Fig. 1

CXCL12/CXCR4 transactivation of growth factor receptors occurs in lipid raft microdomains. a C4-2B and PC3 cells were cultured in the presence or absence of CXCL12. Total cell lysates were analyzed for phosphorylated and total HER2 and EGFR. b C4-2B and PC3 cells were cultured in the presence or absence of CXCL12. Cell lysates were immunoprecipated with anti-HER2 and EGFR antibodies and immunoblotted with total and phosphorylated HER2 and EGFR antibodies. c C4-2B and PC3 cells were cultured in the presence or absence of CXCL12. Lipid raft membrane microdomains were isolated using a successive detergent solubilization method; both lipid rafts and cellular membranes and cytosol fractions were immunoblotted with phosphorylated and native HER2 and EGFR antibodies. d PC3 cells were lysed in a detergent-free lysis buffer and fractionated by sucrose density gradient centrifugation. Fractions were immunoblotted with lipid raft marker Flotillin-2 and cytosol marker β-Tubulin. The lipid raft enriched fraction (#5) was immunoblotted with HER2 and phosphorylated HER2 antibodies