Somatic Mutations in miRNA Genes in Lung Cancer—Potential Functional Consequences of Non-Coding Sequence Variants

Abstract

A growing body of evidence indicates that miRNAs may either drive or suppress oncogenesis. However, little is known about somatic mutations in miRNA genes. To determine the frequency and potential consequences of miRNA gene mutations, we analyzed whole exome sequencing datasets of 569 lung adenocarcinoma (LUAD) and 597 lung squamous cell carcinoma (LUSC) samples generated in The Cancer Genome Atlas (TCGA) project. Altogether, we identified 1091 somatic sequence variants affecting 522 different miRNA genes and showed that half of all cancers had at least one such somatic variant/mutation. These sequence variants occurred in most crucial parts of miRNA precursors, including mature miRNA and seed sequences. Due to our findings, we hypothesize that seed mutations may affect miRNA:target interactions, drastically changing the pool of predicted targets. Mutations may also affect miRNA biogenesis by changing the structure of miRNA precursors, DROSHA and DICER cleavage sites, and regulatory sequence/structure motifs. We identified 10 significantly overmutated hotspot miRNA genes, including the miR-379 gene in LUAD enriched in mutations in the mature miRNA and regulatory sequences. The occurrence of mutations in the hotspot miRNA genes was also shown experimentally. We present a comprehensive analysis of somatic variants in miRNA genes and show that some of these genes are mutational hotspots, suggesting their potential role in cancer.

Pan-Cancer analysis of somatic mutations in miRNA genes

“…We also confirmed the recurrence of mutations in hsa-miR-21 as previously identified with the Annotative Database of miRNA Elements (ADmiRe) 48 . Not surprisingly, the current results overlap almost perfectly with our earlier results obtained for LUAD and LUSC 31 . Minor discrepancies result from some differences in the technical approach (see Materials and Methods).…”

“…An additional indication of miRNA gene sensitivity to genetic alteration is their general high conservation and the decreased density of common SNPs in miRNA hairpin sequences [28][29][30] , which resembles the commonly known phenomenon of the decreased level of nonsynonymous SNPs in protein-coding sequences. In our recent analysis of somatic mutations in lung cancers, we confirmed that seed mutations affect the vast majority of the predicted targets and showed that mutations in miRNA genes often alter the predicted structure of miRNA precursors 31 . miRNAs in cancers were also considered in the context of somatic mutations found in mRNAs that affect mRNA-miRNA and competing endogenous RNA (ceRNA)-miRNA interactions 32,33 .…”

“…Five of the mutations are located in a single hotspot position (chr1:209432167C>T[+]) that is the first position of the seed sequence of miR-205-5p (guide miRNA). As we have shown before, the mutation may substantially affect target recognition, disrupting 250/288 (87%) predicted miR-205-5p targets and creating 471 new targets 31 . The disrupted targets include many validated miR-205-5p targets, including the oncogenes VEGFA (mediator of angiogenesis) and E2F1…”

“…From the vcf.gz files, we extracted somatic mutation calls with PASS annotation. The extraction was performed with a set of in-house Python scripts (https://github.com/martynaut/pancancer_mirnome), an updated version of scripts used in our earlier research 31 . To avoid duplicating mutations detected in multiple sequencing experiments in the same cancer patient, we combined files summing reads associated with particular mutations.…”

“…Hsa-miR-664b is a highly validated (miRBase) and moderately conserved miRNA gene that substantially overlaps with the SNORA36A H/ACA box small nuclear RNA (snoRNA; snoRNABase) playing a role in the pseudouridylation of rRNAs and snRNAs; therefore, all mutations may also affect the function of snoRNA. In our previous study, we showed that some of the mutations, especially chrX:154768615G>T[+] and chrX:154768652C>A[+], cause serious structural aberrations 31 . miR-664b-5p was also recently shown to act as a cancer suppressor in hepatocellular cancer cell lines 96 .…”