Project Title

Cyclic-di-GMP Regulates Salmonella Typhimurium Infection of Epithelial Cells and Macrophages

Authors' Affiliations

Abdulafiz Musa, Department of Biological Sciences, College of Art and Sciences, East Tennessee State University, Johnson City, TN. Dr. Erik Petersen, Department of Health Sciences, College of Public Health, East Tennessee State University, Johnson City, TN.

Location

Culp Center Rm. 304

Start Date

4-25-2023 10:00 AM

End Date

4-25-2023 10:20 AM

Faculty Sponsor’s Department

Health Sciences

Name of Project's Faculty Sponsor

Erik Petersen

Classification of First Author

Graduate Student-Master’s

Competition Type

Competitive

Type

Oral Presentation

Project's Category

Biological Sciences, Microbiology, Molecular Biology

Abstract or Artist's Statement

Regulation of the bacterial second messenger cyclic-di-GMP in Salmonella Typhimurium allows it to delicately alter phenotypes to optimize invasion and survive intracellularly in epithelial cells and macrophages to become virulent and cause infection. The concentration of cyclic-di-GMP is regulated by the presence of external stimuli, sensor CMEs (diguanylate cyclases, DGCs, and phosphodiesterases, PDEs), and cyclic-di-GMP binding effectors. Previous studies established that maintenance of low cyclic-di-GMP concentrations is required for survival in macrophages and that the deletion of 3 active PDEs reduces this survival. This study aimed to further investigate the regulation of cyclic-di-GMP for survival in macrophages and epithelial cells. Salmonella Typhimurium mutants were generated and used for an infection assay with RAW 264.7 macrophage and HeLa epithelial cell lines to determine active CMEs via intracellular survival. Intracellular survival was quantified by plate counting of cell lysates at 1-, 4-, and 24-hours post-infection. Our result showed that the previously identified 3 PDEs also influenced the infection of epithelial cells. We re-established the decreased survival in the RAW 264.7 macrophage cell line and determined that the cyclic-di-GMP-binding cellulose synthase BcsA was responsible for decreased survival in macrophages. Finally, we identified an active DGC whose deletion within the 3xKO PDEs restores survival levels, suggesting that this enzyme is responsible for the synthesis of cyclic-di-GMP during macrophage infection. Further studies on how cyclic-di-GMP regulates Salmonella Typhimurium intracellular survival could lead to identifying a potential alternative drug target for treating its infections.

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Apr 25th, 10:00 AM Apr 25th, 10:20 AM

Cyclic-di-GMP Regulates Salmonella Typhimurium Infection of Epithelial Cells and Macrophages

Culp Center Rm. 304

Regulation of the bacterial second messenger cyclic-di-GMP in Salmonella Typhimurium allows it to delicately alter phenotypes to optimize invasion and survive intracellularly in epithelial cells and macrophages to become virulent and cause infection. The concentration of cyclic-di-GMP is regulated by the presence of external stimuli, sensor CMEs (diguanylate cyclases, DGCs, and phosphodiesterases, PDEs), and cyclic-di-GMP binding effectors. Previous studies established that maintenance of low cyclic-di-GMP concentrations is required for survival in macrophages and that the deletion of 3 active PDEs reduces this survival. This study aimed to further investigate the regulation of cyclic-di-GMP for survival in macrophages and epithelial cells. Salmonella Typhimurium mutants were generated and used for an infection assay with RAW 264.7 macrophage and HeLa epithelial cell lines to determine active CMEs via intracellular survival. Intracellular survival was quantified by plate counting of cell lysates at 1-, 4-, and 24-hours post-infection. Our result showed that the previously identified 3 PDEs also influenced the infection of epithelial cells. We re-established the decreased survival in the RAW 264.7 macrophage cell line and determined that the cyclic-di-GMP-binding cellulose synthase BcsA was responsible for decreased survival in macrophages. Finally, we identified an active DGC whose deletion within the 3xKO PDEs restores survival levels, suggesting that this enzyme is responsible for the synthesis of cyclic-di-GMP during macrophage infection. Further studies on how cyclic-di-GMP regulates Salmonella Typhimurium intracellular survival could lead to identifying a potential alternative drug target for treating its infections.