SDF-1–mediated ERK activation of HEK/CD4.403 cells expressing wild-type and mutated CXCR4 molecules.

(A) Serum-starved transfected cells were stimulated for 2 minutes with SDF-1 at 125 nM and lysed as described in “Materials and methods.” Protein samples were run on an SDS-polyacrylamide gel and Western blotted with an antiphosphoERK1/2 antibody. Protein loading was controlled by using a total anti–ERK1/2 antibody. (B) Densitometric analysis of phosphorylated ERK expression. Data are presented as fold increases of ERK phosphorylation, in which the amount of ERK phosphorylation in cells that do not express CXCR4 is assigned a value of 1.0 after SDF-1 activation. All the relative intensities were calculated by normalizing the intensity of phosphorylated ERK protein to its ERK protein loading control. Results shown are from 2 to 5 independent experiments; error bars reflect SDs. Statistical analysis was performed as described in “Materials and methods” (*P < .05, **P < .01, ***P < .001). (C) ERK phosphorylation induced by SDF-1 in HEK/CD4.403/CXCR4 cells in the presence or absence of PTX.