Journal: Frontiers in Molecular Biosciences

Article Title: CK2β Is a Gatekeeper of Focal Adhesions Regulating Cell Spreading

doi: 10.3389/fmolb.2022.900947

Figure Lengend Snippet: Upregulation of pTyr proteins in lysates of ΔCK2β MCF10A cells. (A) Lysate s from WT and ΔCK2β MCF10A cells were analyzed for pTyr-containing proteins using the 4G10 antibody. GAPDH was used as a loading control. (B) Cell extracts (500 μg of protein) of WT and ΔCK2β MCF10A cells were loaded on 4G10 resin. After extensive washing, the bound proteins were eluted with 100 mM phenyl phosphate. The collected fractions were analyzed by Western blotting with the 4G10 antibody. (C) Proteins immunopurified from WT and ΔCK2β MCF10A cell lysates were analyzed by Western blotting for p-FAK (Y576/577), p-Src (Y418), p-PAX1(Y118), p-p130CAS (Y410), FAK, Src, PAX1, and p130CAS and (D) for p-EGFR (Y1173), EGFR, p-PLCγ (Y771), and PLCγ.

Article Snippet: Western blot membranes were immunostained with primary antibodies as follows: phosphotyrosine 4G10 clone, #05-321 (Merck); P-FAK1 Y576/577, #3281, P-PAX1 Y118, #2541, P-p130CAS Y410, #4011, and P-EGFR Y1173, #4407 (Cell Signaling, Danvers, MA, United States); P-Src Y418, #11091 (SAB Signaling Antibody, Maryland, United States); Src, #sc-19, and EGFR, #sc-03 (Santa Cruz Biotechnology Inc., Santa Cruz, CA, United States); PAX1, #610052, FAK1, #610088, and p130CAS, #610271 (BD Biosciences, Pont de Claix, France); P-PLCγ Y771, #ab131455, and PLCγ, #ab52200 (Abcam, Paris, France); and GAPDH, #AM4300 (Invitrogen, Thermo Fisher, France).

Techniques: Western Blot