Award Date

5-1-2020

Degree Type

Thesis

Degree Name

Master of Science (MS)

Department

Life Sciences

First Committee Member

Eduardo Robleto

Second Committee Member

Helen Wing

Third Committee Member

Boo Shan Tseng

Fourth Committee Member

Ronald Gary

Number of Pages

68

Abstract

Until the late 1980’s it was widely understood that bacterial variance emerges randomly during growth. Research that followed has convincingly shown evidence for mutations arising in non-growing conditions, a phenomenon known as stationary-phase mutagenesis. In Bacillus subtilis, an elegant mutagenic mechanism has been elucidated in non-growing cells that biases mutations to transcribed regions of a subpopulation. One interesting possibility is that mutations can be further biased to hotspots within genes through alternate DNA structures known as non-B DNA. Non-B DNA have been linked to genomic instability and disease in humans, lesser is known about its role in bacteria. Here we investigate if G4 DNA, a type of non-B DNA, are mutagenic hotspots in non-growing B. subtilis cells. We hypothesize that G4 DNA can block RNA polymerase and trigger gratuitous transcription coupled repair. Gratuitous repair, or repair occurring in the absence of DNA damage, can lead to mutagenesis via repetitive re-synthesis of DNA, which can introduce mutations. In order to test this hypothesis, we constructed strains differing in their ability to form G4 DNA in a gene of interest and measured the effect on mutagenesis. We found that a strain having the potential to form G4 DNA in the coding strand had the highest levels of mutagenesis and this effect was dependent on a transcription coupled repair factor Mfd. Our data adds to the evidence of how B. subtilis avoids genetic load by having an elegant mechanism that biases mutations to distinct regions of genes under selection. Further, elucidating how alternate DNA structures promote genetic instability can lead to a better understanding of bacterial evolution and genetic diseases in humans.

Keywords

Bacillus subtilis; DNA mutagenesis; G4 DNA; Stationary phase mutagenesis; Transcription coupled repair

Disciplines

Biology | Microbiology

File Format

pdf

File Size

0.26 MB

Degree Grantor

University of Nevada, Las Vegas

Language

English

Rights

IN COPYRIGHT. For more information about this rights statement, please visit http://rightsstatements.org/vocab/InC/1.0/

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