Spliceosome-Associated microRNAs Signify Breast Cancer Cells and Portray Potential Novel Nuclear Targets.

MicroRNAs (miRNAs) act as negative regulators of gene expression in the cytoplasm. Previous studies have identified the presence of miRNAs in the nucleus. Here we study human breast cancer-derived cell-lines (MCF-7 and MDA-MB-231) and a non-tumorigenic cell-line (MCF-10A) and compare their miRNA sequences at the spliceosome fraction (SF). We report that the levels of miRNAs found in the spliceosome, their identity, and pre-miRNA segmental composition are cell-line specific. One such miRNA is miR-7704 whose genomic position overlaps HAGLR, a cancer-related lncRNA. We detected an inverse expression of miR-7704 and HAGLR in the tested cell lines. Specifically, inhibition of miR-7704 caused an increase in HAGLR expression. Furthermore, elevated levels of miR-7704 slightly altered the cell-cycle in MDA-MB-231. Altogether, we show that SF-miR-7704 acts as a tumor-suppressor gene with HAGLR being its nuclear target. The relative levels of miRNAs found in the spliceosome fractions (e.g., miR-100, miR-30a, and let-7 family) in non-tumorigenic relative to cancer-derived cell-lines was monitored. We found that the expression trend of the abundant miRNAs in SF was different from that reported in the literature and from the observation of large cohorts of breast cancer patients, suggesting that many SF-miRNAs act on targets that are different from the cytoplasmic ones. Altogether, we report on the potential of SF-miRNAs as an unexplored route for cancerous cell state.

Small RNA sequences derived from pre-microRNAs in the supraspliceosome.

MicroRNAs (miRNAs) are short non-coding RNAs that negatively regulate the expression and translation of genes in healthy and diseased tissues. Herein, we characterize short RNAs from human HeLa cells found in the supraspliceosome, a nuclear dynamic machine in which pre-mRNA processing occurs. We sequenced small RNAs (<200 nt) extracted from the supraspliceosome, and identified sequences that are derived from 200 miRNAs genes. About three quarters of them are mature miRNAs, whereas the rest account for various defined regions of the pre-miRNA, and its hairpin-loop precursor. Out of these aligned sequences, 53 were undetected in cellular extract, and the abundance of additional 48 strongly differed from that in cellular extract. Notably, we describe seven abundant miRNA-derived sequences that overlap non-coding exons of their host gene. The rich collection of sequences identical to pre-miRNAs at the supraspliceosome suggests overlooked nuclear functions. Specifically, the abundant hsa-mir-99b may affect splicing of LINC01129 primary transcript through base-pairing with its exon-intron junction. Using suppression and overexpression experiments, we show that hsa-mir-7704 negatively regulates the level of the lncRNA HAGLR. We claim that in cases of extended base-pairing complementarity, such supraspliceosomal pre-miRNA sequences might have a role in transcription attenuation, maturation and processing.