Hypoxia-driven transcriptional activation of MIR100HG by HIF-1α contributes to adaptive gene regulation in hepatocellular carcinoma

Abstract

Background: Hepatocellular carcinoma (HCC) develops in a chronically hypoxic microenvironment; however, the contribution of hypoxia-regulated long non-coding RNAs (lncRNAs) remains poorly defined. Aim: We aimed to identify MIR100HG as a hypoxia-responsive regulator in hepatocellular carcinoma (HCC) and to define its transcriptional control by hypoxia-inducible factor 1 alpha. Methods: Basal MIR100HG expression was analyzed in four hepatocellular carcinoma cell lines (Hep3B, Huh-7, SNU-398, and SNU-475) and non-tumorigenic Clone-9 hepatocytes. Hypoxia was induced by cobalt (II) chloride. Proliferation, migration, and clonogenicity were evaluated using functional assays. Promoter analysis, electrophoretic mobility shift assay, chromatin immunoprecipitation quantitative polymerase chain reaction, and 5′-deletion luciferase assays were used to map the hypoxia response elements. Transcriptomic profiling was performed using RNA sequencing, followed by gene set enrichment analysis. Gain- and loss-of-function experiments validated the downstream effects. Results: MIR100HG expression was highest in Hep3B cells and was significantly induced under hypoxia in all hepatic models, indicating a conserved, oxygen-sensitive response. Hypoxia suppressed proliferation, migration, and clonogenicity, with the strongest inhibition observed in non-tumorigenic hepatocytes under hypoxic conditions. Functional hypoxia response elements were identified within the −815/+231 promoter region, demonstrating direct transcriptional activation by hypoxia-inducible factor 1 alpha. Transcriptomic analyses revealed the enrichment of hypoxia signaling, oxidative stress, proteasome activity, RNA metabolism, autophagy, and ferroptosis pathways. While hypoxia induced broad transcriptional reprogramming, MIR100HG modulated the expression of selected hypoxia-responsive genes, including the PAGE1 and CTAG2, as demonstrated under hypoxic conditions by RNA-seq and functional perturbation experiments. Conclusions: MIR100HG is a hypoxia-inducible factor 1 alpha-driven long non-coding RNA associated with hypoxiaresponsive transcriptional programs in hepatocellular carcinoma and represents a potential biomarker and therapeutic target.