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rabbit anti fignl1 proteintech 17604 1 ap wb (Proteintech)

Name: rabbit anti fignl1 proteintech 17604 1 ap wb
Brand: Proteintech
Rating: 93 (9 reviews)

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Proteintech rabbit anti fignl1 proteintech 17604 1 ap wb
Rabbit Anti Fignl1 Proteintech 17604 1 Ap Wb, supplied by Proteintech, used in various techniques. Bioz Stars score: 93/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/rabbit anti fignl1 proteintech 17604 1 ap wb/product/Proteintech
Average 93 stars, based on 9 article reviews

rabbit anti fignl1 proteintech 17604 1 ap wb - by Bioz Stars, 2026-02

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Journal: bioRxiv

Article Title: FIGNL1 inhibits homologous recombination in BRCA2 deficient cells by dissociating RAD51 filaments

doi: 10.1101/2024.11.03.621741

Figure Lengend Snippet: (A) Left: Schematics of the results of knocking-out FIGNL1 in RPE1-WT cells. Right: Genotyping results from RPE1-WT and 2 different knockout clones of FIGNL1 . Electropherogram representative figure showing the alignment of the FIGNL1 genomic DNA region at the indicated binding site of sgRNA followed by its PAM sequence. (B) Western blot analysis of BRCA2 with doxycycline induced knock-down and FIGNL1 levels upon CRISPR-Cas9 mediated knockout in RPE1 cells. (C) Left: Schematics of the results of knocking-out Fignl1 in Brca2 proficient (top) and deficient (bottom) mouse mammary tumor cells. Right: Genotyping results from Brca2 proficient (top) and deficient (bottom) mouse mammary tumor cells with 2 different clones of Fignl1 knockouts each. Electropherogram representative figure showing the alignment of Fignl1 genomic DNA region at the indicated binding site of sgRNA followed by its PAM sequence. (D) Western blot analysis of FIGNL1 levels from each clone upon CRISPR-Cas9 mediated knockout in mouse mammary tumor cells. (E) The percentage of cells in the different phases of the cell cycle profile of RPE1 and RPE1 FIGNL1 -KO cells. Error bars are indicative of mean±SD from 2 independent flow cytometry experiments. (F) The percentage of cells in the different phases of the cell cycle profile of mouse mammary tumor cells with Fignl1 -KO. Error bars are indicative of mean±SD from 2 independent flow cytometry experiments.

Article Snippet: Human FIGNL1 sgRNAs (table below) were cloned into pDG458 (Addgene #100900) plasmid at BbsI site on both side of Cas9 expression plasmid.

Techniques: Knock-Out, Clone Assay, Binding Assay, Sequencing, Western Blot, Knockdown, CRISPR, Flow Cytometry

(A) Top: Schematics showing the experimental setup for the high-content, automated RAD51 immunofluorescence (IF) in human RPE1 cells. 2µg/ml of doxycycline for 48 hours to induce the knock-down of BRCA2, followed by EdU labelling and induction of DSB via 10Gy of X-ray treatment. Left: Representative IF images showing EdU and RAD51 foci in untreated (NT) and ionizing radiation treated (IR) cells; scale bar: 20µm. Right: Automated Quantitative Image Based Cytometry (QIBC) plot of the RAD51 foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 2000 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (B) Top: Schematics showing the experimental setup for RAD51-IF in mouse mammary tumor cells expressing AsiSI with HA-tag. 24 hours of doxycycline treatment to induce AsiSI-HA expression with last 5 hours to translocate it to the nucleus by addition of 4-hydroxytamoxifen (4-OHT). Left: Representative IF images of the RAD51 and HA-tag in un-induced (NT) and induced (DOX &4 OHT) cells; scale bar: 20µm. Right: Automated QIBC plots of the RAD51 foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 1500 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (C) Top: The side and zoom-in view of the predicted FIGNL1-RAD51 complex. The FIGNL1 monomers are shown as surface representation (Chain A: Light blue; Chain B: Pale cyan; Chain C: Pale green; Chain D: Light yellow; Chain E: Light Orange; Chain F: Light pink) while RAD51 (beige) is shown in cartoon representation. Bottom: The zoom-in views represent the interacting regions of FIGNL1 and RAD51. Blue inset: the FIGNL1 FRBD (dark blue) interacts with a loop connecting the RAD51 Walker A/B motifs (green). Pink inset: two Pore Loops present in the FIGNL1 ATPase region (dark blue) interact with the RAD51 N-terminal domain (pink). (D) Top: Schematics of the DNA strand exchange assays. The assays were performed by adding 150nM purified RAD51 to the DNA substrates in the presence of increasing concentrations of purified FIGNL1-dNLS. The red asterisk indicates the position of the radioactive label. Bottom: Percentage normalized quantifications showing the band intensity of the exchanged DNA; error bars are indicative of mean±SEM from 3 independent experiments (n=3) P-values from Dunnett’s multiple comparison One-Way ANOVA comparing between indicated groups (*: <0.05; **: <0.01; ***: <0.001). (E) Top: Schematics of the DNA strand exchange assays. The assays were performed by adding 150nM purified RAD51 and 50nM RPA to the DNA substrates. The reactions were then supplemented with 30nM miBRCA2 and increasing concentrations of purified FIGNL1-dNLS. The red asterisk indicates the position of the radioactive label. Bottom: Percentage normalized quantifications showing the band intensity of the exchanged DNA; error bars are indicative of mean±SEM from 3 independent experiments (n=3). P-values from Dunnett’s multiple comparison One-Way ANOVA comparing between indicated groups (***: <0.001). (F) Top: Schematics of DR-GFP assay showing the strategy behind the experiment. Mutant SceGFP containing ISceI recognition site and stop codon is repaired by the downstream iGFP to restore the GFP fluorescence in the cells. Center: U2-OS cells with a stable integrated DR-GFP system were transfected for 72 hours with the indicated siRNA’s and underwent 48 hours of ISceI treatment to measure the levels of GFP fluorescence relative to the WT cells undergoing ISceI treatment. Error bars are representative of the mean±SD from 3 independent experiments (n=3). P-values from unpaired t-test comparing between indicated groups (**: >0.01). Bottom: Western blot analysis showing the levels of FIGNL1 and BRCA2 from the corresponding cells used for FACS analysis.

Journal: bioRxiv

Article Title: FIGNL1 inhibits homologous recombination in BRCA2 deficient cells by dissociating RAD51 filaments

doi: 10.1101/2024.11.03.621741

Figure Lengend Snippet: (A) Top: Schematics showing the experimental setup for the high-content, automated RAD51 immunofluorescence (IF) in human RPE1 cells. 2µg/ml of doxycycline for 48 hours to induce the knock-down of BRCA2, followed by EdU labelling and induction of DSB via 10Gy of X-ray treatment. Left: Representative IF images showing EdU and RAD51 foci in untreated (NT) and ionizing radiation treated (IR) cells; scale bar: 20µm. Right: Automated Quantitative Image Based Cytometry (QIBC) plot of the RAD51 foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 2000 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (B) Top: Schematics showing the experimental setup for RAD51-IF in mouse mammary tumor cells expressing AsiSI with HA-tag. 24 hours of doxycycline treatment to induce AsiSI-HA expression with last 5 hours to translocate it to the nucleus by addition of 4-hydroxytamoxifen (4-OHT). Left: Representative IF images of the RAD51 and HA-tag in un-induced (NT) and induced (DOX &4 OHT) cells; scale bar: 20µm. Right: Automated QIBC plots of the RAD51 foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 1500 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (C) Top: The side and zoom-in view of the predicted FIGNL1-RAD51 complex. The FIGNL1 monomers are shown as surface representation (Chain A: Light blue; Chain B: Pale cyan; Chain C: Pale green; Chain D: Light yellow; Chain E: Light Orange; Chain F: Light pink) while RAD51 (beige) is shown in cartoon representation. Bottom: The zoom-in views represent the interacting regions of FIGNL1 and RAD51. Blue inset: the FIGNL1 FRBD (dark blue) interacts with a loop connecting the RAD51 Walker A/B motifs (green). Pink inset: two Pore Loops present in the FIGNL1 ATPase region (dark blue) interact with the RAD51 N-terminal domain (pink). (D) Top: Schematics of the DNA strand exchange assays. The assays were performed by adding 150nM purified RAD51 to the DNA substrates in the presence of increasing concentrations of purified FIGNL1-dNLS. The red asterisk indicates the position of the radioactive label. Bottom: Percentage normalized quantifications showing the band intensity of the exchanged DNA; error bars are indicative of mean±SEM from 3 independent experiments (n=3) P-values from Dunnett’s multiple comparison One-Way ANOVA comparing between indicated groups (*: <0.05; **: <0.01; ***: <0.001). (E) Top: Schematics of the DNA strand exchange assays. The assays were performed by adding 150nM purified RAD51 and 50nM RPA to the DNA substrates. The reactions were then supplemented with 30nM miBRCA2 and increasing concentrations of purified FIGNL1-dNLS. The red asterisk indicates the position of the radioactive label. Bottom: Percentage normalized quantifications showing the band intensity of the exchanged DNA; error bars are indicative of mean±SEM from 3 independent experiments (n=3). P-values from Dunnett’s multiple comparison One-Way ANOVA comparing between indicated groups (***: <0.001). (F) Top: Schematics of DR-GFP assay showing the strategy behind the experiment. Mutant SceGFP containing ISceI recognition site and stop codon is repaired by the downstream iGFP to restore the GFP fluorescence in the cells. Center: U2-OS cells with a stable integrated DR-GFP system were transfected for 72 hours with the indicated siRNA’s and underwent 48 hours of ISceI treatment to measure the levels of GFP fluorescence relative to the WT cells undergoing ISceI treatment. Error bars are representative of the mean±SD from 3 independent experiments (n=3). P-values from unpaired t-test comparing between indicated groups (**: >0.01). Bottom: Western blot analysis showing the levels of FIGNL1 and BRCA2 from the corresponding cells used for FACS analysis.

Article Snippet: Human FIGNL1 sgRNAs (table below) were cloned into pDG458 (Addgene #100900) plasmid at BbsI site on both side of Cas9 expression plasmid.

Techniques: Immunofluorescence, Knockdown, Cytometry, MANN-WHITNEY, Two Tailed Test, Expressing, Purification, Comparison, Mutagenesis, Fluorescence, Transfection, Western Blot

(A) Left: representative images of high-content, automated RAD51-IF images showing the EdU and RAD51 foci in untreated and ionizing radiation treated mouse mammary tumor cells; scale bar: 20µm. Right: Automated QIBC plots of the RAD51 foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 1500 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (B) Left: representative images of 53BP1-IF images showing the 53BP1 foci in untreated and AsiSI induced mouse mammary tumor cells; scale bar: 20µm. Right: Automated QIBC plots of the 53BP1 foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 1500 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (C) Top Left: Schematics showing the experimental setup for RAD51-IF in U2-OS AsiSI (DIvA) cells. 48 hours of siRNA transfection to knock-down BRCA2 and FIGNL1 with last 5 hours to translocate constitutively expressing cytoplasmic AsiSI to the nucleus by addition of 4-hydroxytamoxifen (4-OHT). Top right: Western blot analysis showing the levels of BRCA2 and FIGNL1 upon siRNA mediated knock-down in DIvA cells. Bottom left: representative images of high-content, automated RAD51-IF images showing the RAD51 and γH2AX foci in untreated and AsiSI induced DIvA cells; scale bar: 5μm. Bottom right: Automated QIBC plots of the RAD51 foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 1500 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001).

Journal: bioRxiv

Article Title: FIGNL1 inhibits homologous recombination in BRCA2 deficient cells by dissociating RAD51 filaments

doi: 10.1101/2024.11.03.621741

Figure Lengend Snippet: (A) Left: representative images of high-content, automated RAD51-IF images showing the EdU and RAD51 foci in untreated and ionizing radiation treated mouse mammary tumor cells; scale bar: 20µm. Right: Automated QIBC plots of the RAD51 foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 1500 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (B) Left: representative images of 53BP1-IF images showing the 53BP1 foci in untreated and AsiSI induced mouse mammary tumor cells; scale bar: 20µm. Right: Automated QIBC plots of the 53BP1 foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 1500 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (C) Top Left: Schematics showing the experimental setup for RAD51-IF in U2-OS AsiSI (DIvA) cells. 48 hours of siRNA transfection to knock-down BRCA2 and FIGNL1 with last 5 hours to translocate constitutively expressing cytoplasmic AsiSI to the nucleus by addition of 4-hydroxytamoxifen (4-OHT). Top right: Western blot analysis showing the levels of BRCA2 and FIGNL1 upon siRNA mediated knock-down in DIvA cells. Bottom left: representative images of high-content, automated RAD51-IF images showing the RAD51 and γH2AX foci in untreated and AsiSI induced DIvA cells; scale bar: 5μm. Bottom right: Automated QIBC plots of the RAD51 foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 1500 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001).

Article Snippet: Human FIGNL1 sgRNAs (table below) were cloned into pDG458 (Addgene #100900) plasmid at BbsI site on both side of Cas9 expression plasmid.

Techniques: MANN-WHITNEY, Two Tailed Test, Transfection, Knockdown, Expressing, Western Blot

$(A) Recombinant FIGNL1-dNLS protein purified from baculovirus infected insect Sf9 cells. The purified protein was separated on a polyacrylamide gel and stained with Coomassie blue. (B) Representative images of electrophoretic mobility shift assay showing the concentration dependent binding to ssDNA (left) or dsDNA (middle); RPA was used as a control for ssDNA binding and yeast Ku as a control for dsDNA binding. The red asterisk indicates the position of the radioactive label. Right: quantification of binding efficiency with ssDNA and dsDNA in the presence or absence of Mg2+ and ATP; error bars are indicative of mean±SEM. (C) Recombinant RPA protein purified from baculovirus infected insect Sf9 cells. The purified protein was separated on a polyacrylamide gel and stained with Coomassie blue. R1: RPA1, R2: RPA2 and R3: RPA3. (D) Representative images of electrophoretic mobility shift assay showing the concentration dependent DNA binding efficiency of FIGNL1-dNLS in the presence of 8nM RPA; Left: low concentration, Right: high concentration of FIGNL1-dNLS. The red asterisk indicates the position of the radioactive label. (E) Recombinant RAD51 protein purified from BL21-(DE3)pLysS E. coli cells. The purified protein was separated on a polyacrylamide gel and stained with Coomassie blue. (F) Top: Schematics of the in vitro interaction assay between FIGNL1-dNLS (immobilized) and RAD51. Bottom: Western blot results indicative of the interaction between FIGNL1 and RAD51 in the in vitro interaction assay. (G) Top: Schematics of the GFPtrap pull-down of the EGFP-FIGNL1 and RAD51 from nuclear extracts. EGFP-FIGNL1 stably over-expressing cells in FIGNL1 -/- RPE1 cells were used for nuclear fractionation and GFPtrap pulled down. Bottom: Western blot results indicative of the interaction between FIGNL1 and RAD51. (H) The side and top view of the FIGNL1 hexamer. The monomers are shown as surface representation (Chain A: Light blue; Chain B: Pale cyan; Chain C: Pale green; Chain D: Light yellow; Chain E: Light Orange; Chain F: Light pink). (I) The top and side view of FIGNL1 showing the interacting domains participating in formation of the hexamer (N-terminal: Blue, ATPase: Dark green, and C-terminal: maroon). (J) Top: Quantification of ssDNA degradation was done by normalizing the ssDNA substrate band intensity against the band intensity of the no-protein product; error bars are indicative of mean±SEM from 4 independent experiments (n=4). P-values from Ordinary One-Way ANOVA comparing between indicated groups (**: <0.01; ****: <0.0001). The red asterisk indicates the position of the radioactive label. Bottom: representative gel picture for ssDNA protection assay showing the protection efficiency of RAD51 against degradation by DNA2 nuclease in the presence of increasing concentrations of FIGNL1-dNLS. (K) Top: Schematics of miBRCA2 constructed with BRC repeats BRC1 & 2, DNA binding domain (DBD) and C-terminal domain (C-ter) compared to the full-length BRCA2. Recombinant miBRCA2 protein purified from baculovirus infected insect Sf9 cells. The purified protein was separated on a polyacrylamide gel and stained with Coomassie blue.$

Journal: bioRxiv

Article Title: FIGNL1 inhibits homologous recombination in BRCA2 deficient cells by dissociating RAD51 filaments

doi: 10.1101/2024.11.03.621741

Figure Lengend Snippet: (A) Recombinant FIGNL1-dNLS protein purified from baculovirus infected insect Sf9 cells. The purified protein was separated on a polyacrylamide gel and stained with Coomassie blue. (B) Representative images of electrophoretic mobility shift assay showing the concentration dependent binding to ssDNA (left) or dsDNA (middle); RPA was used as a control for ssDNA binding and yeast Ku as a control for dsDNA binding. The red asterisk indicates the position of the radioactive label. Right: quantification of binding efficiency with ssDNA and dsDNA in the presence or absence of Mg2+ and ATP; error bars are indicative of mean±SEM. (C) Recombinant RPA protein purified from baculovirus infected insect Sf9 cells. The purified protein was separated on a polyacrylamide gel and stained with Coomassie blue. R1: RPA1, R2: RPA2 and R3: RPA3. (D) Representative images of electrophoretic mobility shift assay showing the concentration dependent DNA binding efficiency of FIGNL1-dNLS in the presence of 8nM RPA; Left: low concentration, Right: high concentration of FIGNL1-dNLS. The red asterisk indicates the position of the radioactive label. (E) Recombinant RAD51 protein purified from BL21-(DE3)pLysS E. coli cells. The purified protein was separated on a polyacrylamide gel and stained with Coomassie blue. (F) Top: Schematics of the in vitro interaction assay between FIGNL1-dNLS (immobilized) and RAD51. Bottom: Western blot results indicative of the interaction between FIGNL1 and RAD51 in the in vitro interaction assay. (G) Top: Schematics of the GFPtrap pull-down of the EGFP-FIGNL1 and RAD51 from nuclear extracts. EGFP-FIGNL1 stably over-expressing cells in FIGNL1 -/- RPE1 cells were used for nuclear fractionation and GFPtrap pulled down. Bottom: Western blot results indicative of the interaction between FIGNL1 and RAD51. (H) The side and top view of the FIGNL1 hexamer. The monomers are shown as surface representation (Chain A: Light blue; Chain B: Pale cyan; Chain C: Pale green; Chain D: Light yellow; Chain E: Light Orange; Chain F: Light pink). (I) The top and side view of FIGNL1 showing the interacting domains participating in formation of the hexamer (N-terminal: Blue, ATPase: Dark green, and C-terminal: maroon). (J) Top: Quantification of ssDNA degradation was done by normalizing the ssDNA substrate band intensity against the band intensity of the no-protein product; error bars are indicative of mean±SEM from 4 independent experiments (n=4). P-values from Ordinary One-Way ANOVA comparing between indicated groups (**: <0.01; ****: <0.0001). The red asterisk indicates the position of the radioactive label. Bottom: representative gel picture for ssDNA protection assay showing the protection efficiency of RAD51 against degradation by DNA2 nuclease in the presence of increasing concentrations of FIGNL1-dNLS. (K) Top: Schematics of miBRCA2 constructed with BRC repeats BRC1 & 2, DNA binding domain (DBD) and C-terminal domain (C-ter) compared to the full-length BRCA2. Recombinant miBRCA2 protein purified from baculovirus infected insect Sf9 cells. The purified protein was separated on a polyacrylamide gel and stained with Coomassie blue.

Article Snippet: Human FIGNL1 sgRNAs (table below) were cloned into pDG458 (Addgene #100900) plasmid at BbsI site on both side of Cas9 expression plasmid.

Techniques: Recombinant, Purification, Infection, Staining, Electrophoretic Mobility Shift Assay, Concentration Assay, Binding Assay, Control, In Vitro, Western Blot, Stable Transfection, Expressing, Fractionation, Construct

Journal: bioRxiv

Article Title: FIGNL1 inhibits homologous recombination in BRCA2 deficient cells by dissociating RAD51 filaments

doi: 10.1101/2024.11.03.621741

Figure Lengend Snippet:

Article Snippet: Human FIGNL1 sgRNAs (table below) were cloned into pDG458 (Addgene #100900) plasmid at BbsI site on both side of Cas9 expression plasmid.

Techniques:

(A) Schematic representation of strategy behind deletion of BRCA2 in mouse embryonic stem cells. (B) Left: representative southern blot images showing the 4.8kb conditional knock-out allele (upper band) and 2.2kb deleted allele (lower band) of BRCA2. Stars in the shFIGNL1 clones represents the viable clones in the absence of BRCA2. Right: Percentage quantification of rescued clones. (C) Left: representative RAD51-IF images in mouse embryonic stem cells (scale bar: 20μm). Brca2 R2336H a Brca2 cell line that has a hypomorphic allele (p.arg2336His) of BRCA2 that is defective in HR. Right: Semi-automated QIBC plots of the RAD51 foci per nucleus in WT and FIGNL1 knock down clones; scatter box plot showing the foci distribution between 10-90th percentile (n=150). The experiment was performed two times. P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (D) Representative metaphase spread image showing different aberrations analyzed for genomic instability. Scale bar (whole metaphase): 5 µm; scale bar (individual aberrations): 1 µm. (E) Quantification of chromosomal aberrations in mouse mammary tumor cells in response to Olaparib and cisplatin from 3 independent experiments in which 50 individual metaphases were analyzed for each sample per experiment. P values from Ordinary two-way ANOVA (****: <0.0001) (F) Percentage survivability of mouse mammary tumor cells in response to Olaparib; quantification was performed by normalization to non-treated cells. P-values from Dunnett’s multiple comparison One-Way ANOVA comparing between indicated groups (*: <0.05, **: <0.01). (G) Percentage survivability of mouse mammary tumor cells in response to cisplatin; quantification was performed by normalization to non-treated cells. P-values from Dunnett’s multiple comparison One-Way ANOVA comparing between indicated groups (*: <0.05, **: <0.01).

Journal: bioRxiv

Article Title: FIGNL1 inhibits homologous recombination in BRCA2 deficient cells by dissociating RAD51 filaments

doi: 10.1101/2024.11.03.621741

Figure Lengend Snippet: (A) Schematic representation of strategy behind deletion of BRCA2 in mouse embryonic stem cells. (B) Left: representative southern blot images showing the 4.8kb conditional knock-out allele (upper band) and 2.2kb deleted allele (lower band) of BRCA2. Stars in the shFIGNL1 clones represents the viable clones in the absence of BRCA2. Right: Percentage quantification of rescued clones. (C) Left: representative RAD51-IF images in mouse embryonic stem cells (scale bar: 20μm). Brca2 R2336H a Brca2 cell line that has a hypomorphic allele (p.arg2336His) of BRCA2 that is defective in HR. Right: Semi-automated QIBC plots of the RAD51 foci per nucleus in WT and FIGNL1 knock down clones; scatter box plot showing the foci distribution between 10-90th percentile (n=150). The experiment was performed two times. P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (D) Representative metaphase spread image showing different aberrations analyzed for genomic instability. Scale bar (whole metaphase): 5 µm; scale bar (individual aberrations): 1 µm. (E) Quantification of chromosomal aberrations in mouse mammary tumor cells in response to Olaparib and cisplatin from 3 independent experiments in which 50 individual metaphases were analyzed for each sample per experiment. P values from Ordinary two-way ANOVA (****: <0.0001) (F) Percentage survivability of mouse mammary tumor cells in response to Olaparib; quantification was performed by normalization to non-treated cells. P-values from Dunnett’s multiple comparison One-Way ANOVA comparing between indicated groups (*: <0.05, **: <0.01). (G) Percentage survivability of mouse mammary tumor cells in response to cisplatin; quantification was performed by normalization to non-treated cells. P-values from Dunnett’s multiple comparison One-Way ANOVA comparing between indicated groups (*: <0.05, **: <0.01).

Article Snippet: Human FIGNL1 sgRNAs (table below) were cloned into pDG458 (Addgene #100900) plasmid at BbsI site on both side of Cas9 expression plasmid.

Techniques: Southern Blot, Knock-Out, Clone Assay, Knockdown, MANN-WHITNEY, Two Tailed Test, Comparison

(A) Quantification of chromosomal aberrations in FIGNL1 -/- RPE1 clones in the presence or absence of BRCA2 in response to Olaparib and cisplatin from 3 independent experiments in which 50 individual metaphases were analyzed for each sample per experiment. P values from Ordinary two-way ANOVA (****: <0.0001) (B) Quantification of chromosomal aberrations in mouse embryonic stem cells with FIGNL1 knock down clones in the presence or absence of Brca2 in response to Olaparib from 3 independent experiments in which 50 individual metaphases were analyzed for each sample per experiment. P values from Ordinary two-way ANOVA (****: <0.0001) (C) Representative clonogenic images of mouse mammary tumor cells in response to Olaparib. (D) Representative clonogenic images of mouse mammary tumor cells in response to cisplatin. (E) Percentage survivability of mouse embryonic stem cells in response to Olaparib; quantification was performed by normalization to non-treated cells. P-values from Dunnett’s multiple comparison One-way ANOVA comparing between indicated groups (*: <0.05, ***: <0.001). (F) Percentage survivability of mouse embryonic stem cells in response to cisplatin; quantification was performed by normalization to non-treated cells. P-values from Dunnett’s multiple comparison One-way ANOVA comparing between indicated groups (*: <0.05, **: <0.01).

Journal: bioRxiv

Article Title: FIGNL1 inhibits homologous recombination in BRCA2 deficient cells by dissociating RAD51 filaments

doi: 10.1101/2024.11.03.621741

Figure Lengend Snippet: (A) Quantification of chromosomal aberrations in FIGNL1 -/- RPE1 clones in the presence or absence of BRCA2 in response to Olaparib and cisplatin from 3 independent experiments in which 50 individual metaphases were analyzed for each sample per experiment. P values from Ordinary two-way ANOVA (****: <0.0001) (B) Quantification of chromosomal aberrations in mouse embryonic stem cells with FIGNL1 knock down clones in the presence or absence of Brca2 in response to Olaparib from 3 independent experiments in which 50 individual metaphases were analyzed for each sample per experiment. P values from Ordinary two-way ANOVA (****: <0.0001) (C) Representative clonogenic images of mouse mammary tumor cells in response to Olaparib. (D) Representative clonogenic images of mouse mammary tumor cells in response to cisplatin. (E) Percentage survivability of mouse embryonic stem cells in response to Olaparib; quantification was performed by normalization to non-treated cells. P-values from Dunnett’s multiple comparison One-way ANOVA comparing between indicated groups (*: <0.05, ***: <0.001). (F) Percentage survivability of mouse embryonic stem cells in response to cisplatin; quantification was performed by normalization to non-treated cells. P-values from Dunnett’s multiple comparison One-way ANOVA comparing between indicated groups (*: <0.05, **: <0.01).

Article Snippet: Human FIGNL1 sgRNAs (table below) were cloned into pDG458 (Addgene #100900) plasmid at BbsI site on both side of Cas9 expression plasmid.

Techniques: Clone Assay, Knockdown, Comparison

(A) Schematics of the FIGNL1 mutant proteins (B) Western blot analysis of FIGNL1 mutants fusion with GFP protein. (C) Quantification of chromosomal aberrations in FIGNL1 -/- RPE1 cells reconstituted with FIGNL1 mutants in response to Olaparib and cisplatin from 3 independent experiments in which 50 individual metaphases were analyzed for each sample per experiment. P values from Ordinary two-way ANOVA (ns-nonsignificant; *: <0.05; **: <0.01).

Journal: bioRxiv

Article Title: FIGNL1 inhibits homologous recombination in BRCA2 deficient cells by dissociating RAD51 filaments

doi: 10.1101/2024.11.03.621741

Figure Lengend Snippet: (A) Schematics of the FIGNL1 mutant proteins (B) Western blot analysis of FIGNL1 mutants fusion with GFP protein. (C) Quantification of chromosomal aberrations in FIGNL1 -/- RPE1 cells reconstituted with FIGNL1 mutants in response to Olaparib and cisplatin from 3 independent experiments in which 50 individual metaphases were analyzed for each sample per experiment. P values from Ordinary two-way ANOVA (ns-nonsignificant; *: <0.05; **: <0.01).

Article Snippet: Human FIGNL1 sgRNAs (table below) were cloned into pDG458 (Addgene #100900) plasmid at BbsI site on both side of Cas9 expression plasmid.

Techniques: Mutagenesis, Western Blot

$(A) Top: representative images of high-content, automated RAD51-IF images showing the EdU and RAD51 foci in untreated and ionizing radiation treated human RPE1 cells; scale bar: 5µm. Bottom: Automated QIBC plots of the RAD51 foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 1500 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (B) Left: The side and top view of FIGNL1-MMS22L-TONSL complex. The FIGNL1 hexamer is shown as surface representation (Light blue) while MMS22L (teal) and TONSL (light pink) are shown in cartoon representation. Right: Surface representation of ternary complex showing the major binding interface of ternary complex. MMS22L-TONSL interacts through its heterodimeric interface (maroon) with FIGNL1 FRBD and NLS-FRBD connecting loop (dark blue). (C) Top: Schematics of the in vitro interaction assay between recombinant FIGNL1-dNLS (immobilized) and MMS22L-TONSL. Bottom: Western blot results show that FIGNL1-dNLS interacts with MMS22L-TONSL in the in vitro interaction assay. (D) Top: Schematics of the GFPtrap pull-down of the EGFP-FIGNL1 and MMS22L-TONSL from nuclear extracts. EGFP-FIGNL1 stably over-expressing cells in FIGNL1 -/- RPE1 cells were used for nuclear fractionation and GFPtrap pulled down. Bottom: Western blot results indicative of the interaction between FIGNL1 and MMS22L-TONSL. (E) (Top) The side and top of the FIGNL1-RAD51-MMS22L-TONSL complex. The FIGNL1 hexamer, (Light blue), MMS22L (teal), and TONSL (light pink) are shown as surface representation, while RAD51 (beige) is shown in cartoon representation. (Middle) Surface representation and zoom in view of ternary complex showing the binding interfaces of FIGNL1− FRBD and pore loops are represented in dark blue; while RAD51 interacting domains, N-terminal and loop connecting Walker A/B motifs are represented in pink and green respectively. The non-interacting MMS22L-TONSL is marked in grey. (Bottom) Side and top view of the overlay of FIGNL1-MMS22L-TONSL in ternary (raspberry/tMMS22L-TONSL/without RAD51) and quaternary (grey/qMMS22L-TONSL/with RAD51) complexes. The outward movement of the MMS22L-TONSL heterodimeric interface away from the FIGNL1 pocket (shown by black arrow) represents the loss of FIGNL1-MMS22L-TONSL binding upon FIGNL1-RAD51 interaction. (F) Top: Schematics of the in vitro interaction assay between recombinant FIGNL1-dNLS (immobilized), MMS22L-TONSL and RAD51-K133R in the presence or absence of ssDNA. Bottom: Percentage normalized quantifications showing the band intensity of the bound MMS22L-TONSL to FIGNL1-dNLS; error bars are indicative of mean±SEM from 4 independent experiments (n=4). P-values from Ordinary One-Way ANOVA comparing between indicated groups (****: <0.0001). Western blot results indicate the interactions between the proteins involved. (G) Top: Schematics of the GFPtrap pull-down of the EGFP-FIGNL1 and MMS22L-TONSL in the presence or absence of BRCA2 (+DOX), from nuclear extracts. EGFP-FIGNL1 stably over-expressing cells in FIGNL1 -/- RPE1 cells were used for nuclear fractionation and GFPtrap pulled down. Bottom: Western blot results indicative of the interaction between FIGNL1 and MMS22L-TONSL.$

Journal: bioRxiv

Article Title: FIGNL1 inhibits homologous recombination in BRCA2 deficient cells by dissociating RAD51 filaments

doi: 10.1101/2024.11.03.621741

Figure Lengend Snippet: (A) Top: representative images of high-content, automated RAD51-IF images showing the EdU and RAD51 foci in untreated and ionizing radiation treated human RPE1 cells; scale bar: 5µm. Bottom: Automated QIBC plots of the RAD51 foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 1500 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (B) Left: The side and top view of FIGNL1-MMS22L-TONSL complex. The FIGNL1 hexamer is shown as surface representation (Light blue) while MMS22L (teal) and TONSL (light pink) are shown in cartoon representation. Right: Surface representation of ternary complex showing the major binding interface of ternary complex. MMS22L-TONSL interacts through its heterodimeric interface (maroon) with FIGNL1 FRBD and NLS-FRBD connecting loop (dark blue). (C) Top: Schematics of the in vitro interaction assay between recombinant FIGNL1-dNLS (immobilized) and MMS22L-TONSL. Bottom: Western blot results show that FIGNL1-dNLS interacts with MMS22L-TONSL in the in vitro interaction assay. (D) Top: Schematics of the GFPtrap pull-down of the EGFP-FIGNL1 and MMS22L-TONSL from nuclear extracts. EGFP-FIGNL1 stably over-expressing cells in FIGNL1 -/- RPE1 cells were used for nuclear fractionation and GFPtrap pulled down. Bottom: Western blot results indicative of the interaction between FIGNL1 and MMS22L-TONSL. (E) (Top) The side and top of the FIGNL1-RAD51-MMS22L-TONSL complex. The FIGNL1 hexamer, (Light blue), MMS22L (teal), and TONSL (light pink) are shown as surface representation, while RAD51 (beige) is shown in cartoon representation. (Middle) Surface representation and zoom in view of ternary complex showing the binding interfaces of FIGNL1− FRBD and pore loops are represented in dark blue; while RAD51 interacting domains, N-terminal and loop connecting Walker A/B motifs are represented in pink and green respectively. The non-interacting MMS22L-TONSL is marked in grey. (Bottom) Side and top view of the overlay of FIGNL1-MMS22L-TONSL in ternary (raspberry/tMMS22L-TONSL/without RAD51) and quaternary (grey/qMMS22L-TONSL/with RAD51) complexes. The outward movement of the MMS22L-TONSL heterodimeric interface away from the FIGNL1 pocket (shown by black arrow) represents the loss of FIGNL1-MMS22L-TONSL binding upon FIGNL1-RAD51 interaction. (F) Top: Schematics of the in vitro interaction assay between recombinant FIGNL1-dNLS (immobilized), MMS22L-TONSL and RAD51-K133R in the presence or absence of ssDNA. Bottom: Percentage normalized quantifications showing the band intensity of the bound MMS22L-TONSL to FIGNL1-dNLS; error bars are indicative of mean±SEM from 4 independent experiments (n=4). P-values from Ordinary One-Way ANOVA comparing between indicated groups (****: <0.0001). Western blot results indicate the interactions between the proteins involved. (G) Top: Schematics of the GFPtrap pull-down of the EGFP-FIGNL1 and MMS22L-TONSL in the presence or absence of BRCA2 (+DOX), from nuclear extracts. EGFP-FIGNL1 stably over-expressing cells in FIGNL1 -/- RPE1 cells were used for nuclear fractionation and GFPtrap pulled down. Bottom: Western blot results indicative of the interaction between FIGNL1 and MMS22L-TONSL.

Article Snippet: Human FIGNL1 sgRNAs (table below) were cloned into pDG458 (Addgene #100900) plasmid at BbsI site on both side of Cas9 expression plasmid.

Techniques: MANN-WHITNEY, Two Tailed Test, Binding Assay, In Vitro, Recombinant, Western Blot, Stable Transfection, Expressing, Fractionation

(A) Predictome analysis highlighting the FIGNL1 interacting partners involved in genome maintenance. (B) Recombinant MMS22L and TONSL heterodimer purified from baculovirus infected insect Sf9 cells. Purified proteins were separated on a polyacrylamide gel and stained with Coomassie blue. (C) Recombinant RAD51-K133R protein purified from BL21-(DE3)pLysS strain in E. coli cells. Purified proteins were separated on a polyacrylamide gel and stained with Coomassie blue.

Journal: bioRxiv

Article Title: FIGNL1 inhibits homologous recombination in BRCA2 deficient cells by dissociating RAD51 filaments

doi: 10.1101/2024.11.03.621741

Figure Lengend Snippet: (A) Predictome analysis highlighting the FIGNL1 interacting partners involved in genome maintenance. (B) Recombinant MMS22L and TONSL heterodimer purified from baculovirus infected insect Sf9 cells. Purified proteins were separated on a polyacrylamide gel and stained with Coomassie blue. (C) Recombinant RAD51-K133R protein purified from BL21-(DE3)pLysS strain in E. coli cells. Purified proteins were separated on a polyacrylamide gel and stained with Coomassie blue.

Article Snippet: Human FIGNL1 sgRNAs (table below) were cloned into pDG458 (Addgene #100900) plasmid at BbsI site on both side of Cas9 expression plasmid.

Techniques: Recombinant, Purification, Infection, Staining

(A) Left: Representative high-content, automated IF images showing EdU and MMS22L foci in untreated and ionizing radiation treated cells; scale bar: 5µm. Right: Automated QIBC plots of the MMS22L foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 2000 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (B) Top: Schematics of the DNA strand exchange assay. The strand exchange assays were performed by adding 400nM purified RAD51 and 75nM MMS22L-TONSL to the DNA substrate. The assay was then supplemented by increasing concentrations of purified FIGNL1-dNLS. The red asterisk indicates the position of the radioactive label. Bottom: Percentage normalized quantifications showing the band intensity of the exchanged DNA; error bars are indicative of mean±SEM from 3 independent experiments (n=3). P-values from ordinary One-way ANOVA comparing between indicated groups (*: <0.05). (C) Top: Representative high-content, automated IF images showing EdU and RAD51 foci in untreated and ionizing radiation treated human RPE1 cells; scale bar: 5µm. Bottom: Automated QIBC plots of the RAD51 foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 1500 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (D) DR-GFP system stably integrated U2-OS cells transfected with 72 hours of indicated siRNAs overlapping with 48 hours of ISceI to measure the relative levels of GFP fluorescence to the WT cells. Error bars are representative of the mean±SD from 3 independent experiments (n=3). P-values from unpaired t-test comparing between indicated groups (****: <0.0001). (E) Quantification of chromosomal aberrations in RPE1 FIGNL1 knock-out clones with siRNAs against MMS22L and TONSL in response to Olaparib and cisplatin from 3 independent experiments in which 50 individual metaphases were analyzed for each sample per experiment. P values from Ordinary two-way ANOVA (*: <0.05; **: <0.01; ****: <0.0001). (F) Percentage survivability of mouse mammary tumor cells in response to Olaparib; quantification was performed by normalization to non-treated cells. P-values from Dunnett’s multiple comparison One-Way ANOVA comparing between indicated groups (ns: 0.1234; *: <0.05, **: <0.01).

Journal: bioRxiv

Article Title: FIGNL1 inhibits homologous recombination in BRCA2 deficient cells by dissociating RAD51 filaments

doi: 10.1101/2024.11.03.621741

Figure Lengend Snippet: (A) Left: Representative high-content, automated IF images showing EdU and MMS22L foci in untreated and ionizing radiation treated cells; scale bar: 5µm. Right: Automated QIBC plots of the MMS22L foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 2000 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (B) Top: Schematics of the DNA strand exchange assay. The strand exchange assays were performed by adding 400nM purified RAD51 and 75nM MMS22L-TONSL to the DNA substrate. The assay was then supplemented by increasing concentrations of purified FIGNL1-dNLS. The red asterisk indicates the position of the radioactive label. Bottom: Percentage normalized quantifications showing the band intensity of the exchanged DNA; error bars are indicative of mean±SEM from 3 independent experiments (n=3). P-values from ordinary One-way ANOVA comparing between indicated groups (*: <0.05). (C) Top: Representative high-content, automated IF images showing EdU and RAD51 foci in untreated and ionizing radiation treated human RPE1 cells; scale bar: 5µm. Bottom: Automated QIBC plots of the RAD51 foci per nucleus; scatter box plot showing the foci distribution between 10-90th percentile from 1500 cells (experiment was repeated 3 times). P-values from Mann-Whitney test (two-tailed) comparing between indicated groups (****: <0.0001). (D) DR-GFP system stably integrated U2-OS cells transfected with 72 hours of indicated siRNAs overlapping with 48 hours of ISceI to measure the relative levels of GFP fluorescence to the WT cells. Error bars are representative of the mean±SD from 3 independent experiments (n=3). P-values from unpaired t-test comparing between indicated groups (****: <0.0001). (E) Quantification of chromosomal aberrations in RPE1 FIGNL1 knock-out clones with siRNAs against MMS22L and TONSL in response to Olaparib and cisplatin from 3 independent experiments in which 50 individual metaphases were analyzed for each sample per experiment. P values from Ordinary two-way ANOVA (*: <0.05; **: <0.01; ****: <0.0001). (F) Percentage survivability of mouse mammary tumor cells in response to Olaparib; quantification was performed by normalization to non-treated cells. P-values from Dunnett’s multiple comparison One-Way ANOVA comparing between indicated groups (ns: 0.1234; *: <0.05, **: <0.01).

Article Snippet: Human FIGNL1 sgRNAs (table below) were cloned into pDG458 (Addgene #100900) plasmid at BbsI site on both side of Cas9 expression plasmid.

Techniques: MANN-WHITNEY, Two Tailed Test, Purification, Stable Transfection, Transfection, Fluorescence, Knock-Out, Clone Assay, Comparison

Journal: bioRxiv

Article Title: FIGNL1 inhibits homologous recombination in BRCA2 deficient cells by dissociating RAD51 filaments

doi: 10.1101/2024.11.03.621741

Article Snippet: Human FIGNL1 sgRNAs (table below) were cloned into pDG458 (Addgene #100900) plasmid at BbsI site on both side of Cas9 expression plasmid.

Techniques: MANN-WHITNEY, Two Tailed Test, Western Blot

(A) Upon induction of DSBs, the break ends are resected in the 5’to 3’ direction, exposing a 3’ssDNA overhang. (B) BRCA2 plays a key role by loading and stabilizing RAD51 on the resected single-stranded ssDNA, facilitating RAD51-mediated strand invasion. Besides facilitating RAD51 loading, BRCA2 is crucial for maintaining RAD51 filament stability by preventing its premature dissociation caused by the anti-recombinase FIGNL1. In the presence of BRCA2, the limited availability of free RAD51 allows the hexameric FIGNL1 to interact with the MMS22L-TONSL complex. This interaction ensures that RAD51 levels are finely regulated during HR in wild-type cells. (C) However, in the absence of BRCA2, the elevated concentration of free RAD51 results in the dissociation of the FIGNL1-MMS22L-TONSL complex. This could lead to in dynamic cycles of RAD51 loading and dissociation between the MMS22L-TONSL complex and FIGNL1, ultimately causing RAD51 loading defect and increased genome instability. (D) The loss of FIGNL1 in BRCA2-deficient cells restores productive RAD51 loading by the MMS22L-TONSL complex, rescuing HR efficiency, which in turn restores the genome instability observed upon loss of BRCA2.

Journal: bioRxiv

Article Title: FIGNL1 inhibits homologous recombination in BRCA2 deficient cells by dissociating RAD51 filaments

doi: 10.1101/2024.11.03.621741

Figure Lengend Snippet: (A) Upon induction of DSBs, the break ends are resected in the 5’to 3’ direction, exposing a 3’ssDNA overhang. (B) BRCA2 plays a key role by loading and stabilizing RAD51 on the resected single-stranded ssDNA, facilitating RAD51-mediated strand invasion. Besides facilitating RAD51 loading, BRCA2 is crucial for maintaining RAD51 filament stability by preventing its premature dissociation caused by the anti-recombinase FIGNL1. In the presence of BRCA2, the limited availability of free RAD51 allows the hexameric FIGNL1 to interact with the MMS22L-TONSL complex. This interaction ensures that RAD51 levels are finely regulated during HR in wild-type cells. (C) However, in the absence of BRCA2, the elevated concentration of free RAD51 results in the dissociation of the FIGNL1-MMS22L-TONSL complex. This could lead to in dynamic cycles of RAD51 loading and dissociation between the MMS22L-TONSL complex and FIGNL1, ultimately causing RAD51 loading defect and increased genome instability. (D) The loss of FIGNL1 in BRCA2-deficient cells restores productive RAD51 loading by the MMS22L-TONSL complex, rescuing HR efficiency, which in turn restores the genome instability observed upon loss of BRCA2.

Article Snippet: Human FIGNL1 sgRNAs (table below) were cloned into pDG458 (Addgene #100900) plasmid at BbsI site on both side of Cas9 expression plasmid.

Techniques: Concentration Assay

High FIGNL1 expression in human CRC samples. (a) The expression of FIGNL1 in the TCGA database. (b) Immunohistochemical detection of FIGNL1 expression in clinical CRC samples. FIGNL1, fidgetin-like 1; CRC, colorectal cancer.

Journal: Balkan Medical Journal

Article Title: Exploring the Effect of Fidgetin-Like 1 on Colorectal Cancer Through Tissue Chip and In Vitro Experiments

doi: 10.4274/balkanmedj.galenos.2024.2024-7-9

Figure Lengend Snippet: High FIGNL1 expression in human CRC samples. (a) The expression of FIGNL1 in the TCGA database. (b) Immunohistochemical detection of FIGNL1 expression in clinical CRC samples. FIGNL1, fidgetin-like 1; CRC, colorectal cancer.

Article Snippet: Details of primer antibodies used in this study are as follows: FIGNL1 (17604-1-AP, Proteintech, Wuhan, China), GADPH (60004-1-Ig, Proteintech, Wuhan, China), p-JNK (80024-1-AP, Proteintech, Wuhan, China), T-JNK (24164-1-AP, Proteintech, Wuhan, China), p-ERK (28733-1-AP, Proteintech, Wuhan, China), T-ERK (11257-1-AP, Proteintech, Wuhan, China), p-P38 (28796-1-AP, Proteintech, Wuhan, China), T-P38 (14064-1-AP, Proteintech, Wuhan, China), SPIDR (HPA041582, Sigma-Aldrich, Missouri, USA).

Techniques: Expressing, Immunohistochemical staining

FIGNL1 knockdown inhibited Caco-2 cell proliferation, migration, and invasion. (a) Representative blot demonstrating FIGNL1 expression in NCM460, Colo-205, and Caco-2 cells. (b) Bar chart illustrating FIGNL1 mRNA expression following FIGNL1 siRNA treatment in Caco-2 cells. (c) Representative blot depicting FIGNL1 expression following FIGNL1 siRNA treatment in Caco-2 cells. (d) Line chart exhibiting Caco-2 cell proliferation with FIGNL1 siRNA treatment. (e) Cell migration and invasion were examined using transwell assay after FIGNL1 siRNA transfection in Caco-2 cells. Scale bar: 500 µm. * p < 0.05 and ** p < 0.01 versus control siRNA group. Con, control; OD, optical density; FIGNL1, fidgetin-like 1; CRC, colorectal cancer; siRNA, Small interfering RNA.

Journal: Balkan Medical Journal

Article Title: Exploring the Effect of Fidgetin-Like 1 on Colorectal Cancer Through Tissue Chip and In Vitro Experiments

doi: 10.4274/balkanmedj.galenos.2024.2024-7-9

Figure Lengend Snippet: FIGNL1 knockdown inhibited Caco-2 cell proliferation, migration, and invasion. (a) Representative blot demonstrating FIGNL1 expression in NCM460, Colo-205, and Caco-2 cells. (b) Bar chart illustrating FIGNL1 mRNA expression following FIGNL1 siRNA treatment in Caco-2 cells. (c) Representative blot depicting FIGNL1 expression following FIGNL1 siRNA treatment in Caco-2 cells. (d) Line chart exhibiting Caco-2 cell proliferation with FIGNL1 siRNA treatment. (e) Cell migration and invasion were examined using transwell assay after FIGNL1 siRNA transfection in Caco-2 cells. Scale bar: 500 µm. * p < 0.05 and ** p < 0.01 versus control siRNA group. Con, control; OD, optical density; FIGNL1, fidgetin-like 1; CRC, colorectal cancer; siRNA, Small interfering RNA.

Techniques: Knockdown, Migration, Expressing, Transwell Assay, Transfection, Control, Small Interfering RNA

FIGNL1 overexpression promoted Colo-205 cell proliferation, migration, and invasion. (a) Bar chart displaying FIGNL1 mRNA expression following FIGNL1 overexpression lentivirus infection in Colo-205 cells. (b) Representative blot depicting FIGNL1 expression after FIGNL1 overexpression-inducing lentivirus infection in Colo-205 cells. (c) Line chart exhibiting the Colo-205 cell proliefration after FIGNL1 overexpression-inducing lentivirus infection. (d) Cell migration and invasion were assessed using transwell assay after FIGNL1 overexpression-inducing lentivirus infection in Colo-205 cells. Scale bar: 200 µm. * p < 0.05 and ** p < 0.01 versus control overexpression group. FIGNL1, fidgetin-like 1; Con, control; OD, optical density; OE, overexpression.

Journal: Balkan Medical Journal

Article Title: Exploring the Effect of Fidgetin-Like 1 on Colorectal Cancer Through Tissue Chip and In Vitro Experiments

doi: 10.4274/balkanmedj.galenos.2024.2024-7-9

Figure Lengend Snippet: FIGNL1 overexpression promoted Colo-205 cell proliferation, migration, and invasion. (a) Bar chart displaying FIGNL1 mRNA expression following FIGNL1 overexpression lentivirus infection in Colo-205 cells. (b) Representative blot depicting FIGNL1 expression after FIGNL1 overexpression-inducing lentivirus infection in Colo-205 cells. (c) Line chart exhibiting the Colo-205 cell proliefration after FIGNL1 overexpression-inducing lentivirus infection. (d) Cell migration and invasion were assessed using transwell assay after FIGNL1 overexpression-inducing lentivirus infection in Colo-205 cells. Scale bar: 200 µm. * p < 0.05 and ** p < 0.01 versus control overexpression group. FIGNL1, fidgetin-like 1; Con, control; OD, optical density; OE, overexpression.

Techniques: Over Expression, Migration, Expressing, Infection, Transwell Assay, Control

FIGNL1 promoted CRC progression by activating the P38 pathway in vitro. (a) Representative blots demonstrating ERK/JNK/P38 MAPK expression following FIGNL1 siRNA treatment in Caco-2 cells. (b) Representative blots depicting ERK/JNK/P38 MAPK expression with lentivirus infection-related FIGNL1 overexpression in Colo-205 cells. (c) Line chart illustrating the Colo-205 cell proliferation following sb203580 treatment after FIGNL1 overexpression and control lentivirus infection. (d) Cell migration and invasion were examined by transwell assay after FIGNL1 overexpression-inducing lentivirus infection and sb203580 treatment in Colo-205 cells. Scale bar: 200 µm. * p < 0.05 versus control overexpression group. # p < 0.05 versus FIGNL1 overexpression group. FIGNL1, Fidgetin-like 1; Con, control; OD, optical density, OE, overexpression; siRNA, Small interfering RNA

Journal: Balkan Medical Journal

Article Title: Exploring the Effect of Fidgetin-Like 1 on Colorectal Cancer Through Tissue Chip and In Vitro Experiments

doi: 10.4274/balkanmedj.galenos.2024.2024-7-9

Figure Lengend Snippet: FIGNL1 promoted CRC progression by activating the P38 pathway in vitro. (a) Representative blots demonstrating ERK/JNK/P38 MAPK expression following FIGNL1 siRNA treatment in Caco-2 cells. (b) Representative blots depicting ERK/JNK/P38 MAPK expression with lentivirus infection-related FIGNL1 overexpression in Colo-205 cells. (c) Line chart illustrating the Colo-205 cell proliferation following sb203580 treatment after FIGNL1 overexpression and control lentivirus infection. (d) Cell migration and invasion were examined by transwell assay after FIGNL1 overexpression-inducing lentivirus infection and sb203580 treatment in Colo-205 cells. Scale bar: 200 µm. * p < 0.05 versus control overexpression group. # p < 0.05 versus FIGNL1 overexpression group. FIGNL1, Fidgetin-like 1; Con, control; OD, optical density, OE, overexpression; siRNA, Small interfering RNA

Techniques: In Vitro, Expressing, Infection, Over Expression, Control, Migration, Transwell Assay, Small Interfering RNA

FIGNL1 activated P38 phosphorylation via SPIDR. (a) Prediction of proteins interacting with FIGNL1. (b) Gene enrichment was attained through gene ontology analysis. (c) The collective scores of various indicators regarding the interaction between FIGNL1 and the targeted proteins. (d) Representative blots depicting the binding between FIGNL1 and SPIDR. (e) Representative blots demonstrating P38 phosphorylation in FIGNL1-overexpression Colo-205 cells after SPIDR siRNA treatment. FIGNL1, Fidgetin-like 1; Con, control; OE, overexpression; IP, immunoprecipitation.

Journal: Balkan Medical Journal

Article Title: Exploring the Effect of Fidgetin-Like 1 on Colorectal Cancer Through Tissue Chip and In Vitro Experiments

doi: 10.4274/balkanmedj.galenos.2024.2024-7-9

Figure Lengend Snippet: FIGNL1 activated P38 phosphorylation via SPIDR. (a) Prediction of proteins interacting with FIGNL1. (b) Gene enrichment was attained through gene ontology analysis. (c) The collective scores of various indicators regarding the interaction between FIGNL1 and the targeted proteins. (d) Representative blots depicting the binding between FIGNL1 and SPIDR. (e) Representative blots demonstrating P38 phosphorylation in FIGNL1-overexpression Colo-205 cells after SPIDR siRNA treatment. FIGNL1, Fidgetin-like 1; Con, control; OE, overexpression; IP, immunoprecipitation.

Techniques: Phospho-proteomics, Binding Assay, Over Expression, Control, Immunoprecipitation

FIGNL1 interacting with SPIDR and facilitating CRC cell proliferation, migration, and invasion by activating the P38 pathway. FIGNL1, Fidgetin-like 1; CRC, colorectal cancer.

Journal: Balkan Medical Journal

Article Title: Exploring the Effect of Fidgetin-Like 1 on Colorectal Cancer Through Tissue Chip and In Vitro Experiments

doi: 10.4274/balkanmedj.galenos.2024.2024-7-9

Figure Lengend Snippet: FIGNL1 interacting with SPIDR and facilitating CRC cell proliferation, migration, and invasion by activating the P38 pathway. FIGNL1, Fidgetin-like 1; CRC, colorectal cancer.

Techniques: Migration

Journal: Balkan Medical Journal

Article Title: Exploring the Effect of Fidgetin-Like 1 on Colorectal Cancer Through Tissue Chip and In Vitro Experiments

doi: 10.4274/balkanmedj.galenos.2024.2024-7-9

Article Snippet: Following embedding, slicing, dewaxing, and hydration, the slides were stained with the primary antibody against FIGNL1 (1:200, 17604-1-AP, Proteintech, Wuhan, China).

Techniques: Expressing, Immunohistochemical staining

Journal: Balkan Medical Journal

Article Title: Exploring the Effect of Fidgetin-Like 1 on Colorectal Cancer Through Tissue Chip and In Vitro Experiments

doi: 10.4274/balkanmedj.galenos.2024.2024-7-9

Article Snippet: Following embedding, slicing, dewaxing, and hydration, the slides were stained with the primary antibody against FIGNL1 (1:200, 17604-1-AP, Proteintech, Wuhan, China).

Techniques: Knockdown, Migration, Expressing, Transwell Assay, Transfection, Control, Small Interfering RNA

Journal: Balkan Medical Journal

Article Title: Exploring the Effect of Fidgetin-Like 1 on Colorectal Cancer Through Tissue Chip and In Vitro Experiments

doi: 10.4274/balkanmedj.galenos.2024.2024-7-9

Article Snippet: Following embedding, slicing, dewaxing, and hydration, the slides were stained with the primary antibody against FIGNL1 (1:200, 17604-1-AP, Proteintech, Wuhan, China).

Techniques: Over Expression, Migration, Expressing, Infection, Transwell Assay, Control

Journal: Balkan Medical Journal

Article Title: Exploring the Effect of Fidgetin-Like 1 on Colorectal Cancer Through Tissue Chip and In Vitro Experiments

doi: 10.4274/balkanmedj.galenos.2024.2024-7-9

Article Snippet: Following embedding, slicing, dewaxing, and hydration, the slides were stained with the primary antibody against FIGNL1 (1:200, 17604-1-AP, Proteintech, Wuhan, China).

Techniques: In Vitro, Expressing, Infection, Over Expression, Control, Migration, Transwell Assay, Small Interfering RNA

Journal: Balkan Medical Journal

Article Title: Exploring the Effect of Fidgetin-Like 1 on Colorectal Cancer Through Tissue Chip and In Vitro Experiments

doi: 10.4274/balkanmedj.galenos.2024.2024-7-9

Article Snippet: Following embedding, slicing, dewaxing, and hydration, the slides were stained with the primary antibody against FIGNL1 (1:200, 17604-1-AP, Proteintech, Wuhan, China).

Techniques: Phospho-proteomics, Binding Assay, Over Expression, Control, Immunoprecipitation

Journal: Balkan Medical Journal

Article Title: Exploring the Effect of Fidgetin-Like 1 on Colorectal Cancer Through Tissue Chip and In Vitro Experiments

doi: 10.4274/balkanmedj.galenos.2024.2024-7-9

Article Snippet: Following embedding, slicing, dewaxing, and hydration, the slides were stained with the primary antibody against FIGNL1 (1:200, 17604-1-AP, Proteintech, Wuhan, China).

Techniques: Migration

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