MST3 Regulates AMPK and YAP-Hippo Signaling in Cell Models Relevant to Renal Fibrosis.

YAP is a transcription cofactor in the Hippo pathway that interacts with the TEAD family of transcription factors in the nucleus to promote CTGF expression and stimulate cell growth. YAP hyperactivation is frequently observed in fibrotic diseases. The main kinases in the Hippo pathway, MST1/2, a member of the STE20 family, promote Lats phosphorylation, leading to YAP phosphorylation, which prevents its nuclear entry and thus inhibits cell growth. High cell density induces Lats phosphorylation, causing YAP phosphorylation and its exclusion from the nucleus. Additionally, energy stress, such as glucose deprivation, induces AMPK phosphorylation, which also prevents YAP from entering the nucleus. MST3, another member of the STE20 family, has been shown to regulate cell apoptosis, migration, polarization, and ion homeostasis in previous studies. We hypothesized that MST3 is involved in Hippo pathway-mediated fibrosis. To test this, we overexpressed HA-tagged MST3 (HA-MST3) and a kinase-dead mutant (HA-MST3-KD) in MDCK cells. When cells reached a high density, HA-MST3 was activated to phosphorylate YAP, promoting its nuclear exit and inhibiting cell growth. In contrast, HA-MST3-KD cells showed reduced phosphorylated YAP, resulting in YAP retention in the nucleus, continuous cell growth, and NIH/3T3 cell fibrosis. Interestingly, YAP did not exit the nucleus in HA-MST3-KD cells treated with the YAP inhibitor verteporfin, but it did exit under metformin treatment due to energy stress, accompanied by increased AMPK and YAP phosphorylation, which inhibited MST3-KD-mediated fibrosis. These findings suggest that metformin-induced AMPK activation could provide a therapeutic approach for MST3-KD-mediated fibrosis.