Telomeric RNAs as a novel player in telomeric integrity

Telomeres protect linear chromosome ends from being recognized and processed as double-strand breaks by DNA repair activities. This protective function of telomeres is essential for chromosome stability. Until recently, telomeres have been considered to be transcriptionally silent. This notion was overturned in a series of recent papers that describe the existence of telomeric repeat-containing RNAs (TERRAs) in vertebrates and yeast. Here, we summarize recent developments in this field of telomere research, in particular the possible mechanisms that control TERRA expression

A Yeast Model of FUS/TLS-Dependent Cytotoxicity

FUS/TLS is a nucleic acid binding protein that, when mutated, can cause a subset of familial amyotrophic lateral sclerosis (fALS). Although FUS/TLS is normally located predominantly in the nucleus, the pathogenic mutant forms of FUS/TLS traffic to, and form inclusions in, the cytoplasm of affected spinal motor neurons or glia. Here we report a yeast model of human FUS/TLS expression that recapitulates multiple salient features of the pathology of the disease-causing mutant proteins, including nuclear to cytoplasmic translocation, inclusion formation, and cytotoxicity. Protein domain analysis indicates that the carboxyl-terminus of FUS/TLS, where most of the ALS-associated mutations are clustered, is required but not sufficient for the toxicity of the protein. A genome-wide genetic screen using a yeast over-expression library identified five yeast DNA/RNA binding proteins, encoded by the yeast genes ECM32, NAM8, SBP1, SKO1, and VHR1, that rescue the toxicity of human FUS/TLS without changing its expression level, cytoplasmic translocation, or inclusion formation. Furthermore, hUPF1, a human homologue of ECM32, also rescues the toxicity of FUS/TLS in this model, validating the yeast model and implicating a possible insufficiency in RNA processing or the RNA quality control machinery in the mechanism of FUS/TLS mediated toxicity. Examination of the effect of FUS/TLS expression on the decay of selected mRNAs in yeast indicates that the nonsense-mediated decay pathway is probably not the major determinant of either toxicity or suppression.Fidelity Biosciences (Firm)Fidelity Biosciences (Firm) (Research Inititative)ALS Therapy AllianceNational Institutes of Health (U.S.) (NIH 1RC1NS06839)National Institutes of Health (U.S.) (NIH U01NS05225-03)National Institutes of Health (U.S.) (NIH R01NS050557-05)National Institutes of Health (U.S.) (NIH 1RC2NS070342-01)Pierre L. de Bourgknecht ALS Research FoundationNational Science Foundation (U.S.) (NS614192

Investigation of an Evolutionarily Conserved Staufen-mediated mRNA Decay Pathway that Involves SINEs and lncRNAs

Thesis (Ph.D.)--University of Rochester. School of Medicine & Dentistry. Dept. of Biochemistry and Biophysics, 2014.The mammalian double stranded (ds)RNA-binding protein Staufen (STAU)1, is a multi-functional protein that regulates gene expression at different levels. One example is STAU1-mediated mRNA decay (SMD). STAU1, when bound to a STAU1-binding site (SBS) in the 3'-untranslated region (3'UTR) of mRNAs, functions in complex with UPF1 to elicit the decay of targeted mRNAs in a way that depends on mRNA translation. Human SBSs can be formed by intramolecular or intermolecular base-paring, the latter of which is between an mRNA 3'UTR Alu element and a partially complementary Alu element within long noncoding RNAs (lncRNAs) called ½-sbsRNAs. Alu elements are a type of short interspersed element (SINE). Even though Alu elements are confined to primates, I have shown that SINE-dependent intermolecular SBS formation is not. My computational analyses demonstrated that 13.2% of annotated mouse mRNAs contain a single 3'UTR SINE, and 28.4% of known mouse lncRNAs contain one or more SINEs. Further experiments provide evidence that SMD occurs in mouse cells via partially complementary mRNA−lncRNA base-pairing. I have shown the physiological relevance of these interactions by demonstrating that mouse (m)½-sbsRNA-triggered SMD regulates C2C12-cell myogenesis. I undertook a series of computational analyses to pursue the mechanistic similarity between SINE- and ½-sbsRNA-dependent SMD in human and mouse cells despite the distinct evolutionary origins of their SINEs. These analyses demonstrate that 4.3% of the orthologous genes in human and mouse contain at least one 3'UTR SINE, a commonality of which indicates that SINEs have been positively selected for after integration. Subsequent deep sequencing of RNA in human and mouse skeletal muscle cell lines identified mRNA orthologs whose levels are elevated upon downregulation of STAU1 and, independently UPF1. Statistical analyses confirmed that the percentage of 3'UTR SINE-containing transcripts in orthologous SMD target pairs is significantly higher than that in the sum total of orthologous mRNA pairs. This indicates that SMD contributes, at least in part, to the selection and/or maintenance of SINEs in mRNA 3'UTRs. We speculate that such a selection was favored due to its involvement in conserved functions, which implies a convergent evolution of 3'UTR SINEs mediated by SMD

mRNA-mRNA duplexes that autoelicit Staufen1-mediated mRNA decay

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