Downregulation of PRMT6 promotes the Warburg effect in hepatocellular carcinoma via PKM2

Abstract

Hepatocellular carcinoma (HCC) is one of the top ranked causes of cancer death in the world, accounting for more than 740,000 deaths in 2012. Treatment of HCC is hindered by recurrence and resistance to chemotherapy. Altered metabolism in cancer has been linked to increased tumorigenicity and resistance to chemotherapeutics. One of the main features of altered metabolism is the Warburg effect, which is widely observed in solid tumor including HCC. Recently, our group have identified protein arginine methyltransferase 6 (PRMT6) as a tumor suppressor protein in HCC. Protein arginine methylation is a post-translational modification implicated in a variety of cellular functions including signaling and regulation of gene expression.Downregulation of PRMT6 expression was observed in HCC patients and knockdown of PRMT6 in HCC cell lines increased the tumorigenicity and sensitivity to both chemo- and molecular-targeted therapies. Other members of the Type 1 PRMT protein has previously been reported to be involved in glucose metabolism. Microarray profiling of HCC cells stably knockdown and overexpressed of PRMT6 showed enrichment of genes in glycolysis. We therefore hypothesize that reduced expression of PRMT6 in HCC drives the Warburg effect to support cancer progression. Metabolome profiling of HCC cells stably knockdown and overexpressed of PRMT6 showed enrichment of metabolites in glycolysis. Using Seahorse Extracellular Flux Analyzer, we show that stable knockdown of PRMT6 in HCC cell line drives glycolysis. This enhanced glycolysis rate is coupled with increased uptake of glucose, lactate production and pyruvate kinase activity. mRNA expression level of several glycolysis-related genes was screened by real-time quantitative PCR and we identified PKM2, a well-known regulator of the Warburg effect, as a potential link between PRMT6 and enhanced glycolysis. Knockdown of PRMT6 leads to an increase in mRNA and protein expression level of PKM2, but not PKM1 and PKLR, suggesting that PKM2 is the sole contributor for enhanced pyruvate kinase activity. In addition, by immunofluorescence staining, PKM2 is shown to accumulate in the nucleus following PRMT6 knockdown, where it has been shown to drive the expression of other glycolysis-related genes. In conclusion, PRMT6 regulates the expression level PKM2 and glycolysis of HCC cell. Ongoing studies are in progress to delineate the underlying mechanisms by which PRMT6 regulates PKM2 to drive Warburg effect.