Integrated analyses identify a poor-prognosis subtype of hepatocellular carcinoma regulated by a core microRNA regulatory circuitry

Abstract

Cancer stem cells (CSCs) cause tumor heterogeneity, relapse, and resistance to therapy. The underpinnings of CSCs remain to be elucidated, especially the underlying gene regulatory network. We here conducted integrated analyses and identified a miRNA-regulatory network defining a stemness subtype with poor-prognosis from TCGA hepatocellular carcinoma (HCC) cohort with independent validations. The poor-prognosis subtype was characterized by the signature expression pattern of CSCs orchestrated by two miRNAs and their mRNA targets that formed a core regulatory circuitry (CRC). Within the CRC, miR483-3p bound a complementary sequence on SOX9 promoter, facilitating the recruitment of RNA polymerase II and STAT3, which was essential for SOX9 transcription activation. SOX9 can further activate SOX4 expression. Both SOX4 and its associated activator lncSOX4 were the direct targets of miR204-5p. SOX4 and miR204-5p formed double-negative feedback loop through mutual inhibition. The expression level of miR204-5p was tightly modulated by miR483-3p, whose promoter was significantly de-methylated in the stemness subtype. Activation of the CRC essential for the self-renewal and maintenance of liver CSCs culminated in downregulation of miR204-5p and upregulation of miR483-3p, SOX9, and SOX4. Functional significance of the CRC for HCC metastasis and drug resistance was further demonstrated with various in vitro and in vivo assays.