I studied three biological problems in my dissertation research. The problems involved flow of information into the cells from outside, the regulation of information flow by the ribosomes in protein synthesis, and the disruption of information flow due to microsatellite repeat expansions leading to a human disease myotonic dystrophy. In the first study, I built a conceptual basis for interpreting and understanding the cellular responses to multiple concurrent stimuli. A gene represents the inherent information of the cells while a stimulus represents the information outside their boundary. Since a gene and a stimulus are both packets of information, they can be considered analogues. Therefore, the concepts of gene interactions can be applied to the study of modulation of cellular processes by stimuli. This assumption allowed me to define the concepts of environmental interactions and environmental epistasis in terms of gene interactions and genetic epistasis. I used proteomic and transcriptomic changes in Saccharomyces cerevisiae to test the conceptual framework. In the second study, I designed and performed experiments to test the ribosome filter hypothesis. The ribosome filter hypothesis says that the amount of information flow from a transcript to a protein is regulated by the compositions of the subpopulations of ribosomes in a cell. The composition of a ribosome determines its interactions with the mRNA and accessory factors, which in turn determine the efficiency of translation of a transcript. Therefore, to efficiently translate the proteome required for growing in one environmental condition would require a specific complement of ribosomes with different compositions. The required complement of ribosomes will be different for a cell growing in a different environmental condition. A difference in the protein composition of ribosomes from cells growing in two different conditions would be evidence supporting the ribosome filter hypothesis. It would allow identification of candidate ribosomal proteins, or their post-translation modifications that regulate information flow from specific transcripts. I used growth of S. cerevisiae with fermentable carbon source, glucose, and non-fermentable carbon source, glycerol, as two conditions. I used iTRAQ labeling based quantitative proteomics as well as, in collaboration with the Joachim Frank lab, cryo-electron microscopy to measure the changes in protein composition of ribosomes. I used yeast genetics and polysome profiling to measure the effect of loss of function of a candidate ribosomal protein, Rpl8a or Rpl8b, on translation. In the third project, I studied the changes introduced in the skeletal muscle proteome of myotonic dystrophy patients, both type 1 and 2, due to the disruption of information flow by microsatellite repeat expansions in the non-coding regions of mRNA transcripts. I used iTRAQ labeling based quantitative proteomics analysis to quantitate the changes in the skeletal muscle proteome of DM patients compared to healthy volunteers. I identified differentially present proteins and used pathway analysis to understand their role in the pathogenesis. I have identified a number of candidate proteins that are interesting targets for more in depth genetic and biochemical studies including a ribosomal protein RPL13A, previously implicated in regulating information flow by translational inhibition of transcripts containing the GAIT sequence motif. In summary, I have studied three different ways the information content of cells and tissues are affected.
|Advisor:||Link, Andrew J.|
|Commitee:||Link, Andrew J., Ohi, Melanie D., Reiter, Nicholas, Schey, Kevin L., Tansey, William P.|
|School Location:||United States -- Tennessee|
|Source:||DAI-B 80/07/(E), Dissertation Abstracts International|
|Subjects:||Molecular biology, Biochemistry, Bioinformatics|
|Keywords:||Environmental epistasis, Environmental interactions, Myotonic dystrophy, Ribosome filter hypothesis, Specialized ribosome, Translational control|
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