Abstract 2665: SHOT-RNAs: A novel class of tRNA-derived functional RNAs expressed in hormone-dependent cancers

Abstract

Abstract Sex hormones and their receptors play critical roles in the genesis and progression of breast and prostate cancers. We here present that sex hormone signaling pathways promote the expression of specific tRNA halves termed Sex HOrmone-dependent TRNA-derived RNAs (SHOT-RNAs). Screenings using 94 cancer cell lines indicated that SHOT-RNAs are specifically and abundantly expressed in estrogen receptor-positive (ER+) breast cancer and androgen receptor-positive (AR+) prostate cancer. None of the ER− breast cancer, AR− prostate cancer, or other examined cancer cell lines from other tissues exhibited abundant expression of SHOT-RNAs. ER-dependent accumulation of SHOT-RNAs is not limited to a cell culture system, but it also occurs in luminal-type breast cancer patient tissues. The expression specificity allowed us to hypothesize that sex hormones and their receptors are the crucial molecular factors for SHOT-RNA generation. Our hypothesis has been confirmed by the following three observations: (1) siRNA-mediated reduction of ER or AR decreased SHOT-RNA levels, (2) hormone-depletion from the culture medium decreased SHOT-RNA levels, and (3) addition of the sex hormone, estrogen or androgen, to the medium increased SHOT-RNA levels. These results indicate the clear dependency of SHOT-RNA expressions on sex hormones and their receptors. SHOT-RNAs are produced from aminoacylated mature tRNAs by angiogenin (ANG)-mediated anticodon cleavage. Resultant 5′- and 3′-SHOT-RNAs, corresponding to 5′- and 3′-tRNA halves, bear 3′-terminal modifications, cyclic phosphate (cP) and amino acid, respectively. Thus, SHOT-RNAs would not be captured by standard RNA-seq methods because the modification inhibits adapter ligation steps. This could be the reason why SHOT-RNAs had not been discovered in spite of their abundance and expression specificity. To identify a comprehensive repertoire of SHOT-RNAs, we developed a method named “cP-RNA-seq” in which 3′-termini of all RNAs, except for those containing a 3′-terminal cP, were cleaved by a periodate treatment, thereby selectively amplifying and sequencing only cP-containing RNAs. The application of cP-RNA-seq to breast cancer cells identified eight cytoplasmic tRNA species as major sources of 5′-SHOT-RNAs. We further investigated the functional significance of SHOT-RNA expression. Intriguingly, siRNA-directed reduction of each one of the three different species of 5′-SHOT-RNA severely impaired cell proliferation, whereas 3′-SHOT-RNA reduction showed no impact. These results strongly suggest that at least 5′-SHOT-RNAs are expressed as functional RNA molecules promoting cell growth. Our results have unveiled a novel tRNA-engaged pathway in tumorigenesis of hormone-dependent cancers and implicate SHOT-RNAs as potential candidates for biomarkers and therapeutic targets. We will discuss our working model and future challenges. Citation Format: Shozo Honda, Megumi Shigematsu, Phillipe Loher, Juan P. Palazzo, Issei Imoto, Isidore Rigoutsos, Yohei Kirino. SHOT-RNAs: A novel class of tRNA-derived functional RNAs expressed in hormone-dependent cancers. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2665.