STK3 is a transcriptional target of YAP1 and a hub component in the crosstalk between Hippo and Wnt signaling pathways during gastric carcinogenesis

Background: Serine/threonine kinase 3 (STK3) is recognized as a key regulator in Hippo pathway and a tumor-suppressing gene in various cancer types. However, its non-canonical role has been gradually revealed in cancer development. Methods: Our objective is to elucidate the upregulation pattern and molecular mechanisms of STK3 in advancing gastric cancer (GC) progression. The regulation of YAP1 on STK3 was assessed through a combination of bulk and single-cell RNA-sequencing, Western blot, ChIP-qPCR, gene knockout mouse models, and functional rescue assays. The oncogenic roles of STK3 were confirmed through subcutaneous xenograft formation models and functional assays including spheroid formation and organoid growth. The phosphorylated target of STK3 was revealed by co-immunoprecipitation and in vitro kinase assays. STK3-targeted drugs were screened out by molecular docking and cellular thermal shift assay (CETSA). Results: Reduction of YAP1 significantly impaired STK3 expression at both mRNA and protein levels, and deletion of STK3 partially attenuated the oncogenic activity of YAP1. Notably, MNNG-induced tumors in Yap1 ^−/−Taz ^−/− mice exhibited decreased STK3 expression. Knockdown of STK3 led to reduced expression of stemness markers and xenograft growth, while sensitizing GC organoids and xenografts to 5-fluorouracil treatment. Mechanistically, the direct interaction between STK3 and GSK-3β promoted GSK-3β phosphorylation and β-catenin nuclear accumulation, and thus the activation of Wnt signaling. Furthermore, aminopterin demonstrates as a promising STK3-targeted small molecule with remarkable effectiveness in inhibiting GC cell malignance and xenograft growth. Conclusions: STK3 was identified as a transcriptional target of YAP1, leading to enhanced DNA repair ability and stemness acquisition during GC progression by activating Wnt/β-catenin activity through GSK-3β degradation. Moreover, STK3-targeted therapy offered a novel approach to concur acquired chemo-resistance in GC patients.