Browsing by Subject "MicroRNA"
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Item Energy balance effects on microRNA expression in a mouse model of pancreatic cancer(2010-12) Goldberg, Jason Asher; Lashinger, Laura M.; Hursting, Stephen D.Pancreatic cancer is the fourth leading cause of cancer death in the United States, with a five-year survival rate under 5%. Given the disease’s deadliness, increasing our understanding of the molecular nature of the pancreatic cancer is key to developing more effective preventive measures and treatments. Dietary energy restriction (DER) has been shown to have potent anticancer effects in pancreatic cancer, but the mechanism of action has yet to be completely elucidated. Here we investigate the potential of altered microRNA expression as a mechanism by which DER exerts its anticancer effect. Using the Exiqon microRNA Array, we identified several microRNAs of interest for further study. This includes microRNA (mir) 669c, a known regulator of glutathione-S transferases (linked to carcinogen metabolism and oxidative stress) that increases with age. To our knowledge, this is the first exploration of the effects of DER (which is known to suppress oxidative stress and other processes associated with aging and cancer) on microRNA expression. These findings may provide the initial steps towards identifying novel targets for pancreatic cancer prevention or treatment.Item Functionality and efficiency improvement of miRCheck, a popular program for microRNA structure prediction(2014-12) Bandyopadhyay, Parika; Roux, Stanley J.Plant and animals both contain non-coding small RNAs that play important roles in their growth, development and responses to biotic and abiotic stresses. MicroRNAs are short (20-24nt), endogenously expressed, and a well characterized class of small RNAs, which are derived from processing longer, hairpin-like precursors. Discovery of most miRNAs relies upon either of two methods: i) molecular cloning of small RNAs or ii) prediction of miRNA genes based on conserved sequences, and secondary structures of known miRNAs using computational tools. miRCheck is a popular computational tool chain comprised of various Perl scripts for identifying and profiling plant miRNA genes. The program serves two purposes: 1. Identifying miRNA homologs in target genomic or cDNA sequences using a given small RNA library, 2. Searching for all potential miRNA precursor loci across the target genome based on evolutionarily-conserved structural features without any reference small RNA library to compare with. miRCheck builds upon several popular tools like patscan, RNAfold, einverted, and connects them to provide a complete tool chain for identifying miRNAs. Although miRCheck is a very well designed tool chain, it still has a few issues that need to be addressed to enhance its functionality and efficiency. This work analyzes the working mechanism of miRCheck, proposes some methods to enhance its efficiency and functionality, and implements those in a modified tool chain, py-miRCheck, in Python. To process a long genome sequence, miRCheck looks at small segments and serially evaluates them leading to long run times. Even in a high performance computing node, it takes days to process a standard sized reference genome obtained from NCBI repository. It highlights the inefficiency in the program. On the functionality side, there are several issues that need to be addressed for usability improvements: i) lack of parameterized design, ii) procedural design, iii) lack of GUI interface for running the tool chain, and iv) deployment-related problems. In this work, I address all these areas and also parallelize the tool chain to improve its efficiency by over a factor of 3. I also provide a Django-based prototype web front end to submit queries on a genome sequence. In summary, this work improves the usability of this tool chain to a great extent.Item The role of a viral microRNA and RNA interference during viral replication in mammalian cells(2012-12) Seo, Gil Ju; Sullivan, Christopher S.RNA interference (RNAi) is an evolutionarily conserved process that regulates gene expression. Host cells and viruses interact in many ways, including through miRNAs and RNAi. Viral miRNAs are encoded when viruses, specially including the the polyoma and herpes families, are transcribed in the nucleus. Some viral miRNAs function to regulate host or viral gene expression. Most viral miRNAs’ functions are not known, however, in great detail. A miRNA can be encoded late during infection, as it is by SV40, a model polyomavirus. This downregulates early viral gene expression by directing mRNA RISC-mediated cleavage. As more polyomaviruses are discovered that are associated with human disease, it becomes more important to understand their function and to uncover whether these emerging viruses encode miRNAs. The work presented here shows the discovery of several viral miRNAs in human polyomaviruses—JCV, BKV, and MCV. In addition, I found that viral miRNAs have the evolutionarily conserved function of negatively regulating viral early gene transcripts at a late stage in the infection. During viral replication, viruses utilize the miRNA components of RNAi. However, in invertebrate organisms RNAi also actively defends against viral infection. It is still being debated, though, whether RNAi plays an antiviral role in mammalian cells. Should it be true that RNAi is an antiviral response in mammalian cells, then what is predicted by such a scenario is inconsistent with my studies. I have found that RNAi is strongly inhibited in the early stages after viral infection. Studies with a chemical mimic of viral infection (poly I:C) imply that the innate cellular immune response is responsible for this inhibition. I investigated the molecular changes, in response to viral infection, (e.g. poly ADP-ribosylation of Ago2) in the RNA-induced silencing complex (RISC). I determined that the inhibition of RNAi is brought about by components of the innate response. Completion of this study details a previously unknown “cross talk” between RNAi and the host innate immune response in mammalian cells. Furthermore, I found mir-17 family attenuates a subclass of interferon-stimulated genes. An understanding of viral miRNA and RNAi offers a clue as to we can use molecular intervention for viral infections.