Abstract A29: Gender-specific and race-specific regulatory noncoding RNAs are prevalent in healthy and in cancerous tissues

In this study we show that the cellular profiles of microRNA (miRNA) isoforms and of transfer RNA fragments (tRFs) depend strongly on a person’s gender, race, and population-membership as well as on tissue, disease, and disease subtype. MiRNAs and tRFs are two of many known categories of regulatory non-protein-coding RNAs (ncRNAs) that are expressed in a wide spectrum of cell types and regulate numerous processes in health and disease. Our findings suggest that we need to explicitly incorporate these molecules in all future studies of the post-transcriptional regulatory mechanisms that underlie the onset and progression of cancers. MiRNAs were originally discovered more than 20 years ago and have since been linked to development, homeostasis, and many human conditions including cancers. For many years, each miRNA precursor molecule was thought to produce a single mature miRNA (“archetype”) from one or both of its two arms. The archetype miRNA sequences for the various miRNA loci were collected in curated public repositories. It is now known that a miRNA precursor arm can produce multiple isoforms at the same time. These isoforms are known as “isomiRs,” have distinct endpoints and relative abundances, and enter the RNA interference (RNAi) pathway through loading on the Argonaute silencing complex. Analysis of transcriptomes from hundreds of healthy individuals from the 1,000 Genomes Project (1KG) and hundreds of normal breast and breast cancer (BRCA) samples from The Cancer Genome Atlas (TCGA) repository allowed us to show that attributes such as a person’s gender, population, and race modulate the active population of isomiRs in a tissue. Tissue type, tissue state, and disease subtype also modulate a cell’s isomiR profile. Interestingly, we found that co-expressed isomiRs from the same miRNA locus are generally not well correlated. Gene expression analyses following over-expression experiments of isomiRs from the same cancer-linked miRNA precursor arm showed effectively non-overlapping targetomes, despite the shared origin of the tested isomiRs. Similarly to the case of miRNAs, the genomic loci coding for transfer RNAs (tRNAs) were believed to only produce the mature clover-leaf-shaped tRNA that is a key ingredient of the translation of mRNAs into amino acid sequences. It is now known that tRNA transcripts will also give rise to tRNA fragments, the tRFs, which co-exist with the corresponding full-length mature tRNA. Individual tRFs have been linked to cell growth, cell proliferation, response to DNA damage, translation initiation, response to stress, etc. In complete analogy to the isomiRs, some tRFs have been shown to load on Argonaute and thus to also participate in RNAi. By studying the same transcriptomic datasets from the 1KG and TCGA projects mentioned above, we showed that the composition and abundance profiles of tRNA fragments depend on a person’s gender, population, race, tissue type, tissue state, and disease subtype. Notably we showed that tRFs are produced from both nuclear and from mitochondrial tRNAs and have distinct length profiles that are characteristic of the tRF source (nucleus vs. mitochondrion) and of the tissue at hand. Our studies also uncovered a new category of tRFs, the i-tRFs, that are wholly internal to the respective mature tRNA, straddle the anticodon, and contribute most of the differences observed across tissues, genders, populations, and races. Using two different fragment-specific PCR techniques we validated two i–tRF molecules in 21 of 22 tested breast tumor and adjacent normal samples, and in eight breast cancer cell lines. Our isomiR and tRF results show that the molecular signatures of human tissues, in health and disease, comprise many previously unsuspected functional non-coding RNAs. The newly identified molecules have unexpected complex dependencies on patient attributes as well as on tissue and disease. Citation Format: Isidore Rigoutsos, Aristeidis Telonis, Phillipe Loher, Eric Londin. Gender-specific and race-specific regulatory noncoding RNAs are prevalent in healthy and in cancerous tissues. [abstract]. In: Proceedings of the Fourth AACR International Conference on Frontiers in Basic Cancer Research; 2015 Oct 23-26; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2016;76(3 Suppl):Abstract nr A29.