Browsing College of Computer, Mathematical & Natural Sciences by Subject "-1 PRF"
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- ItemCharacterization of programmed -1 ribosomal frameshift signals in the interleukin 2 receptor gamma mRNA(2016) Flickinger, Zachary Flickinger; Dinman, Jonathan D; Biology; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Programmed -1 ribosomal frameshift (-1 PRF) signals are predicted to be present in ~10% of eukaryotic mRNA, suggesting a conserved role for -1 PRF in eukaryotic mRNA translation. This work focuses on -1 PRF in the interleukin 2 receptor gamma (IL2RG) mRNA. IL2RG is a component of receptors for six cytokines responsible for lymphocyte proliferation and differentiation. Altered expression of IL2RG is linked to immunodeficiency and lymphoma. We verified that the IL2RG mRNA has one potential -1 PRF signal in exon 3 that stimulates -1 frameshifting and a second in exon 8 that is definitively a -1 PRF signal. Both of these signals redirect the ribosome to a premature termination codon, suggesting -1 PRF alters IL2RG expression via NMD. Testing the effect of cancer patient associated mutations discovered in the exon 8 -1 PRF signal may elucidate a role for IL2RG -1 PRF in normal physiology and pathological phenotypes.
- ItemESTABLISHING LINK BETWEEN TRANSLATIONAL RECODING AND HUMAN DISEASE(2015) Advani, Vivek Manoharlal; Dinman, Jonathan D; Cell Biology & Molecular Genetics; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Gene expression can be controlled at the level of mRNA stability, and prior studies from our laboratory have explained how Programmed -1 Ribosomal Frameshifting (-1 PRF) fits within this paradigm. Computational analyses suggest that 10-15% of eukaryotic mRNAs contain at least one potential -1 PRF signal. The overwhelming majority of predicted "genomic" -1 PRF events are predicted to direct translating ribosomes to premature termination codons. We have demonstrated that these can function as mRNA destabilizing elements through the Nonsense-Mediated mRNA Decay (NMD) pathway. In published work we have explored the biological significance of the connection between -1 PRF and NMD on telomere maintenance in yeast. More recently we extended this line of inquiry to human cells, demonstrating that a sequence element in the mRNA encoding Ccr5p harbors a -1 PRF signal which functions as an mRNA destabilizing element through NMD. In the current work we are exploring the link between global changes in -1 PRF rates and human health using yeast and human cell-based models of two diseases, X-linked Dyskeratosis Congenita (X-DC) and Spinocerebellar ataxia 26 family (SCA26) as models. Preliminary findings suggest these genetically inherited defects result in translational fidelity defects (i.e. changes in rates of -1 PRF, +1 PRF, and stop codon recognition), with attendant effects on mRNA abundance, gene expression and telomere maintenance. These studies establish a paradigm for understanding the linkage between translational fidelity and human disease.
- ItemTHE IMPACT OF TRANSLATIONAL FIDELITY ON HUMAN HEALTH(2018) Marques dos Santos Vieira, Carolina; Dinman, Jonathan D; Cell Biology & Molecular Genetics; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Ribosomopathies are a class of diseases resulting from mutations in genes encoding ribosomal proteins and ribosome biogenesis factors. Pleiotropic clinical presentations of different ribosomopathies has been taken as evidence of specialized ribosomes. Alternatively, gene dosage effects have been proposed to account for the observed differences. A yeast genetics approach was used to address this issue. Due to a historical gene duplication event, S. cerevisiae cells harbor two ohnologs for most ribosomal proteins. Deletion of one yeast ribosomal protein ohnolog was used to mimic haploinsufficiency in diploid cells (i.e. pseudo-haploinsufficient yeast). Further, insertion of a second copy of the undeleted ohnolog into the locus of the deleted ohnolog enabled separation of effects due to gene dosage from those due to ribosomal protein ohnolog identity. We found that significant changes in translational fidelity in the ribosomal protein ohnolog deletion strains were corrected by ohnolog duplication. Changes in gene dosage, particularly as they may affect the abundance of an enzyme as central as the ribosome, can impart stress through far reaching effects on cellular metabolism. Thus, as an orthogonal approach, we also examined the stress profiles of cells harboring the cbf5-D95A allele (model of X-linked Dyskeratosis Congenita) and the rps23a-R69K allele (model of MacInnes Syndrome). RNA-seq analysis revealed increased expression of proteins involved in response to oxidative stress in cbf5-D95A cells. Growth curve analysis revealed a longer plateau of the cbf5-D95A cells upon reaching stationary phase, suggestive of a pre-adaptive stress response. Decreased ROS abundance, tunicamycin resistance and increased basal levels of HAC1 mRNA in the mutant cells support this hypothesis. Similar results were observed with regard to the ohnolog deletion strains. Taken as a whole, these data support the gene dosage model as opposed to the specialized ribosome hypothesis with the caveat that this conclusion is limited to yeast cells growing in rich medium.