Alternative splicing (AS) allows generation of cell type–specific mRNA transcripts and contributes to hallmarks of cancer. Genome‐wide analysis for AS in human hepatocellular carcinoma (HCC), however, is limited. We sought to obtain a comprehensive AS landscape in HCC and define tumor‐associated variants. Single‐molecule real‐time long‐read RNA sequencing was performed on patient‐derived HCC cells, and presence of splice junctions was defined by SpliceMap‐LSC‐IDP algorithm. We obtained an all‐inclusive map of annotated AS variants and further discovered 362 alternative spliced variants that are not previously reported in any database (neither RefSeq nor GENCODE). They were mostly derived from intron retention and early termination codon with an in‐frame open reading frame in 81.5%. We corroborated many of these predicted unannotated and annotated variants to be tumor specific in an independent cohort of primary HCC tumors and matching nontumoral liver. Using the combined Sanger sequencing and TaqMan junction assays, unique and common expressions of spliced variants including enzyme regulators (ARHGEF2, SERPINH1), chromatin modifiers (DEK, CDK9, RBBP7), RNA‐binding proteins (SRSF3, RBM27, MATR3, YBX1), and receptors (ADRM1, CD44v8‐10, vitamin D receptor, ROR1) were determined in HCC tumors. We further focused functional investigations on ARHGEF2 variants (v1 and v3) that arise from the common amplified site chr.1q22 of HCC. Their biological significance underscores two major cancer hallmarks, namely cancer stemness and epithelial‐to‐mesenchymal transition–mediated cell invasion and migration, although v3 is consistently more potent than v1. Conclusion: Alternative isoforms and tumor‐specific isoforms that arise from aberrant splicing are common during the liver tumorigenesis. Our results highlight insights gained from the analysis of AS in HCC.
Apart from b-catenin accumulation, loss of 3p21 is one of the most frequent genetic alterations in numerous malignancies including nasopharyngeal carcinoma (NPC). Herein, we characterized a novel candidate tumor suppressor gene (TSG) CAC-NA2D3, a voltage-dependent subunit alpha 2 delta 3 of a calcium channel complex. Downregulation of CACNA2D3 was frequently detected in primary NPCs and NPC cell lines compared with their nontumorigenic counterparts. Attenuated CACNA2D3 expression may be associated with loss of heterozygosity (LOH) at intragenic single-nucleotide polymorphism sites (rs589281, rs1449325 and rs6797113) and/or epigenetic silencing by methylation and histone deacetylation. Given the extensive effects of calcium in cancer, we then investigated the tumor suppressive role and underlying mechanism of CACNA2D3 in the development and progression of NPC. CACNA2D3 was stably transfected into NPC cell lines (C666 and SUNE1) at levels comparative with the normal nasopharynx, alongside siRNA-mediated silencing in an immortalized nasopharyngeal epithelial cell line (NP69) to conduct in vivo and in vitro functional assays. Our findings show that CACNA2D3-mediated increase in intracellular calcium (Ca21) can induce mitochondrial-mediated apoptosis and activation of NLK (through the Wnt/Ca21 pathway) to antagonize Wnt signaling-mediated anchorage-dependent and independent cell proliferation (via CCND1 and CMYC), invasion (via MMP7) and epithelial-to-mesynchemal transition (via SNAIL). As the expression pattern of calcium channels and their degree of functionality can change with the progression of cancer, CACNA2D3 may indeed be a promising biomarker for NPC. Our study also warrants further exploration in the potential therapeutic use of existing epigenetic targeting drugs (e.g., 5-azacytidine, SAHA) to reconstitute CACNA2D3-associated tumor suppression in NPC.
Co-infection of Cytomegalovirus and Epstein-Barr Virus Diminishes the Frequency of CD56 dim NKG2A + KIR − NK Cells and Contributes to Suboptimal Control of EBV in
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