rabbit-anti-p-fak (y397)  (Cell Signaling Technology Inc)

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: The oncolytic avian reovirus p17 protein suppresses invadopodia formation via disruption of TKs5 complexes and oncogenic signaling pathways

doi: 10.3389/fcimb.2025.1603124

Figure Lengend Snippet: The ARV p17 protein promotes PTEN expression in HeLa and A549 cells and inhibits FAK phosphorylation at Y397. (A) The expression levels of PTEN, phosphorylated FAK (p-FAK Y397), and total FAK were analyzed in p17-transfected cancer cells and untreated control cells. Whole-cell lysates were collected at 0, 6, 12, 18, and 24 hours post-transfection, followed by Western blot analysis. β-actin was used as a loading control. The fold changes in activation and inactivation indicated below each lane, were normalized against those in the mock-transfected cells. Protein levels in the mock group were set as 1-fold for comparison. (B) Signals for all blots were quantified using ImageJ software. Data in panel A are means and SE from three independent experiments.

Article Snippet: Rabbit-anti-p-FAK (Y397) , 8556 , , 1500 , Cell Signaling.

Techniques: Expressing, Phospho-proteomics, Transfection, Control, Western Blot, Activation Assay, Comparison, Software

Journal: Translational Cancer Research

Article Title: CSE1L in enhancing the effect of defactinib on gastric cancer cells via the inhibition of FAK phosphorylation

doi: 10.21037/tcr-24-2049

Figure Lengend Snippet: CSE1L upregulated the focal adhesion pathway. (A) A volcano plot illustrating the differential genes between CSE1L KO and CSE1L NC based on transcriptome data. (B) KEGG pathway enrichment analysis of the differential genes between CSE1L KO and CSE1L NC . (C) Heatmap showing the differential genes associated with the focal adhesion pathway between CSE1L KO and CSE1L NC . (D) Quantitative real-time PCR showed that the expression of ITGB4, PTK2, β-actin, and MYL9 decreased in MGC-803 cells upon knockout of CSE1L. The experiments were performed in triplicate. (***, P<0.001; ****, P<0.0001 by t -test). (E) Quantitative real-time PCR showed that the expression of ITGB4, PTK2, β-actin, and MYL9 increased in BGC-823 cells upon overexpression of CSE1L. The experiments were performed in triplicate. (*PTK2: P=0.02; *β-actin: P=0.02; *MYL9: P=0.03; ***, P<0.001 by t -test). (F) WB showed that compared with the CSE1L NC , CSE1L KO significantly inhibited the expression of ITGB4, p-FAK, FAK, β-actin, and MYL9. (G) WB showed that compared with the CSE1L NC , CSE1L OE increased the expression of ITGB4, p-FAK, FAK, β-actin, and MYL9. KO, knockout; NC, negative control; CT, cycle threshold; p-FAK, phosphorylated FAK; FAK, focal adhesion kinase; Y397, tyrosine 397; CSE1L, chromosome segregation 1 like; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; KEGG, Kyoto Encyclopedia of Genes and Genomes; PCR, polymerase chain reaction; WB, western bolt.

Article Snippet: Antibodies against FAK (66258-1-Ig; Proteintech), p-FAK (Y397) (T55587; Abmart, Shanghai, China), CSE1L (PA5-21468; Invitrogen), β-actin (66009-1-Ig; Proteintech), GAPDH (60004-1-Ig; Proteintech), integrin beta-4 (ITGB4) (21738-1-AP; Proteintech), and myosin regulatory light polypeptide 9 (MYL9) (15354-1-AP; Proteintech) were used according to respective the manufacturer’s instructions.

Techniques: Real-time Polymerase Chain Reaction, Expressing, Knock-Out, Over Expression, Negative Control, Polymerase Chain Reaction, Western Blot

Journal: Translational Cancer Research

Article Title: CSE1L in enhancing the effect of defactinib on gastric cancer cells via the inhibition of FAK phosphorylation

doi: 10.21037/tcr-24-2049

Figure Lengend Snippet: The expression of CSE1L influenced the efficacy of defactinib. (A) CCK8 assay showed that the knockout of CSE1L significantly enhanced the killing effect of defactinib on MGC-803 cells. Conversely, the overexpression of CSE1L weakened the killing effect of defactinib on BGC-823 cells (*CSE1L NC vs. CSE1L KO : P=0.03; **CSE1L NC + defactinib vs. CSE1L KO + defactinib : P=0.001; **CSE1L OE + defactinib vs. CSE1L NC + defactinib : P=0.002; ***, P<0.001; ****, P<0.0001 by t -test). (B) WB revealed that compared with the CSE1L KO , overexpression of the CSE1L could promote FAK (Y397) phosphorylation. To test the inhibition of FAK (Y397) phosphorylation via defactinib in MGC-803 cells and BGC-823 cells, the cells were exposed to defactinib at a dose of 10 μM for 16 hours before WB assays were performed. (C) Representative images of immunofluorescence staining of p-FAK (Y397) in CSE1L NC and CSE1L KO MGC-803 cells and in CSE1L NC and CSE1L OE BGC-823 cells as assessed by confocal microscopy (from three independent experiments). DAPI = blue, pFAK Y397 = green, and phalloidin = red. Scale bar: 10 μm. (D) The expression of CSE1L and FAK in GC cell lines (MGC-803, BGC-823, SGC-7901, HGC-27, MKN-45, MKN-74, AGS, and N87) was detected using WB. (E) FAK expression was positively associated with CSE1L expression according to the protein volume data from WB (P=0.001; R=0.952). (F) PTK2 expression was positively associated with CSE1L expression in clinical GC samples. Gene correlation analysis was based on the TCGA gastric carcinoma dataset. (G) CCK8 assay showed that defactinib significantly killed BGC-823 cells but that the killing effect on MGC-803 cells was weak. Cells were treated with defactinib at a dose of 10 μM for 18 hours before CCK8 assays were performed. The experiment was performed in triplicate. (*MGC-803: P=0.03; **AGS: P=0.007; **MKN-74: P=0.005; ***, P<0.001; ****, P<0.0001 by t -test). CSE1L, chromosome segregation 1 like; NC, negative control; KO, knockout; OD, optical density; OE, overexpression; p-FAK, phosphorylated FAK; FAK, focal adhesion kinase; Y397, tyrosine 397; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; DAPI, 4',6-diamidino-2-phenylindole; CCK8, Cell Counting Kit 8; WB, western bolt; GC, gastric cancer; TCGA, The Cancer Genome Atlas.

Article Snippet: Antibodies against FAK (66258-1-Ig; Proteintech), p-FAK (Y397) (T55587; Abmart, Shanghai, China), CSE1L (PA5-21468; Invitrogen), β-actin (66009-1-Ig; Proteintech), GAPDH (60004-1-Ig; Proteintech), integrin beta-4 (ITGB4) (21738-1-AP; Proteintech), and myosin regulatory light polypeptide 9 (MYL9) (15354-1-AP; Proteintech) were used according to respective the manufacturer’s instructions.

Techniques: Expressing, CCK-8 Assay, Knock-Out, Over Expression, Inhibition, Immunofluorescence, Staining, Confocal Microscopy, Negative Control, Cell Counting, Western Blot