GSE171729
Multiplexed functional genomic analysis of somatic 5' untranslated region mutations across the spectrum of human prostate cancer (RNASeq)
The functional consequences of genetic variants within 5' untranslated regions (UTRs) on a genome-wide scale are poorly understood in disease. We developed a high-throughput multi-layer massively parallel sequencing-based method called PLUMAGE (Pooled full-length UTR Multiplex Assay on Gene Expression) to quantify the molecular consequences of somatic 5' UTR mutations found across 229 prostate cancer patients with localized to metastatic disease. We show that 5' UTR mutations can control both transcript levels and mRNA translation rates through the creation of new DNA binding elements or RNA-based cis-regulatory motifs. We also discover that single point mutations can simultaneously impact both transcript levels and mRNA translation of the same gene. Using gene editing technology, we validate that a single point mutation in the oncogenic CKS2 5' UTR can increase mRNA specific translation. Turning to a molecular pathway critical for cancer, we provide evidence that functional 5' UTR mutations in the MAP kinase signaling pathway can upregulate pathway-specific gene expression and are associated with distinct clinical outcomes. Our study reveals the diverse mechanisms by which the mutational landscape of 5' UTRs can co-opt multiple levels of gene expression and demonstrates that single nucleotide alternations within leader sequences are functional in cancer. Overall design: Next-generation sequencing of metastatic castration-resistant prostate cancer (mCRPC) tumors using Illumina TruSeq Stranded mRNA Library prep.