Part I: Mycobacterium tuberculosis, the etiological agent of tuberculosis, is a difficult pathogen to treat, requiring a lengthy treatment course with numerous antibiotics. It is believed that a robust regulation of gene expression contributes to high tolerance to antibiotics and other stressors. mRNA concentration is one physical factor that may impact mRNA half-life, a contributing factor to overall gene expression. Previous work in M. tuberculosis and other bacteria indicates a lack of consensus regarding whether mRNA abundance and mRNA half-life show a strong, negative correlation or a weak, positive correlation. Additionally, mRNA abundance may impact protein abundance in a non-linear fashion. However, there is a lack of consensus regarding the relationship between mRNA abundance and protein abundance. By understanding the impact of mRNA abundance on regulating gene expression, we sought to gain a greater understanding of how M. tuberculosis is able to effectively respond to stress. Using a tetracycline-inducible gene expression system in the model organism Mycolicibacterium smegmatis, we tested various combinations of concentrations of anhydrotetracycline (aTc) and induction times to determine conditions that would provide the widest range of mRNA and protein expression levels. We established that a range of aTc concentrations from 0 ng/mL to 50 ng/mL at a 4-hour induction time provided a wide range of gfpmut3 expression. The degradation data were too noisy to determine half-life and make meaningful conclusions regarding the relationship between mRNA abundance and mRNA half-life. Additionally, our system could not be used to investigate the relationship between mRNA abundance and protein abundance due to a loss of inducer-based expression for undetermined reasons. Part II: Metabolic pathways are complex, multi-structured organizations of proteins and metabolites that promote the continuation and survival of a living system. Through regulation of the proteins that compose these pathways, a robust system enables greater survival in a variety of stresses and conditions. M. tuberculosis, the causative agent of tuberculosis, is one such organism. This makes treatment of the pathogen difficult. Generating and using biovisualizations is important for culminating a greater understanding of M. tuberculosis and its mechanisms of regulating gene expression. While there are an abundance of biovisualizations mapping metabolic pathways and protein-protein interactions, there is little work involving the ability to visualize changes in protein expression and impacts of regulations on systems. Additionally, many of these visualizations and other existing visualization tools are based on reference pathways, and are not specific to M. tuberculosis. In this paper, a prototype biovisualization of M. tuberculosis metabolic pathways was constructed consisting of three panels: one visualizing metabolic pathways, one displaying protein-protein interactions, and one showing changes in gene expression from a user uploaded file. The tasks and design components were made based on interviews with M. tuberculosis researchers. This work highlights a prospective idea for future metabolic biovisualization work.

• This report represents the work of one or more WPI undergraduate students submitted to the faculty as evidence of completion of a degree requirement. WPI routinely publishes these reports on its website without editorial or peer review.

Creator

• Le, Van
• Orszulak, Adrian

Publisher

• Worcester Polytechnic Institute

Identifier

• 65831
• E-project-042822-160139

Keyword

• biovisualization
• metabolic pathways
• mRNA abundance
• gene expression
• mRNA half-life
• Mycolicibacterium smegmatis

Advisor

• Shell, Scarlet
• Harrison, Lane T.

Year

• 2022

Date created

• 2022-04-28

Resource type

• Major Qualifying Project

Major

• Biology & Biotechnology

Rights statement

• In Copyright