Target-site Dynamics and Alternative Polyadenylation Explain a Large Share of Apparent MicroRNA Differential Expression

Abstract

MicroRNA (miRNA) abundance reflects a dynamic balance between biogenesis, target engagement, and decay, yet differential expression analyses typically ignore changes in target-site availability driven by alternative polyadenylation (APA). We introduce MIRNAPEX, an expression-stratification-based machine learning framework that quantifies miRNA regulatory effect sizes from RNA-seq data by integrating target-gene expression with 3'UTR isoform usage to infer effective binding-site dosage. Using pan-cancer training sets, we train models that learn relationships between transcriptomic features and miRNA log-fold changes, with APA patterns providing predictive information beyond gene expression alone. When applied to knockdowns of core APA regulators, MIRNAPEX captured widespread 3'UTR shortening and accurately anticipated miRNA-specific shifts whose direction and magnitude mirrored APA-driven changes in binding-site availability. Analysis of target-directed miRNA degradation interactions further showed that loss of distal decay-trigger sites coincides with increased miRNA abundance, consistent with reduced degradation. Together, these findings demonstrate that apparent miRNA differential expression can arise from dynamic target-site landscapes rather than altered miRNA transcription, and that neglecting this dimension can lead to misestimation of regulatory effect sizes.