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. 2014 Feb 5;9(2):e88405. doi: 10.1371/journal.pone.0088405

Figure 4. PKCα binds to Nox5 and regulates its activity through direct phosphorylation of Ser490, Thr494 and Ser498.

Figure 4

(A) COS-7 cells expressing Nox5 were lysed and immunoprecipitated using either control IgG or anti-HA antibody. Immune complexes were immunoblotted for HA-Nox5 or PKCα. (B) Nox5 was immunoprecipitated from COS-7 cells transduced with HA-Nox5 adenovirus and subject to an in vitro kinase assay. Phosphorylated samples were immunoblotted for phosphorylated Ser490, Thr494, and Ser498 versus total Nox5. Superoxide release from COS-7 cells cotransfected with HA-Nox5 WT or triple mutant (Nox5 S490A,T494A,S498A) and either control (RFP) or myr-PKCα (C). Lysates were immunoblotted for PKCα, MEK1/2, ERK1/2 phosphorylation, phosphorylated S490, T494 and S498 relative to total Nox5 (HA) expression (D). Results are representative of at least 3 separate experiments, presented as means ± S.E., * p<0.05 versus Vehicle.