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A Fast and Reliable Pipeline for Bacterial Transcriptome Analysis Case study: Serine-dependent Gene Regulation in Streptococcus pneumoniae
Published on: April 25, 2015
Synthetic gene-regulatory networks in the opportunistic human pathogen Streptococcus pneumoniae
Robin A Sorg1, Clement Gallay2, Laurye Van Maele3
1Molecular Genetics Group, Groningen Biomolecular Sciences and Biotechnology Institute, Centre for Synthetic Biology, University of Groningen, 9747 AG, Groningen, The Netherlands.
Abstract:
Streptococcus pneumoniae can cause disease in various human tissues and organs, including the ear, the brain, the blood, and the lung, and thus in highly diverse and dynamic environments. It is challenging to study how pneumococci control virulence factor expression, because cues of natural environments and the presence of an immune system are difficult to simulate in vitro. Here, we apply synthetic biology methods to reverse-engineer gene expression control in S. pneumoniae A selection platform is described that allows for straightforward identification of transcriptional regulatory elements out of combinatorial libraries. We present TetR- and LacI-regulated promoters that show expression ranges of four orders of magnitude. Based on these promoters, regulatory networks of higher complexity are assembled, such as logic AND gates and IMPLY gates. We demonstrate single-copy genome-integrated toggle switches that give rise to bimodal population distributions. The tools described here can be used to mimic complex expression patterns, such as the ones found for pneumococcal virulence factors. Indeed, we were able to rewire gene expression of the capsule operon, the main pneumococcal virulence factor, to be externally inducible (YES gate) or to act as an IMPLY gate (only expressed in absence of inducer). Importantly, we demonstrate that these synthetic gene-regulatory networks are functional in an influenza A virus superinfection murine model of pneumonia, paving the way for in vivo investigations of the importance of gene expression control on the pathogenicity of S. pneumoniae.
Insights
Synthetic biology tools were developed to control gene expression in Streptococcus pneumoniae. These tools enable the study of virulence factor regulation and function in vivo, advancing our understanding of pneumococcal disease.
Area of Science:
- Synthetic Biology
- Microbial Pathogenesis
- Gene Regulation
Background:
- Streptococcus pneumoniae causes diverse infections, but studying its virulence factor expression in vitro is challenging due to the lack of natural environmental cues and immune system presence.
- Understanding pneumococcal virulence factor control is crucial for developing effective treatments and interventions against this pathogen.
Purpose of the Study:
- To apply synthetic biology methods for reverse-engineering gene expression control in Streptococcus pneumoniae.
- To develop tools for identifying regulatory elements and assembling complex gene-regulatory networks.
- To investigate the function of these synthetic networks in vivo, particularly concerning virulence factor expression.
Main Methods:
- Developed a selection platform for identifying transcriptional regulatory elements from combinatorial libraries.
- Engineered TetR- and LacI-regulated promoters with a four-orders-of-magnitude expression range.
- Assembled complex regulatory networks, including AND and IMPLY gates, and single-copy genome-integrated toggle switches.
Main Results:
- Demonstrated TetR- and LacI-regulated promoters enabling precise control over gene expression.
- Successfully created synthetic gene-regulatory networks, including logic gates and toggle switches, leading to bimodal population distributions.
- Rewired the capsule operon (a key virulence factor) to be externally inducible (YES gate) or conditionally expressed (IMPLY gate).
- Validated the functionality of these synthetic networks in a murine model of influenza A virus superinfection-induced pneumonia.
Conclusions:
- Synthetic biology provides powerful tools to engineer and study gene expression control in Streptococcus pneumoniae.
- The developed tools allow for mimicking complex expression patterns relevant to pneumococcal virulence.
- These findings pave the way for in vivo investigations into the role of gene expression regulation in pneumococcal pathogenicity.
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