Mitochondrial tRNA-Derived Fragments as Candidate Metastasis-Modifying RNA.

How mitochondrial DNA (mtDNA) polymorphisms influence complex phenotypes remains poorly understood. Using mitochondrial-nuclear exchange mice, we previously showed that mtDNA single-nucleotide polymorphisms (SNP) modify metastasis, cardiovascular disease, and epigenetic marks independently of metabolic differences. The only mtDNA SNP correlating with these phenotypes resides in the gene encoding mitochondrial transfer RNA (tRNA)-arginine [mt-tRNAArg (UCG), mt-TR], suggesting a role for non-protein-coding loci. In this study, we identify and preliminarily characterize previously undescribed tRNA-derived fragments (tRF) generated from mt-TRs. Northern blotting revealed distinct tRF that are differentially expressed among mtDNA SNPs, between lung and liver, and between sexes. Surprisingly, small RNA sequencing of untreated RNA did not detect the same tRFs in high abundance. However, demethylating and restoring 5'-OH and 3'-PO4 termini allowed detection of sequences consistent with the northern blot bands. Enforcing exact matching to the mitochondrial genome and normalizing to their parental molecule revealed putative tRF sequences with shared cleavage sites. Based on connections among mtDNA SNPs, the resulting SNP-dependent tRF, and SNP-metastasis correlation, we propose that these tRFs may function as metastasis modifiers. These data also expand the functional output of the mitochondrial genome that can contribute to phenotype modification. SIGNIFICANCE: This study describes identification and initial characterization of mitochondrial tRNA fragments that are suspected to regulate metastasis efficiency.