC803 cells were transfected with plasmids encoding dominant‑negative mutant CDK1 (DN‑CDK1) or CDK1, and lysate was analyzed by immunoblotting with antibodies against the indicated proteins. c MKN28 cells expressing wild-type ISL1 or ISL1-S269A were transfected with plasmid encoding CDK1 or control vector. Cell lysates were immunoblotted with antibodies against the indicated proteins. d MKN28 cells were transfected with plasmids encoding wild-type ISL1 or ISL1-S269A and treated with CDK1 inhibitor RO-3306 or DMSO. Lysates were analyzed by immunoblotting with antibodies against the indicated proteins. GAPDH served as the internal control. e Representative IHC staining of ISL1-S269-p expression in GC (right, n = 60) and paired adjacent tissues (left, n = 60). f Relationship between expression of ISL1-S629-p and overall survival in gastric cancer

    Journal: Molecular Medicine

    Article Title: Phosphorylation of islet-1 serine 269 by CDK1 increases its transcriptional activity and promotes cell proliferation in gastric cancer

    doi: 10.1186/s10020-021-00302-6

    Figure Lengend Snippet: CDK1 mediates ISL1 phosphorylation at serine 269. a MGC803 cells transfected with plasmid encoding CDK1 or control vector (left), negative control or CDK1 si-RNA (right). Lysate was analyzed by immunoblotting with antibodies against the indicated proteins. b MGC803 cells were transfected with plasmids encoding dominant‑negative mutant CDK1 (DN‑CDK1) or CDK1, and lysate was analyzed by immunoblotting with antibodies against the indicated proteins. c MKN28 cells expressing wild-type ISL1 or ISL1-S269A were transfected with plasmid encoding CDK1 or control vector. Cell lysates were immunoblotted with antibodies against the indicated proteins. d MKN28 cells were transfected with plasmids encoding wild-type ISL1 or ISL1-S269A and treated with CDK1 inhibitor RO-3306 or DMSO. Lysates were analyzed by immunoblotting with antibodies against the indicated proteins. GAPDH served as the internal control. e Representative IHC staining of ISL1-S269-p expression in GC (right, n = 60) and paired adjacent tissues (left, n = 60). f Relationship between expression of ISL1-S629-p and overall survival in gastric cancer

    Article Snippet: Monoclonal anti-CDK1 (#9116), GAPDH (#2118), rabbit polyclonal anti-CDK1-T161P (#9114), and mouse monoclonal anti-cyclin B1 (#4135) were purchased from Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Transfection, Plasmid Preparation, Negative Control, Western Blot, Dominant Negative Mutation, Expressing, Immunohistochemistry

    Phosphorylated ISL1 showed improved localization on cyclinB1 and cyclinB2 promoters. a ChIP assay analysis and PCR gel electrophoresis analysis of ISL-1 recruitment to the cyclinB1 and cyclinB2 promoters in MGC803 cells. b ChIP-re-IP assay was performed with anti-ISL-1 or rabbit IgG antibodies and then with anti-CDK1 or IgG antibodies for immunoprecipitation using chromatin harvested from MGC803 cells. c , d Quantitative chromatin immunoprecipitation analysis of cyclinB1 and cyclinB2 promoters in MKN28 cell line transfected with control vector or expressing wild-type ISL1 or ISL1-S269A plasmids. Data are expressed as mean ± s. d. from three individual experiments. (* p < 0.05, WT vs. control, # p < 0.05, S269A vs. control, $ p < 0.05, WT vs. S269A)

    Journal: Molecular Medicine

    Article Title: Phosphorylation of islet-1 serine 269 by CDK1 increases its transcriptional activity and promotes cell proliferation in gastric cancer

    doi: 10.1186/s10020-021-00302-6

    Figure Lengend Snippet: Phosphorylated ISL1 showed improved localization on cyclinB1 and cyclinB2 promoters. a ChIP assay analysis and PCR gel electrophoresis analysis of ISL-1 recruitment to the cyclinB1 and cyclinB2 promoters in MGC803 cells. b ChIP-re-IP assay was performed with anti-ISL-1 or rabbit IgG antibodies and then with anti-CDK1 or IgG antibodies for immunoprecipitation using chromatin harvested from MGC803 cells. c , d Quantitative chromatin immunoprecipitation analysis of cyclinB1 and cyclinB2 promoters in MKN28 cell line transfected with control vector or expressing wild-type ISL1 or ISL1-S269A plasmids. Data are expressed as mean ± s. d. from three individual experiments. (* p < 0.05, WT vs. control, # p < 0.05, S269A vs. control, $ p < 0.05, WT vs. S269A)

    Article Snippet: Monoclonal anti-CDK1 (#9116), GAPDH (#2118), rabbit polyclonal anti-CDK1-T161P (#9114), and mouse monoclonal anti-cyclin B1 (#4135) were purchased from Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Nucleic Acid Electrophoresis, Immunoprecipitation, Chromatin Immunoprecipitation, Transfection, Plasmid Preparation, Expressing

    Phosphorylation Ser 269 of ISL1 increases transactivation potency. a , b Effect of CDK1 activity on the transcriptional activity of wild-type ISL1 or S269A ISL1. Transcriptional activity of wild-type ISL1 and S269A ISL1 on cyclinB1-luc and cyclinB2-luc determined by luciferase reporter assay in MKN28 cells. c Cell proliferation was studied by CCK-8 analysis of MKN28 cells transfected with plasmids encoding control vector and wild-type ISL1 or ISL1-S269A. d Cell cycle distributions were analyzed by flow cytometry in MKN28 transfected with plasmids encoding control vector and wild-type ISL1 or ISL1-S269A. The data represent 3 independent experiments, each performed in triplicate. Each bar represents mean ± SD. (* p < 0.05, WT vs. control, # p < 0.05, S269A vs. control, $ p < 0.05, WT vs. S269A)

    Journal: Molecular Medicine

    Article Title: Phosphorylation of islet-1 serine 269 by CDK1 increases its transcriptional activity and promotes cell proliferation in gastric cancer

    doi: 10.1186/s10020-021-00302-6

    Figure Lengend Snippet: Phosphorylation Ser 269 of ISL1 increases transactivation potency. a , b Effect of CDK1 activity on the transcriptional activity of wild-type ISL1 or S269A ISL1. Transcriptional activity of wild-type ISL1 and S269A ISL1 on cyclinB1-luc and cyclinB2-luc determined by luciferase reporter assay in MKN28 cells. c Cell proliferation was studied by CCK-8 analysis of MKN28 cells transfected with plasmids encoding control vector and wild-type ISL1 or ISL1-S269A. d Cell cycle distributions were analyzed by flow cytometry in MKN28 transfected with plasmids encoding control vector and wild-type ISL1 or ISL1-S269A. The data represent 3 independent experiments, each performed in triplicate. Each bar represents mean ± SD. (* p < 0.05, WT vs. control, # p < 0.05, S269A vs. control, $ p < 0.05, WT vs. S269A)

    Article Snippet: Monoclonal anti-CDK1 (#9116), GAPDH (#2118), rabbit polyclonal anti-CDK1-T161P (#9114), and mouse monoclonal anti-cyclin B1 (#4135) were purchased from Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Activity Assay, Luciferase, Reporter Assay, CCK-8 Assay, Transfection, Plasmid Preparation, Flow Cytometry

    Phosphorylation of ISL1 increases self-stability but does not increase its interaction with CDK1. a NIH3T3 cells were transfected with wild-type ISL1 or ISL1-S269A plasmids. After 48 h, cells were treated with CHX (100 µg/ml) for 12 h. Whole cell extracts were harvested for western blotting. b Co-IP assays were performed in NIH3T3 cells to detect the interaction between ISL1 (wild-type or mutants) and CDK1. GAPDH served as a loading and negative control. c Schematic representation of positive feedback regulation by the ISL1-CDK1/cyclin B1/2 complex in GC. Full arrow: confirmed; broken arrow: to be verified

    Journal: Molecular Medicine

    Article Title: Phosphorylation of islet-1 serine 269 by CDK1 increases its transcriptional activity and promotes cell proliferation in gastric cancer

    doi: 10.1186/s10020-021-00302-6

    Figure Lengend Snippet: Phosphorylation of ISL1 increases self-stability but does not increase its interaction with CDK1. a NIH3T3 cells were transfected with wild-type ISL1 or ISL1-S269A plasmids. After 48 h, cells were treated with CHX (100 µg/ml) for 12 h. Whole cell extracts were harvested for western blotting. b Co-IP assays were performed in NIH3T3 cells to detect the interaction between ISL1 (wild-type or mutants) and CDK1. GAPDH served as a loading and negative control. c Schematic representation of positive feedback regulation by the ISL1-CDK1/cyclin B1/2 complex in GC. Full arrow: confirmed; broken arrow: to be verified

    Article Snippet: Monoclonal anti-CDK1 (#9116), GAPDH (#2118), rabbit polyclonal anti-CDK1-T161P (#9114), and mouse monoclonal anti-cyclin B1 (#4135) were purchased from Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Transfection, Western Blot, Co-Immunoprecipitation Assay, Negative Control