Arsenic exposure is a global health problem. Millions of people encounter arsenic through contaminated drinking water, consumption, and inhalation. Specific to Arkansas is the presence of arsenic in locally grown rice and contaminated ground water from mining natural gas rich shale deposits. The arsenic response locus in budding yeast is responsible for the detoxification of arsenic and its removal from the cell. This locus constitutes a conserved pathway ranging from prokaryotes to higher eukaryotes. The goal of this dissertation was to identify how the arsenic response locus is regulated in an arsenic dependent manner.
CRISPR-Chromatin Affinity Purification with Mass Spectrometry (CRISPR-ChAP-MS) enrichment was increased by 20-fold compared to previously published results. Optimized CRISPR-ChAP-MS was applied to the arsenic response locus to identify what regulatory machinery must be present for activation of the locus. The use of FASP in combination with off-line bRP-UPLC fractionation identified protein complexes, SAGA and SWI/SNF, previously unattributed to the regulation of the arsenic response locus. Functional assays and arsenite sensitivity screens, identified the HAT activity of SAGA and the ATPase chromatin remodeling activity of SWI/SNF are required for activation of the arsenic response locus. 11 other identified proteins also exhibited both arsenite sensitive phenotypes and a reduction in ACR2 transcription that could each be further interrogated. The SWI/SNF and SAGA complexes have been implicated in cancer and may serve as biomarkers for effectiveness of arsenite trioxide treatment and therapeutic targets.
|Advisor:||Tackett, Alan J.|
|Commitee:||Abbott, Karen L., Raney, Kevin D., Taverna, Sean D., Wahls, Wayne P.|
|School:||University of Arkansas for Medical Sciences|
|Department:||Biochemistry and Molecular Biology|
|School Location:||United States -- Arkansas|
|Source:||DAI-B 79/10(E), Dissertation Abstracts International|
|Subjects:||Molecular biology, Biochemistry, Environmental science|
|Keywords:||Arsenic, Chromatin, Mass spectrometry|
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