Researchers from the Helmholtz Institute for RNA-based Infection Research (HIRI) in Würzburg have utilized CRISPR interference for the first time to determine the function of the mutualistic gut bacterium Bacteroides thetaiotaomicron. They have identified a small RNA molecule (sRNA) that regulates microbial growth in the presence of bile acids.
The findings of this study are expected to contribute to a better understanding of the bacterium’s lifestyle in its natural environment, the human digestive system, and to the development of clinical applications. The study has been published in the Proceedings of the National Academy of Sciences.
Bacteroides thetaiotaomicron is a highly abundant component of the human gut microbiota. This bacterium aids in the digestion of polysaccharides and is crucial for human health, but it can also cause or contribute to infections. B. thetaiotaomicron is being increasingly studied as a model organism, but there is still limited knowledge about the functions of its genes. This particularly applies to genes encoding non-coding types of nucleic acids, which are transcribed as small non-coding RNAs (sRNAs) and are not translated into proteins.
“Our gut mutualists have an impact on our health, but we have very little knowledge about the functions of non-coding genes,” says Alexander Westermann.
Considering the important roles of non-coding DNA regions in infections by pathogens, it can be assumed that they play a similar crucial role in beneficial bacteria like Bacteroides, says Westermann.
Westermann initiated the study and leads a research group at the Helmholtz Institute for RNA-based Infection Research (HIRI) located in Würzburg, a branch of the Helmholtz Center for Infection Research in Braunschweig, in collaboration with the University of Würzburg.
“The limitations of microbiota-related interventions result from an incomplete understanding of the functions of the genes of many species that make up the microbiota. Although the significance of sRNA genes in bacteria has been recognized, tools for their global functional characterization are lacking,” says Westermann.
In collaboration with Chase Beisel’s lab at HIRI, scientists from Westermann’s laboratory used a biochemically engineered tool called CRISPR interference (CRISPRi) to address this challenge. Using specifically designed RNA guides, CRISPRi blocks the expression of selected genes, essentially turning them off.
“Despite ongoing efforts, our best knowledge is that CRISPRi had not been previously used for systematic functional screening of Bacteroides thetaiotaomicron genes,” says Gianluca Prezza, the first author of the study and a doctoral candidate in Westermann’s lab.
Analogous to restoring genes to their normal state, the researchers used CRISPRi to generate a targeted collection of disruptions of sRNAs in these important gut bacteria. In a subsequent screening study, previously unknown sRNAs were identified that regulate genes involved in the construction of Bacteroides cell surfaces and provide increased susceptibility to bile salts. Prezza states, “Silencing the identified sRNA, called BatR, increases the resistance of Bacteroides to bile stress.”
Overall, the presented collection of RNA guides holds potential for systematically discovering the functions of Bacteroides genes under various experimental conditions. “Our work demonstrates the advantages of CRISPRi for the functional characterization of sRNAs and serves as a foundation for targeted gene knockouts in these numerous members of the human microbiota,” says Westermann. Moreover, this study lays the groundwork for replicating such an approach in other bacterial species.
More information:
Gianluca Prezza et al, CRISPR-based screening of small RNA modulators of bile susceptibility in Bacteroides thetaiotaomicron, Proceedings of the National Academy of Sciences (2024). DOI: 10.1073/pnas.2311323121. www.pnas.org/doi/10.1073/pnas.2311323121
FAQ Section based on the main topics and information presented in the article:
1. What is the significance of the study conducted by the Helmholtz Institute for RNA-based Infection Research in Würzburg?
The study conducted by the Helmholtz Institute for RNA-based Infection Research in Würzburg utilized CRISPR interference for the first time to determine the function of the mutualistic gut bacterium Bacteroides thetaiotaomicron. The results of the study will contribute to a better understanding of the bacterium’s lifestyle in its natural environment, the human digestive system, and the development of clinical applications.
2. What is Bacteroides thetaiotaomicron?
Bacteroides thetaiotaomicron is a highly abundant component of the human gut microbiota. This bacterium aids in the digestion of polysaccharides and is crucial for human health, but it can also cause or contribute to infections.
3. What are sRNAs?
sRNAs are small non-coding RNAs that are transcribed as part of non-coding genes. In the case of the study on Bacteroides thetaiotaomicron, sRNAs regulate microbial growth in the presence of bile acids.
4. What is the significance of CRISPRi in the study?
CRISPR interference (CRISPRi) was used in the study to disrupt selected genes in Bacteroides thetaiotaomicron. This tool enables researchers to understand the functions of sRNA genes and create collections of sRNA disruptions for experimental purposes.
5. What are the potential applications of the presented collection of RNA guides?
The presented collection of RNA guides has the potential for systematically discovering the functions of Bacteroides thetaiotaomicron genes under various experimental conditions. This approach can be replicated in other bacterial species.
6. What is the role of non-coding genes in bacterial infections?
Non-coding genes play an important role in both pathogens and beneficial bacteria. The study on Bacteroides thetaiotaomicron aims to better understand the functions of non-coding genes in this gut bacterium, which plays a significant role in human health.
7. Where was the study published?
The study was published in the journal Proceedings of the National Academy of Sciences.
Links to relevant primary domains (without subpages):
– Helmholtz Institute for RNA-based Infection Research (HIRI)
– University of Würzburg
– Proceedings of the National Academy of Sciences
The source of the article is from the blog bitperfect.pe