Function of a complex of p‐ Y42 RhoA GTPase and pyruvate kinase M2 in EGF signaling pathway in glioma cells

Abstract Epidermal growth factor (EGF) is known to be a critical stimulant for inducing the proliferation of glioma cancer cells. In our study, we observed that GST‐RhoA binds to pyruvate kinase M2 (PKM2) in vitro. While EGF reduced the levels of RhoA protein, it significantly increased p‐Y42 RhoA, as well as PKM1 and PKM2 in LN18 glioma cell line. We determined that RhoA undergoes degradation through ubiquitination involving SCF1 and Smurf1. Interestingly, we observed that p‐Y42 RhoA binds to PKM2, while the dephosphomimetic form, RhoA Y42F, did not. Additionally, our observation revealed that PKM2 stabilized both RhoA and p‐Y42 RhoA. Importantly, RhoA, p‐Y42 RhoA, and PKM2, but not RhoA‐GTP, were localized in the nucleus upon EGF stimulation. Knockdown of RhoA with siRNA resulted in the reduced levels of phosphoglycerate kinase1 (PGK1) and microtubule affinity‐regulating kinase 4 (MARK). Furthermore, we found that the promoter of PGK1 was associated with β‐catenin and YAP. Notably, p‐Y42 RhoA and PKM2 co‐immunoprecipitated with β‐catenin and YAP. Based on these findings, we proposed a novel mechanism by which p‐Y42 RhoA and PKM2, in conjunction with β‐catenin and YAP, regulate PGK1 expression, contributing to the progression of glioma upon EGF. image