Investigating Protein-Protein Interactions between Bacteriophage Cain and its Host Mycobacterium smegmatis
Session Title
Final Oral Competition
Faculty Mentor
Victoria J. Frost, Ph.D
College
College of Arts and Sciences
Department
Biology
Abstract
In the face of the antibiotic resistance crisis, bacteriophages have proven to be useful tools for overcoming bacterial defenses in medicine. However, 65% of phage genes have no predicted function, which limits the ability to utilize phages to their maximum potential. In order to effectively manipulate and apply phages therapeutically, we must close this gap in functional knowledge. After using a phenotypic assay to screen mycobacteriophage Cain gene products for interaction with the host cell, Mycobacterium smegmatis, a bacterial 2-hybrid (B2H) assay was used to identify which host proteins were being targeted by phage gene products. To achieve this, a complex array of specifically designed expression vectors, test promoters and cell lines, were used to report potential pairwise interactions between phage gene products and the host’s proteome. With no predicted function, and significant homology with well-studied phages in the same K cluster, Cain gp55 was investigated first. Results revealed interactions with host proteins NusA and GntR which are both transcriptional regulators thought to play various roles including transcript elongation/termination (NusA), cell motility, metabolism, and even virulence (GntR). Another phage gene of no predicted function, Cain gp2, also interacted with several host cell transcriptional regulators, including TetR, AraC, and LysR, thought to be involved in the regulation of metabolic genes. Identifying these interactions can give us hints about a phage gene’s role in manipulating its host. While the B2H assay gives us a starting point, more specific experimentation is needed to identify potentially new and useful functions of undescribed genes.
Previously Presented/Performed?
Winthrop University Showcase of Undergraduate Research and Creative Endeavors, Rock Hill, SC, April 2023.
Type of Presentation
Oral presentation
Grant Support?
Supported by an SC-INBRE grant from the National Institute for General Medical Sciences (P20GM103499) and the Howard Hughes Medical Institute
Start Date
15-4-2023 12:00 PM
Investigating Protein-Protein Interactions between Bacteriophage Cain and its Host Mycobacterium smegmatis
In the face of the antibiotic resistance crisis, bacteriophages have proven to be useful tools for overcoming bacterial defenses in medicine. However, 65% of phage genes have no predicted function, which limits the ability to utilize phages to their maximum potential. In order to effectively manipulate and apply phages therapeutically, we must close this gap in functional knowledge. After using a phenotypic assay to screen mycobacteriophage Cain gene products for interaction with the host cell, Mycobacterium smegmatis, a bacterial 2-hybrid (B2H) assay was used to identify which host proteins were being targeted by phage gene products. To achieve this, a complex array of specifically designed expression vectors, test promoters and cell lines, were used to report potential pairwise interactions between phage gene products and the host’s proteome. With no predicted function, and significant homology with well-studied phages in the same K cluster, Cain gp55 was investigated first. Results revealed interactions with host proteins NusA and GntR which are both transcriptional regulators thought to play various roles including transcript elongation/termination (NusA), cell motility, metabolism, and even virulence (GntR). Another phage gene of no predicted function, Cain gp2, also interacted with several host cell transcriptional regulators, including TetR, AraC, and LysR, thought to be involved in the regulation of metabolic genes. Identifying these interactions can give us hints about a phage gene’s role in manipulating its host. While the B2H assay gives us a starting point, more specific experimentation is needed to identify potentially new and useful functions of undescribed genes.
