Fig. 4.

PKCγ directly phosphorylates CaMKIIβ at S315. (A) Detection of PKCγ-mediated phosphorylation of GST-CaMKIIβ (GST-β) (272–407 aa) with anti–pS315-CaMKIIβ antibody. GST or GST-β (272–407 aa) were phosphorylated by PKCγ in vitro and subjected to immunoblot analysis. (B) Immunoblot analysis of WT and S315A mutant form of HA-tagged CaMKIIβ that were cotransfected with PKCγ-Myc in HeLa cells. Transfected cells were treated with or without 100 nM PMA for 30 min and subjected to immunoblot analysis with the indicated antibodies. (C) Immunocytochemistry of S315 phosphorylation of WT or S315A mutant form of HA-CaMKIIβ cotransfected with PKCγ-flag in HeLa cells. The cells were treated with (+) or without (−) PMA and stained with the indicated antibodies. (Scale bar, 100 μm.)