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Investigating RNA toxicity using sister sRNAs OmrA & OmrB

Black, Johnathan (2023) MOBN03 20222
Degree Projects in Molecular Biology
Abstract
Bacteria have harnessed a variety of self-acting toxic mechanisms to regulate plasmid maintenance, phage defense, expression of virulence factors, and antimicrobial resistance. It was thought that the expression of toxic proteins was the sole source of toxicity in bacteria. Recent studies have shown the potential for RNA to cause toxic growth phenotypes independent of toxic protein expression. However, the current mechanism of such toxicity is unknown.

Using the sRNA OmrA, this project aims to elucidate a mechanism of RNA-based toxicity, using a variety of standard molecular microbiology methods to test omrA interactions at the transcriptional, post-transcriptional and enzymatic level. Links were discovered between OmrA toxicity and TCA... (More)
Bacteria have harnessed a variety of self-acting toxic mechanisms to regulate plasmid maintenance, phage defense, expression of virulence factors, and antimicrobial resistance. It was thought that the expression of toxic proteins was the sole source of toxicity in bacteria. Recent studies have shown the potential for RNA to cause toxic growth phenotypes independent of toxic protein expression. However, the current mechanism of such toxicity is unknown.

Using the sRNA OmrA, this project aims to elucidate a mechanism of RNA-based toxicity, using a variety of standard molecular microbiology methods to test omrA interactions at the transcriptional, post-transcriptional and enzymatic level. Links were discovered between OmrA toxicity and TCA intermediate 2-oxoglutarate. Additional work was performed to establish a transposon mutagenesis protocol as an unbiased approach to find novel genes related to toxicity. (Less)
Popular Abstract
RNA toxicity in bacteria

Bacteria use toxicity to rapidly regulate their behavior and adapt to their environment. This can be done through expressing virulence factors, preventing phage infections, resisting antibiotic treatment. Understanding the mechanisms of toxicity may provide greater understanding of pathogens, how they cause disease and how to develop better treatment options. It was previously thought that toxic proteins were the sole source of toxicity in bacteria; However, there is increasing evidence that different RNAs can cause toxic phenotypes without the involvement of toxic proteins.

OmrA is small RNAs. Small RNAs are non-coding RNA that regulate protein manufacturing by binding to messenger RNAs and preventing them... (More)
RNA toxicity in bacteria

Bacteria use toxicity to rapidly regulate their behavior and adapt to their environment. This can be done through expressing virulence factors, preventing phage infections, resisting antibiotic treatment. Understanding the mechanisms of toxicity may provide greater understanding of pathogens, how they cause disease and how to develop better treatment options. It was previously thought that toxic proteins were the sole source of toxicity in bacteria; However, there is increasing evidence that different RNAs can cause toxic phenotypes without the involvement of toxic proteins.

OmrA is small RNAs. Small RNAs are non-coding RNA that regulate protein manufacturing by binding to messenger RNAs and preventing them from being translated into protein. Through this mechanism, OmrA can regulate biofilm formation in Escherichia coli. It was discovered by the Wagner group that OmrA can cause toxicity in minimal nutrient conditions.

Using OmrA, this project aims to discover a mechanism of RNA-based toxicity, using a variety of standard molecular microbiology methods to test omrA interactions at the transcriptional, posttranscriptional and enzymatic level.

Results & Discussion
By supplementing minimal media with different supplements, the phenotypical source of OmrA toxicity was linked to an intermediate the TCA cycle, an important process for energy production. Using molecular cloning methods, several different proteins involved in this cycle were tested to see if they were also involved in OmrA toxicity. However, no clear link was established.

An unbiased search for OmrA toxicity sources across the genome was carried out using transposon mutagenesis. This found a variety of genes involved in metabolism, inner membrane proteins and RNA modifications. The specific source and mechanism for OmrA toxicity is intertwined with a complex network of metabolic pathways, and more research is required to untangle this.

Master’s Degree Molecular Biology 60 credits 2023
Department of Biology, Lund University


Advisor: Professor Gerhart Wagner, Dr Thomas Stenum
Department of Cell & Molecular Biology, Uppsala University (Less)
Please use this url to cite or link to this publication:
author
Black, Johnathan
supervisor
organization
course
MOBN03 20222
year
type
H2 - Master's Degree (Two Years)
subject
language
English
id
9140680
date added to LUP
2023-10-31 14:23:21
date last changed
2023-10-31 14:23:21
@misc{9140680,
  abstract     = {{Bacteria have harnessed a variety of self-acting toxic mechanisms to regulate plasmid maintenance, phage defense, expression of virulence factors, and antimicrobial resistance. It was thought that the expression of toxic proteins was the sole source of toxicity in bacteria. Recent studies have shown the potential for RNA to cause toxic growth phenotypes independent of toxic protein expression. However, the current mechanism of such toxicity is unknown.

Using the sRNA OmrA, this project aims to elucidate a mechanism of RNA-based toxicity, using a variety of standard molecular microbiology methods to test omrA interactions at the transcriptional, post-transcriptional and enzymatic level. Links were discovered between OmrA toxicity and TCA intermediate 2-oxoglutarate. Additional work was performed to establish a transposon mutagenesis protocol as an unbiased approach to find novel genes related to toxicity.}},
  author       = {{Black, Johnathan}},
  language     = {{eng}},
  note         = {{Student Paper}},
  title        = {{Investigating RNA toxicity using sister sRNAs OmrA & OmrB}},
  year         = {{2023}},
}