Identification of Deregulated Pathways, Key Regulators, and Novel miRNA-mRNA Interactions in HPV-Mediated Transformation

Abstract

Next to a persistent infection with high-risk human papillomavirus (HPV), molecular changes are required for the development of cervical cancer. To identify which molecular alterations drive carcinogenesis, we performed a comprehensive and longitudinal molecular characterization of HPV-transformed keratinocyte cell lines. Comparative genomic hybridization, mRNA, and miRNA expression analysis of four HPV-containing keratinocyte cell lines at eight different time points was performed. Data was analyzed using unsupervised hierarchical clustering, integrated longitudinal expression analysis, and pathway enrichment analysis. Biological relevance of identified key regulatory genes was evaluated in vitro and dual-luciferase assays were used to confirm predicted miRNA-mRNA interactions. We show that the acquisition of anchorage independence of HPV-containing keratinocyte cell lines is particularly associated with copy number alterations. Approximately one third of differentially expressed mRNAs and miRNAs was directly attributable to copy number alterations. Focal adhesion, TGF-beta signaling, and mTOR signaling pathways were enriched among these genes. PITX2 was identified as key regulator of TGF-beta signaling and inhibited cell growth in vitro, most likely by inducing cell cycle arrest and apoptosis. Predicted miRNA-mRNA interactions miR-221-3p_BRWD3, miR-221-3p_FOS, and miR-138-5p_PLXNB2 were confirmed in vitro. Integrated longitudinal analysis of our HPV-induced carcinogenesis model pinpointed relevant interconnected molecular changes and crucial signaling pathways in HPV-mediated transformation.

Human papillomavirus E6 and E7: What remains?

“…Recently attention has been directed at the contribution of microRNAs (miRs) to the HPV life-cycle and HPV-induced carcinogenesis. A wide range of miRs have been found to have altered expression in the presence of HPV E6/E7, affecting a wide range of centrally important cellular signalling pathways [ 159 , 160 ], including those involved in control of cell growth [ [161] , [162] , [163] , [164] ] and apoptosis [ 165 , 166 ].…”

Transcriptome sequencing reveals pathways and genes dysregulated in HPV infected cervical cancer

“…While the mRNA levels of ATG13 and ULK1 encoding key factors in the autophagy pathway were also upregulated, though the activity of their protein products is negatively regulated by mTORC1 [29], revealing other factors may exist to regulate autophagy besides mTOR. Recently, a relevant study by Babion et al indicated that mTOR signaling was one of the three altered pathways in HPV-induced carcinogenesis, in which dysregulation was induced by the copy number changes of respective genes [30]. This provides a novel strategy to study the dysregulated expression of key genes in tumors.…”

Integrative meta‑analysis of gene expression profiles identifies FEN1 and ENDOU as potential diagnostic biomarkers for cervical squamous cell carcinoma

“…The diagnostic accuracy for each candidate transcript was assessed individually by constructing receiver operating characteristic (ROC) curves and calculating the area under the curve (AUC), using an independent cohort, GSE138080 (65). The cohort was chosen due to the balanced number of subjects in the respective groups.…”