Characterizing the Francisella tularensis Universal Stress Protein

Author

Benjamin Girardo, University of Nebraska Medical CenterFollow

ORCID ID

0009-0003-9050-5802

Graduation Date

Winter 12-20-2024

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Programs

Interdisciplinary Graduate Program in Biomedical Sciences

First Advisor

Dr. Marilynn A. Larson

Abstract

Francisella tularensis is an extremely virulent pathogen responsible for the zoonotic disease tularemia. F. tularensis employs various defense mechanisms to survive and persist under diverse environmental and host-induced stress conditions. Despite its significance, the full spectrum of stress factors that enable F. tularensis persistence remain inadequately understood. In many bacterial species, universal stress proteins (Usp) play a crucial role in managing adverse conditions. The aim of this study was to elucidate the role of Usp in F. tularensis as a means of clarifying the stress response mechanisms of this pathogen. The F. tularensis Usp is encoded by a single, highly conserved gene among different F. tularensis strains. Transcriptional analysis demonstrated that F. tularensis usp mRNA has a half-life exceeding 30 minutes, with protein and transcript levels remaining consistently high under various stress conditions, including nutrient deprivation, low pH, hydrogen peroxide, and paraquat exposure. Notably, the F. tularensis usp deletionmutant (Δusp) displayed impaired growth and recovery under paraquat-induced oxidative stress compared to the wildtype (WT) strain. Transcriptional studies further revealed that Usp supports the expression of oxyR and katG, key antioxidant defense genes in F. tularensis. A serendipitous discovery was then made when recombinant F. tularensis Usp (rUsp/His6) produced in Escherichia coli exhibited an apparent molecular weight of 33 kDa, surpassing the predicted 30 kDa. While this discrepancy was attributed primarily to a stop codon readthrough event resulting in an additional 20 amino acids at the C-terminus, the additional molecular weight was also attributed to the presence of post-translational modifications (PTM) on multiple rUsp/His₆ residues, including lysine acetylation and glutamine polyamination. Together, these findings highlight Usp’s protective role in F. tularensis oxidative stress response, while providing insights into potential regulatory functions of Usp afforded by its observed PTMs.

Comments

2024 Copyright, the authors

Recommended Citation

Girardo, Benjamin, "Characterizing the Francisella tularensis Universal Stress Protein" (2024). Theses & Dissertations. 881.
https://digitalcommons.unmc.edu/etd/881