GRK6 expression is negatively regulated by GF/PDGFR. (A) Quantitative real time RT-PCR was performed to determine the expression of GRK6 mRNA in medulloblastoma cells with normal vs. downregulated PDGFR expression. (A) GRK6 mRNA is increased 1.5 folds in B9 (PDGFRβ down-regulated by shRNA in D556, shown below left panel) and 1.5 folds in A4 (PDGFRβ down-regulated by shRNA in Daoy, shown below right panel) respectively, compared to the parental cells (D556 NC1 or Daoy NC1), P < 0.05 (N = 3). (B) Protein level of GRK6 is increased in D556 (B9) and Daoy (A4) with or without CXCL12 treatment. (C) D556 or Daoy was cultured in EMEM medium containing 10% serum (GF+) or serum-free EMEM (GF-) for 24 h, then added 10% serum to the cells cultured in serum-free medium (GF add-back) and cells grown for 24-72 h. Cell lysates were harvested for Western Blot. GF withdrawal results in robust increase of GRK6 in D556 and Daoy (left and middle panels). In right panel, Daoy cells were starved for 24 h and then treated with 10 ng/ml PDGF-BB for 24 h. GRK6 level determined by Western blot shows marked decrease with PDGF treatment.