Characterization and validation of PRKCA as a component in SH4-dependent protein transport. (A) HeLa cells expressing SH4-HASPB-GFP and SH4-YES1-mCherry were transfected with a scrambled siRNA (a–c), and a siRNA directed against PRKCA (d–f), and subjected to live-cell confocal microscopy to determine the localization of both SH4 fusion proteins. (B) Equal protein amounts of total cell lysates from transfected and induced cells were subjected to SDS-PAGE and Western blot analysis with antibodies directed against GFP, PRKCA, and GAPDH. (C) Stable HeLa cells expressing SH4-HASPB-GFP (a,c), or SH4-YES1-GFP (b,d), were cultured in the presence of 10 μM protein kinase C inhibitor GÖ 6983 for 16 h (c,d), and analyzed by live-cell confocal microscopy for subcellular localization of both SH4 fusion proteins.
