Human cells contain myriad excised linear intron RNAs with links to gene regulation and potential utility as biomarkers

• Published in: bioRxiv
• Main Authors: Yao, Jun, Winans, Shelby, Xu, Hengyi, Wu, Douglas C, Ferrick-Kiddie, Elizabeth A, Ares, Manuel, Lambowitz, Alan M
• Format: Paper
• Language: English
• Published: Cold Spring Harbor Cold Spring Harbor Laboratory Press 09.03.2022; Cold Spring Harbor Laboratory
• Edition: 1.5
• Subjects: Alternative splicing; Argonaute 2 protein; Binding sites; Biomarkers; Breast cancer; Cellular stress response; Chromatin remodeling; Enzymes; Gene regulation; Intellectual property; Introns; miRNA; Molecular Biology; Patent applications; Proteins; Ribonucleic acid; RNA; RNA-binding protein; RNA-directed DNA polymerase; Transcription factors; Tumor cell lines; United States > US; Texas; ribosomal proteins; mitochondrial function; ribosome assembly; gene regulation; RNA splicing
• ISSN: 2692-8205; 2692-8205
• DOI: 10.1101/2020.09.07.285114

By using TGIRT-seq, we identified >8,500 short full-length excised linear intron (FLEXI) RNAs in human cells. Subsets of FLEXIs accumulated in a cell-type specific manner, and ~200 corresponded to agotrons or mirtrons or encoded snoRNAs. Analysis of CLIP-seq datasets identified potential interactions between FLEXIs and >100 different RNA-binding proteins (RBPs), 53 of which had binding sites in ≥30 different FLEXIs. In addition to proteins that function in RNA splicing, these 53 RBPs included transcription factors, chromatin remodeling proteins, and cellular growth regulators that impacted FLEXI host gene alternative splicing and/or mRNA levels in knockdown datasets. We computationally identified six groups of RBPs whose binding sites were enriched in different subsets of FLEXIs: AGO1-4 and DICER associated with agotrons and mirtrons; AATF, DKC1, NOLCI, and SMNDC1 associated with snoRNA-encoding FLEXIs; two different combinations of alternative splicing factors found in stress granules; and two novel RBP-intron combinations, one including LARP4 and PABC4, which function together in the cytoplasm to regulate ribosomal protein translation. Our results suggest a model in which proteins involved in transcriptional regulation, alternative splicing, or post-splicing secondary functions bind and stabilize cell-type specific subsets of FLEXIs that perform different biological functions and have potential utility as biomarkers. Competing Interest Statement Thermostable group II intron reverse transcriptase (TGIRT) enzymes and methods for their use are the subject of patents and patent applications that have been licensed by the University of Texas and East Tennessee State University to InGex, LLC. A.M.L., some former and present members of the Lambowitz laboratory, and the University of Texas are minority equity holders in InGex and receive royalty payments from the sale of TGIRT enzymes and kits and from the sublicensing of intellectual property by InGex to other companies. A.M.L., J.Y., H.X. and D.C.W. are inventors on a patent application filed by the University of Texas at Austin for the use of full-length excised intron RNAs and intron RNA fragments as biomarkers. S.W., E.A.F.K. and M.A. have no competing interests.