Ribosomal s6 kinase (RSK) is a mediator of aquaporin 2 S256 phosphorylation and membrane accumulation after EGFR inhibition with erlotinib

Vasopressin (VP) activates PKA, resulting in phosphorylation and membrane accumulation of aquaporin-2 (AQP2). Epidermal growth factor receptor (EGFR) inhibition with erlotinib also induces AQP2 membrane trafficking with a phosphorylation pattern similar to VP, but without increasing PKA activity. Here, we identify the ribosomal s6 kinase (RSK) as the final mediator phosphorylating AQP2 in this novel, erlotinib-induced pathway. We found that RSK was expressed in medullary principal cells in rat kidneys. RSK inhibition with BI-D1870 or siRNA blocked erlotinib-induced AQP2 S256 phosphorylation and membrane trafficking. CRISPR-generated RSK knockout cells failed to show increased S256 phosphorylation in response to erlotinib. Like PKA, RSK was able to phosphorylate AQP2 S256 in vitro . Stimulation of PDK1, a known activator of RSK, caused AQP2 S256 phosphorylation and membrane accumulation similar to erlotinib and VP. We conclude that RSK is the terminal kinase phosphorylating AQP2 at S256 upon EGFR inhibition by erlotinib.