Dysregulation of mRNA expression by miR-186 overexpression in arsenic-induced skin carcinogenesis

Dysregulated miRNA expression contributes to development of arsenic-induced cutaneous squamous cell carcinoma (cSCC). hsa-miR-186 (called “miR-186” henceforth) is overexpressed in arsenical cSCC tissues as well as in preclinical cell line model of arsenical cSCC. Simultaneous miR-186 overexpression and chronic inorganic trivalent arsenite (iAs; 100 nM) exposure transformed human HaCaT cell line preferentially over miR-186 exposure or iAs exposure alone. Both iAs and miR-186 regulate the expression of wide range of mRNA targets. However, how their interaction impact the transcriptome-wide mRNA expression landscape ushering in cancer is unknown. We performed longitudinal RNA-seq analysis in passage-matched HaCaT cells clones (±miR-186 overexpression) with simultaneous chronic iAs exposure (0/100 nM; 29 weeks) at 12 and 29 weeks. We determined the impact of each factor and their interaction towards differential gene expression and pathways dysregulation employing to different statistical approaches (t-statistic and 2-factor ANOVA). We show that a core set of pathways are dysregulated deterministically irrespective of the statistical approach chosen, possibly representing necessary changes for transformation. Our data suggests that each clonal line could take unique routes to dysregulate these core set of pathways necessary for transformation, highlighting the possible role of stochasticity in cancer development. We provide evidence to sift the strengths of weaknesses of each statistical methodology in providing biological understanding of events that play crucial role in carcinogenesis in large datasets with multiple contributing variables.