Transcriptome-Wide 5-Methylcytosine Functional Profiling of Long Non-Coding RNA in Hepatocellular Carcinoma

Growing evidence indicates that methylation status is associated with the pathogenesis of numerous types of cancers. Among these, hepatocellular carcinoma (HCC) is a deadly disease threatening global human health. Although 5-methylcytosine (m5C) has been identified as an important regulatory modification, its distribution in solid tumors, including HCC, remains unclear. The present study aimed to explore the distribution of m5C in HCC.

Six pairs of human HCC tissues and adjacent non-tumor tissues were collected to analyze the transcriptome-wide m5C methylation of long non-coding RNA (lncRNA). RNA MeRIP-seq was performed to identify m5C peaks on lncRNA and differences in m5C distribution between HCC and adjacent tissues. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment pathway analyses were explored to predict the possible roles of m5C.

Using m5C peak sequencing, we observed that a sequence motif was necessary for m5C methylation in HCC lncRNA. Unsupervised hierarchical cluster analysis confirmed that lncRNA m5C methylation occurred more frequently in HCC than adjacent non-tumor tissues. RNA sequencing data demonstrated that more genes were up-regulated by methylation in HCC, while methylation down-regulated more genes in adjacent non-tumor tissues. GO and KEGG pathway analyses revealed that genes having a significant correlation with m5C sites in lncRNA were involved in HCC signaling pathways.

Our results revealed the substantially different amounts and distributions of m5C in HCC compared to adjacent non-tumor tissue. We further predicted the cellular functions in HCC that m5C may participate in to provide evidence implicating m5C lncRNA epigenetic regulation in the tumorigenesis and progression in HCC.