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CsrA-Mediated Translational Activation of the hmsE mRNA Enhances HmsD-Dependent C-di-GMP-Enabled Biofilm Production in Yersinia pestis
The plague bacterium, Yersinia pestis, forms a biofilm-mediated blockage in the flea foregut that enhances its transmission by fleabite. Biofilm formation is positively controlled by cyclic di-GMP (c-di-GMP), which is synthesized by the diguanylate cyclases (DGC), HmsD and HmsT. While HmsD primarily...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Society for Microbiology
2023
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10294631/ https://www.ncbi.nlm.nih.gov/pubmed/37191545 http://dx.doi.org/10.1128/jb.00105-23 |
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author | Silva-Rohwer, Amelia R. Held, Kiara Yakhnin, Helen Babitzke, Paul Vadyvaloo, Viveka |
author_facet | Silva-Rohwer, Amelia R. Held, Kiara Yakhnin, Helen Babitzke, Paul Vadyvaloo, Viveka |
author_sort | Silva-Rohwer, Amelia R. |
collection | PubMed |
description | The plague bacterium, Yersinia pestis, forms a biofilm-mediated blockage in the flea foregut that enhances its transmission by fleabite. Biofilm formation is positively controlled by cyclic di-GMP (c-di-GMP), which is synthesized by the diguanylate cyclases (DGC), HmsD and HmsT. While HmsD primarily promotes biofilm-mediated blockage of fleas, HmsT plays a more minor role in this process. HmsD is a component of the HmsCDE tripartite signaling system. HmsC and HmsE posttranslationally inhibit or activate HmsD, respectively. HmsT-dependent c-di-GMP levels and biofilm formation are positively regulated by the RNA-binding protein CsrA. In this study we determined whether CsrA positively regulates HmsD-dependent biofilm formation through interactions with the hmsE mRNA. Gel mobility shift assays determined that CsrA binds specifically to the hmsE transcript. RNase T1 footprint assays identified a single CsrA binding site and CsrA-induced structural changes in the hmsE leader region. Translational activation of the hmsE mRNA was confirmed in vivo using plasmid-encoded inducible translational fusion reporters and by HmsE protein expression studies. Furthermore, mutation of the CsrA binding site in the hmsE transcript significantly reduced HmsD-dependent biofilm formation. These results suggest that CsrA binding leads to structural changes in the hmsE mRNA that enhance its translation to enable increased HmsD-dependent biofilm formation. Given the requisite function of HmsD in biofilm-mediated flea blockage, this CsrA-dependent increase in HmsD activity underscores that complex and conditionally defined modulation of c-di-GMP synthesis within the flea gut is required for Y. pestis transmission. IMPORTANCE Mutations enhancing c-di-GMP biosynthesis drove the evolution of Y. pestis to flea-borne transmissibility. c-di-GMP-dependent biofilm-mediated blockage of the flea foregut enables regurgitative transmission of Y. pestis by fleabite. The Y. pestis diguanylate cyclases (DGC), HmsT and HmsD, which synthesize c-di-GMP, play significant roles in transmission. Several regulatory proteins involved in environmental sensing, as well as signal transduction and response regulation, tightly control DGC function. An example is CsrA, a global posttranscriptional regulator that modulates carbon metabolism and biofilm formation. CsrA integrates alternative carbon usage metabolism cues to activate c-di-GMP biosynthesis through HmsT. Here, we demonstrated that CsrA additionally activates hmsE translation to promote c-di-GMP biosynthesis through HmsD. This emphasizes that a highly evolved regulatory network controls c-di-GMP synthesis and Y. pestis transmission. |
format | Online Article Text |
id | pubmed-10294631 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | American Society for Microbiology |
record_format | MEDLINE/PubMed |
spelling | pubmed-102946312023-06-28 CsrA-Mediated Translational Activation of the hmsE mRNA Enhances HmsD-Dependent C-di-GMP-Enabled Biofilm Production in Yersinia pestis Silva-Rohwer, Amelia R. Held, Kiara Yakhnin, Helen Babitzke, Paul Vadyvaloo, Viveka J Bacteriol Research Article The plague bacterium, Yersinia pestis, forms a biofilm-mediated blockage in the flea foregut that enhances its transmission by fleabite. Biofilm formation is positively controlled by cyclic di-GMP (c-di-GMP), which is synthesized by the diguanylate cyclases (DGC), HmsD and HmsT. While HmsD primarily promotes biofilm-mediated blockage of fleas, HmsT plays a more minor role in this process. HmsD is a component of the HmsCDE tripartite signaling system. HmsC and HmsE posttranslationally inhibit or activate HmsD, respectively. HmsT-dependent c-di-GMP levels and biofilm formation are positively regulated by the RNA-binding protein CsrA. In this study we determined whether CsrA positively regulates HmsD-dependent biofilm formation through interactions with the hmsE mRNA. Gel mobility shift assays determined that CsrA binds specifically to the hmsE transcript. RNase T1 footprint assays identified a single CsrA binding site and CsrA-induced structural changes in the hmsE leader region. Translational activation of the hmsE mRNA was confirmed in vivo using plasmid-encoded inducible translational fusion reporters and by HmsE protein expression studies. Furthermore, mutation of the CsrA binding site in the hmsE transcript significantly reduced HmsD-dependent biofilm formation. These results suggest that CsrA binding leads to structural changes in the hmsE mRNA that enhance its translation to enable increased HmsD-dependent biofilm formation. Given the requisite function of HmsD in biofilm-mediated flea blockage, this CsrA-dependent increase in HmsD activity underscores that complex and conditionally defined modulation of c-di-GMP synthesis within the flea gut is required for Y. pestis transmission. IMPORTANCE Mutations enhancing c-di-GMP biosynthesis drove the evolution of Y. pestis to flea-borne transmissibility. c-di-GMP-dependent biofilm-mediated blockage of the flea foregut enables regurgitative transmission of Y. pestis by fleabite. The Y. pestis diguanylate cyclases (DGC), HmsT and HmsD, which synthesize c-di-GMP, play significant roles in transmission. Several regulatory proteins involved in environmental sensing, as well as signal transduction and response regulation, tightly control DGC function. An example is CsrA, a global posttranscriptional regulator that modulates carbon metabolism and biofilm formation. CsrA integrates alternative carbon usage metabolism cues to activate c-di-GMP biosynthesis through HmsT. Here, we demonstrated that CsrA additionally activates hmsE translation to promote c-di-GMP biosynthesis through HmsD. This emphasizes that a highly evolved regulatory network controls c-di-GMP synthesis and Y. pestis transmission. American Society for Microbiology 2023-05-16 /pmc/articles/PMC10294631/ /pubmed/37191545 http://dx.doi.org/10.1128/jb.00105-23 Text en Copyright © 2023 Silva-Rohwer et al. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license (https://creativecommons.org/licenses/by/4.0/) . |
spellingShingle | Research Article Silva-Rohwer, Amelia R. Held, Kiara Yakhnin, Helen Babitzke, Paul Vadyvaloo, Viveka CsrA-Mediated Translational Activation of the hmsE mRNA Enhances HmsD-Dependent C-di-GMP-Enabled Biofilm Production in Yersinia pestis |
title | CsrA-Mediated Translational Activation of the hmsE mRNA Enhances HmsD-Dependent C-di-GMP-Enabled Biofilm Production in Yersinia pestis |
title_full | CsrA-Mediated Translational Activation of the hmsE mRNA Enhances HmsD-Dependent C-di-GMP-Enabled Biofilm Production in Yersinia pestis |
title_fullStr | CsrA-Mediated Translational Activation of the hmsE mRNA Enhances HmsD-Dependent C-di-GMP-Enabled Biofilm Production in Yersinia pestis |
title_full_unstemmed | CsrA-Mediated Translational Activation of the hmsE mRNA Enhances HmsD-Dependent C-di-GMP-Enabled Biofilm Production in Yersinia pestis |
title_short | CsrA-Mediated Translational Activation of the hmsE mRNA Enhances HmsD-Dependent C-di-GMP-Enabled Biofilm Production in Yersinia pestis |
title_sort | csra-mediated translational activation of the hmse mrna enhances hmsd-dependent c-di-gmp-enabled biofilm production in yersinia pestis |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10294631/ https://www.ncbi.nlm.nih.gov/pubmed/37191545 http://dx.doi.org/10.1128/jb.00105-23 |
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