The Oncogenic Effects, Pathways, and Target Molecules of JC Polyoma Virus T Antigen in Cancer Cells

Abstract

JC polyoma virus (JCPyV) is a ubiquitous polyoma virus that infects the individual to cause progressive multifocal leukoencephalopathy and malignancies. Here, we found that T-antigen knockdown suppressed proliferation, glycolysis, mitochondrial respiration, migration, and invasion, and induced apoptosis and G₂ arrest. The reverse was true for T-antigen overexpression, with overexpression of Akt, survivin, retinoblastoma protein, β-catenin, β-transducin repeat-containing protein (TRCP), and inhibitor of growth (ING)1, and the underexpression of mammalian target of rapamycin (mTOR), phosphorylated (p)-mTOR, p-p38, Cyclin D1, p21, vascular endothelial growth factor (VEGF), ING2, and ING4 in hepatocellular and pancreatic cancer cells and tissues. In lens tumor cells, T antigen transcriptionally targeted viral carcinogenesis, microRNAs in cancer, focal adhesion, p53, VEGF, phosphoinositide 3 kinase-Akt, and Forkhead box O signaling pathways, fructose and mannose metabolism, ribosome biosynthesis, and choline and pyrimidine metabolism. At a metabolomics level, it targeted protein digestion and absorption, aminoacryl-tRNA biosynthesis, biosynthesis of amino acids, and the AMPK signal pathway. At a proteomic level, it targeted ribosome biogenesis in eukaryotes, citrate cycle, carbon metabolism, protein digestion and absorption, aminoacryl-tRNA biosynthesis, extracellular-matrix-receptor interaction, and biosynthesis of amino acids. In lens tumor cells, T antigen might interact with various keratins, ribosomal proteins, apolipoproteins, G proteins, ubiquitin-related proteins, RPL19, β-catenin, β-TRCP, p53, and CCAAT-enhancer-binding proteins in lens tumor cells. T antigen induced a more aggressive phenotype in mouse and human cancer cells due to oncogene activation, inactivation of tumor suppressors, and disruption of metabolism, cell adhesion, and long noncoding RNA-microRNA-target axes.

Keywords: JC virus T antigen; PI3k-Akt signal pathway; WNT/beta-catenin pathway; oncogenesis; signal pathway.

Figure 1

The effects of T antigen knockdown on the aggressive phenotypes of lens tumor cells. After transfection of short-hairpin (sh)RNA-T antigen, its expression became weak in lens tumor cells as determined by real-time reverse transcription (RT)-PCR (A), Western blot (B), and immunofluorescence (C). Cell viability in lens tumor cells and their transfectants were measured using a cell counting kit-8 (CCK-8) kit (D). Cell cycle, apoptosis, glucose metabolism, and invasion were examined by propidium iodide (PI, E), XF-24 extracellular flux analyzers (F), and transwell assays (G), respectively. ^*p < 0.05, compared with transfectants. NC, negative control.

Figure 2

The effects of T-antigen overexpression on the aggressive phenotypes of cancer cells. After transfection of pEGFP-N1-T antigen, strong expression was observed in QGY7703 and PANC cells as determined by Western blot (A). Cell viability in both cancer cells and their T-antigen transfectants was measured using a cell counting kit-8 (CCK-8) kit (B). Apoptosis, migration, and invasion were measured by Annexin-V staining (C) and transwell chamber (D) assays, respectively. ^*p < 0.05, compared with control and mock.

Figure 3

The effects of T antigen on transcriptome of lens tumor cells. The top signaling pathways for upregulated and downregulated genes after T-antigen knockdown in lens tumor cells are listed according to the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis.

Figure 4

The effects of T antigen on metabolome of lens tumor cells. Relevant Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways after T-antigen knockdown in lens tumor cells were classified into metabolism, genetic information processing, environmental information processing, cell processes, or organismal systems (A). The top signaling pathways are listed according to the KEGG analysis (B).

Figure 5

The effects of T antigen on protein expression in cancer cells and spontaneous tumors. Western blot was employed to examine the distinct protein expression in T-overexpressing QGY-7703 and PANC-1 cells, and in spontaneous hepatocellular carcinoma or pancreatic adenocarcinomas of Alb-cre/T antigen (ALJ) and Pdx1-cre/T antigen (PDJ) transgenic mice. Parental cells and liver and pancreas of wild-type (WT) mice were used as controls.

Figure 6

The partner proteins of JCV T antigen in lens tumor cells. Proteins from T antigen-silenced lens tumor cells were subjected to coimmunoprecipitation (co-IP) by anti-SV40 T-antigen antibody. Co-IP samples were analyzed by Western blot and by electrophoresis and silver staining (A). Protein samples from anti-SV40 T-antigen antibody co-IP were screened for binding proteins to T antigen (B). Protein samples from co-IP using anti-RPL19 antibody were screened for interacting proteins (C). Western blot was used to detect whole cell lysates as an internal control (D). NC, negative control; WCL, whole cell lysates.