Date of Award

5-1-2024

Language

English

Document Type

Master's Thesis

Degree Name

Master of Science (MS)

College/School/Department

Department of Biomedical Sciences

Dissertation/Thesis Chair

Pallavi Ghosh

Committee Members

Joseph T Wade, Janice D Pata

Subject Categories

Biology

Abstract

Mycobacterium abscessus is a non-tuberculous mycobacteria that causes lung infections or skin and soft tissue infections. M. abscessus is innately resistant to antibiotics, requiring a long multi-antibiotic treatment regimen, with a low 45% cure rate for lung infections. The WhiB-like transcription factors are limited to the actinomycete class, which includes Mycobacteria. These proteins regulate multiple cellular processes, including growth and division, responses to oxidative, nitrosative or pH stress, and antibiotic resistance. Our work describes three WhiB-like transcription factors: WhiB7, which regulates antibiotic resistance, WhiB2, with a role in cell division, and Mab1756, an unstudied regulator similar to WhiB2. We used ChIP-seq experiments to locate genomic binding sites of each transcription factor, as well as phenotypic studies to identify the roles of each in the bacterium.WhiB7 binds 56 locations on the M. abscessus genome, directly upregulating acetyltransferases, ribosome-associated proteins, transcriptional regulators, membrane proteins, and biosynthesis and metabolic proteins. WhiB7 binds to an A/T-rich DNA motif found about 23 or 33 bases upstream of the transcription start site. A few of the upregulated proteins showed novel antibiotic resistance effects, in the absence of WhiB7. Mab1756 binds 130 intergenic regions on the M. abscessus genome, WhiB2 binds 158 intergenic regions, and 98 of these regions are bound by both transcription factors. These binding sites likely regulate proteins involved in DNA replication and maintenance, the ribosome and translation, lipid and glycan metabolism, transcription factors and enzymes. Mab1756 binds to a CTTGAC motif, while WhiB2 binds the WhiA motif, GACAC. Cells overexpressing Mab1756 can have a longer length phenotype. The WhiB2 protein has highest expression in cells at mid-log phase, and under nutrient starvation, as was previously documented. This work indicates new roles for WhiB-like transcription factors in M. abscessus. Further studies on WhiB7 may elucidate functions of the biosynthetic or ribosome-associated proteins it upregulates. Further study of Mab1756 and WhiB2 may validate a role in cell division through cell wall remodeling, as well as further roles in DNA replication or translation.

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