HIF2α drives ccRCC metastasis through transcriptional activation of methylation-controlled J protein and enhanced prolegumain secretion.

The role of hypoxia-inducible factor 2α (HIF2α) in clear cell Renal Cell Carcinoma (ccRCC) is still not fully understood. In this study, we identified that urinary prolegumain levels positively correlated with the malignant characteristics of ccRCC. In cultured 786-O and OSRC-2 cells, HIF2α downregulation reduced prolegumain secretion. RNA sequencing assay revealed that HIF2α induces methylation-controlled J (MCJ), a negative regulator on the mitochondrial respiratory chain. Silencing MCJ reduced prolegumain secretion, and MCJ overexpression restored prolegumain secretion inhibited by HIF2α downregulation. Chromatin immunoprecipitation and luciferase assay confirmed MCJ as a transcription target of HIF2α. Furthermore, we showed the ectopic MCJ overexpression reversed the improved mitochondrial damage resulting from HIF2α downregulation, as evidenced by electron microscope, ATP level, GSSG/GSH ratio, MitoSOX, and DHE staining. Through mass spectrometry analysis, we identified oxidation site His343 on the legumain sequence as contributing to the prolegumain secretion. Therapeutically, silencing MCJ or HIF2α or using ROS scavengers Vitamin C or MitoQ alleviated MMP2 activation as well as cell migration and tube formation. In a mouse orthotopic xenograft model of ccRCC, silencing MCJ or administration of MitoQ significantly protected against mitochondrial damage and subsequently reduced the lung metastasis of tumors. Overall, our study identified MCJ as a target molecule of HIF2α in ccRCC. Silencing MCJ or using ROS scavengers like MitoQ can suppress oxidation site His343 in legumain, preventing prolegumain secretion and subsequently reducing metastasis of ccRCC.