Molecular Microbiology最新文献

{"title":"TasA Fibre Interactions Are Necessary for Bacillus subtilis Biofilm Structure","authors":"Natalie C. Bamford, Ryan J. Morris, Alan Prescott, Paul Murphy, Elliot Erskine, Cait E. MacPhee, Nicola R. Stanley-Wall","doi":"10.1111/mmi.15315","DOIUrl":"https://doi.org/10.1111/mmi.15315","url":null,"abstract":"The extracellular matrix of biofilms provides crucial structural support to the community and protection from environmental perturbations. TasA, a key <i>Bacillus subtilis</i> biofilm matrix protein, forms both amyloid and non-amyloid fibrils. Non-amyloid TasA fibrils are formed via a strand-exchange mechanism, whereas the amyloid-like form involves non-specific self-assembly. We performed mutagenesis of the N-terminus to assess the role of non-amyloid fibrils in biofilm development. We find that the N-terminal tail is essential for the formation of structured biofilms, providing evidence that the strand-exchange fibrils are the active form in the biofilm matrix. Furthermore, we demonstrate that fibre formation alone is not sufficient to give structure to the biofilm. We build an interactome of TasA with other extracellular protein components, and identify important interaction sites. Our results provide insight into how protein–matrix interactions modulate biofilm development.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":null,"pages":null},"PeriodicalIF":3.6,"publicationDate":"2024-09-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142330270","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Peptidoglycan Endopeptidase PBP7 Facilitates the Recruitment of FtsN to the Divisome and Promotes Peptidoglycan Synthesis in Escherichia coli","authors":"Xinwei Liu, Gabriela Boelter, Waldemar Vollmer, Manuel Banzhaf, Tanneke den Blaauwen","doi":"10.1111/mmi.15321","DOIUrl":"https://doi.org/10.1111/mmi.15321","url":null,"abstract":"<i>Escherichia coli</i> has many periplasmic hydrolases to degrade and modify peptidoglycan (PG). However, the redundancy of eight PG endopeptidases makes it challenging to define specific roles to individual enzymes. Therefore, the cellular role of PBP7 (encoded by <i>pbpG</i>) is not clearly defined. In this work, we show that PBP7 localizes in the lateral cell envelope and at midcell. The C-terminal α-helix of PBP7 is crucial for midcell localization but not for its activity, which is dispensable for this localization. Additionally, midcell localization of PBP7 relies on the assembly of FtsZ up to FtsN in the divisome, and on the activity of PBP3. PBP7 was found to affect the assembly timing of FtsZ and FtsN in the divisome. The absence of PBP7 slows down the assembly of FtsN at midcell. The Δ<i>pbpG</i> mutant exhibited a weaker incorporation of the fluorescent D-amino acid HADA, reporting on transpeptidase activity, compared to wild-type cells. This could indicate reduced PG synthesis at the septum of the Δ<i>pbpG</i> strain, explaining the slower accumulation of FtsN and suggesting that endopeptidase-mediated PG cleavage may be a rate-limiting step for septal PG synthesis.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":null,"pages":null},"PeriodicalIF":3.6,"publicationDate":"2024-09-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142330267","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Unveiling the Coordinated Action of DesK/DesR and YvfT/YvfU to Control the Expression of an ABC Transporter in Bacillus subtilis","authors":"Pilar Fernández, Lucía Porrini, Julián Ignacio Pereyra, Daniela Albanesi, María Cecilia Mansilla","doi":"10.1111/mmi.15320","DOIUrl":"https://doi.org/10.1111/mmi.15320","url":null,"abstract":"Two-component systems (TCSs) are vital signal transduction pathways ubiquitous among bacteria, facilitating their responses to diverse environmental stimuli. In <i>Bacillus subtilis</i>, the DesK histidine kinase thermosensor, together with the response regulator DesR, constitute a TCS dedicated to membrane lipid homeostasis maintenance. This TCS orchestrates the transcriptional regulation of the <i>des</i> gene, encoding the sole desaturase in these bacteria, Δ5-Des. Additionally, <i>B. subtilis</i> possesses a paralog TCS, YvfT/YvfU, with unknown target gene(s). In this work, we show that YvfT/YvfU controls the expression of the <i>yvfRS</i> operon that codes for an ABC transporter. Interestingly, we found that this regulation also involves the action of DesK/DesR. Notably, opposite to <i>des</i>, <i>yvfRS</i> transcription is induced at 37°C and not at 25°C. Our in vivo and in vitro experiments demonstrate that both YvfU and DesR directly bind to the operon promoter region, with DesR exerting its control over <i>yvfRS</i> expression in its unphosphorylated state. Our study uncovers an intriguing case of cross-regulation where two homologous TCSs interact closely to finely tune gene expression in response to environmental cues. These findings shed light on the complexity of bacterial signal transduction systems and their critical role in bacterial adaptability.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":null,"pages":null},"PeriodicalIF":3.6,"publicationDate":"2024-09-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142330268","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Analysis of bb0556 Expression and Its Role During Borrelia burgdorferi Mammalian Infection","authors":"Sierra George, Connor Waldron, Christina Thompson, Zhiming Ouyang","doi":"10.1111/mmi.15319","DOIUrl":"https://doi.org/10.1111/mmi.15319","url":null,"abstract":"In <i>Borrelia burgdorferi</i>, BB0556 was annotated as a conserved hypothetical protein. We herein investigated gene expression and the importance of this protein during infection. Our data support that <i>bb0556</i> forms an operon with five other genes. A transcriptional start site and the associated σ⁷⁰-type promoter were identified in the sequences upstream of <i>bb0554</i>, and luciferase reporter assays indicated that this promoter is functional in <i>B. burgdorferi</i>. Furthermore, the sequences upstream of <i>bb0556</i> contain an internal promoter to drive gene expression. <i>bb0556</i> expression was affected by various environmental factors such as changes in temperature, pH, and cell density when <i>B. burgdorferi</i> was grown in vitro. Surprisingly, significant differences were observed for <i>bb0556</i> expression between <i>B. burgdorferi</i> strains B31-A3 and CE162, likely due to the different <i>cis-</i> and <i>trans</i>-acting factors in these strains. Moreover, <i>bb0556</i> was found to be highly expressed by <i>B. burgdorferi</i> in infected mice tissues, suggesting that this gene plays an important role during animal infection. To test this hypothesis, we generated a <i>bb0556</i> deletion mutant in a virulent bioluminescent <i>B. burgdorferi</i> strain. The mutant grew normally in the medium and displayed no defect in the resistance to environmental stresses such as reactive oxygen species, reactive nitrogen species, and osmotic stress. However, when the infectivity was compared between the mutant and its parental strain using in vivo bioluminescence imaging as well as analyses of spirochete recovery and bacterial burdens in animal tissues, our data showed that, contrary to the parental strain, the mutant was unable to infect mice. Complementation of <i>bb0556</i> in <i>cis</i> fully restored the infectious phenotype to wild-type levels. Taken together, our study demonstrates that the hypothetical protein BB0556 is a novel virulence factor essential for <i>B. burgdorferi</i> mammalian infection.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":null,"pages":null},"PeriodicalIF":3.6,"publicationDate":"2024-09-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142275659","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Specificity of Membrane-Associated J-Domain Protein, Caj1, in Amphotericin B Tolerance in Budding Yeast","authors":"Preeti Sagarika, Neha Dobriyal, Pakirisamy Deepsika, Avanti Vairagkar, Ankita Das, Chandan Sahi","doi":"10.1111/mmi.15318","DOIUrl":"https://doi.org/10.1111/mmi.15318","url":null,"abstract":"Hsp70:J-domain protein (JDP) machineries play pivotal roles in maintaining cellular proteostasis and governing various aspects of fungal physiology. While Hsp70 is known for its involvement in conferring tolerance to diverse antifungal drugs, the specific contribution of JDPs remains unclear. In this study, we examined the sensitivity of cytosolic JDP deletion strains of budding yeast to amphotericin B (AmB), a polyene antifungal agent widely utilized in fungal disease treatment due to its ability to disrupt the fungal plasma membrane (PM). Deleting Caj1, a PM-associated class II JDP, heightened susceptibility to AmB, and the protection conferred by Caj1 against AmB necessitated both its N-terminal J-domain and C-terminal lipid binding domain. Moreover, Caj1 deficiency compromised PM integrity as evidenced by increased phosphate efflux and exacerbated AmB sensitivity, particularly at elevated temperatures. Notably, phytosphingosine (PHS) addition as well as overexpression of <i>PMP3</i>, a positive PM integrity regulator, significantly rescued AmB sensitivity of <i>caj1Δ</i> cells. Our results align with the notion that Caj1 associates with the PM and cooperates with Hsp70 to regulate PM proteostasis, thereby influencing PM integrity in budding yeast. Loss of Caj1 function at the PM compromises PM protein quality control, thereby rendering yeast cells more susceptible to AmB.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":null,"pages":null},"PeriodicalIF":3.6,"publicationDate":"2024-09-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142237016","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Small Regulatory RNAs of the Rsm Clan in Pseudomonas","authors":"María Trinidad Gallegos, Matías Garavaglia, Claudio Valverde","doi":"10.1111/mmi.15313","DOIUrl":"https://doi.org/10.1111/mmi.15313","url":null,"abstract":"Bacteria of the genus <i>Pseudomonas</i> are ubiquitous on Earth due to their great metabolic versatility and adaptation to fluctuating environments and different hosts. Some groups are important animal/human and plant pathogens, whereas others are studied for their biotechnological applications, including bioremediation, biological control of phytopathogens and plant growth promotion. Notably, their adaptability is mediated by various signal transduction systems, with the post-transcriptional Gac-Rsm cascade playing a key role. This pervasive <i>Pseudomonas</i> pathway controls major transitions at the population level, such as motile/sessile lifestyle, primary/secondary metabolism or replicative/infective behaviour. A hallmark of the Gac-Rsm cascade is the participation of small, regulatory, non-coding RNAs of the Rsm clan. These RNAs are synthetised in response to cell-density-dependent autoinducer signals channelled through the GacS/GacA two-component system, and they counteract, by molecular mimicry, the translational control that RNA-binding proteins of the RsmA family exert over hundreds of mRNAs. Rsm RNAs have been investigated in a few <i>Pseudomonas</i> model species, evidencing the presence of a variable number and families of genes depending on the taxonomic clade. However, the global picture of the distribution of these riboregulators at the genus level was unknown until now. We have undertaken a comprehensive survey and annotation of the vast array of gene sequences encoding members of the Rsm RNA clan in 245 complete genomes that cover 28 phylogenomic clades across the entire genus. The properties of the different families of <i>rsm</i> genes, their phylogenetic radiation, as well as the features of their promoters and adjacent regions, are discussed. The novel insights presented in our manuscript will significantly boost research on the biology of these prevalent RNAs in understudied species of the genus <i>Pseudomonas</i> and closely related genera.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":null,"pages":null},"PeriodicalIF":3.6,"publicationDate":"2024-09-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142234053","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Multiple Effects of L-Leucine in Escherichia coli Lead to L-Leucine-Sensitive Growth in the Absence of Unphosphorylated PtsN","authors":"Neeraj Kumar, Abhijit A. Sardesai","doi":"10.1111/mmi.15317","DOIUrl":"https://doi.org/10.1111/mmi.15317","url":null,"abstract":"In <i>E. coli</i> K-12, the absence of unphosphorylated PtsN (unphospho-PtsN) has been proposed to cause an L-leucine-sensitive growth phenotype (Leu^S) by hyperactivated K⁺ uptake mediated impairment of the expression of the <i>ilvBN</i> operon, encoding subunits of the L-valine (Val)-sensitive acetohydroxyacid synthase I (AHAS I) that renders residual AHAS activity susceptible to inhibition by Leu and K⁺. This leads to AHAS insufficiency and a requirement for L-isoleucine (Ile). Herein, we provide an alternate mechanism for the Leu^S of the ∆<i>ptsN</i> mutant. Genetic and physiological studies with suppressors of the Leu^S indicate that impaired expression of the <i>ilvBN</i> operon jointly caused by the absence of unphospho-PtsN and the presence of Leu coupled to Leu-mediated repression of expression of AHAS III leads to AHAS insufficiency rendering residual AHAS activity susceptible to chronic Val stress that may be generated by exogenous Leu. Hyperactivated K⁺ uptake and an elevated α-ketobutyrate level mediate elevation of <i>ilvBN</i> expression and alleviate the Leu^S. The requirement of unphospho-PtsN as a positive regulator of <i>ilvBN</i> expression may buffer Ile biosynthesis against Leu-mediated AHAS insufficiency and protect AHAS I function from chronic endogenous Val generated by Leu and could be realized in certain environments that impair AHAS function.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":null,"pages":null},"PeriodicalIF":3.6,"publicationDate":"2024-09-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142231820","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The Impact of YabG Mutations on Clostridioides difficile Spore Germination and Processing of Spore Substrates","authors":"Morgan S. Osborne, Joshua N. Brehm, Carmen Olivença, Alicia M. Cochran, Mónica Serrano, Adriano O. Henriques, Joseph A. Sorg","doi":"10.1111/mmi.15316","DOIUrl":"https://doi.org/10.1111/mmi.15316","url":null,"abstract":"YabG is a sporulation‐specific protease that is conserved among sporulating bacteria. <jats:italic>Clostridioides difficile</jats:italic> YabG processes the cortex destined proteins preproSleC into proSleC and CspBA to CspB and CspA. YabG also affects synthesis of spore coat/exosporium proteins CotA and CdeM. In prior work that identified CspA as the co‐germinant receptor, mutations in <jats:italic>yabG</jats:italic> were found which altered the co‐germinants required to initiate spore germination. To understand how these mutations in the <jats:italic>yabG</jats:italic> locus contribute to <jats:italic>C</jats:italic>. <jats:italic>difficile</jats:italic> spore germination, we introduced these mutations into an isogenic background. Spores derived from <jats:italic>C</jats:italic>. <jats:italic>difficile yabG</jats:italic><jats:sub>C207A</jats:sub> (a catalytically inactive allele), <jats:italic>C</jats:italic>. <jats:italic>difficile yabG</jats:italic><jats:sub>A46D</jats:sub>, <jats:italic>C</jats:italic>. <jats:italic>difficile yabG</jats:italic><jats:sub>G37E</jats:sub>, and <jats:italic>C</jats:italic>. <jats:italic>difficile yabG</jats:italic><jats:sub>P153L</jats:sub> strains germinated in response to taurocholic acid alone. Recombinantly expressed and purified preproSleC incubated with <jats:italic>E</jats:italic>. <jats:italic>coli</jats:italic> lysate expressing wild type YabG resulted in the removal of the presequence from preproSleC. Interestingly, only YabG<jats:sub>A46D</jats:sub> showed any activity toward purified preproSleC. Mutation of the YabG processing site in preproSleC (R119A) led to YabG shifting its processing to R115 or R112. Finally, changes in <jats:italic>yabG</jats:italic> expression under the mutant promoters were analyzed using a SNAP‐tag and revealed expression differences at early and late stages of sporulation. Overall, our results support and expand upon the hypothesis that YabG is important for germination and spore assembly and, upon mutation of the processing site, can shift where it cleaves substrates.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":null,"pages":null},"PeriodicalIF":3.6,"publicationDate":"2024-09-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142170937","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Molecular Microbiology of Microbiomes","authors":"Pierre Santucci","doi":"10.1111/mmi.15308","DOIUrl":"https://doi.org/10.1111/mmi.15308","url":null,"abstract":"","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":null,"pages":null},"PeriodicalIF":3.6,"publicationDate":"2024-09-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142170934","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"In Vivo Cross-Linking Sheds Light on the Salmonella Divisome in Which PBP3 and PBP3_SAL Compete for Occupancy.","authors":"Sónia Castanheira, David López-Escarpa, Alberto Paradela, Francisco García-Del Portillo","doi":"10.1111/mmi.15309","DOIUrl":"https://doi.org/10.1111/mmi.15309","url":null,"abstract":"<p><p>Bacterial cell division is orchestrated by proteins that assemble in dynamic complexes collectively known as the divisome. Essential monofunctional enzymes with glycosyltransferase or transpeptidase (TPase) activities, FtsW and FtsI respectively, engage in the synthesis of septal peptidoglycan (sPG). Enigmatically, Salmonella has two TPases that can promote cell division independently: FtsI (PBP3) and the pathogen-specific paralogue PBP3_SAL. How Salmonella regulates the assembly of the sPG synthase complex with these two TPases, is unknown. Here, we characterized Salmonella division complexes in wild-type cells and isogenic mutants lacking PBP3 or PBP3_SAL. The complexes were cross-linked in vivo and pulled down with antibodies recognizing each enzyme. Proteomics of the immunoprecipitates showed that PBP3 and PBP3_SAL do not extensively cross-link in wild type cells, supporting the presence of independent complexes. More than 40 proteins cross-link in complexes in which these two TPases are present. Those identified with high scores include FtsA, FtsK, FtsQLB, FtsW, PBP1B, SPOR domain-containing proteins (FtsN, DedD, RlpA, DamX), amidase activators (FtsX, EnvC, NlpD) and Tol-Pal proteins. Other cross-linked proteins are the protease Prc, the elongasome TPase PBP2 and, D,D-endo- and D,D-carboxypeptidases. PBP3 and PBP3_SAL localize at midcell and compete for occupying the division complex in response to environmental cues. Thus, a catalytic-dead PBP3_SAL-S300A variant impairs cell division in a high osmolarity and acidic condition in which it is produced at levels exceeding those of PBP3. Salmonella may therefore exploit an 'adjustable' divisome to exchange TPases for ensuring cell division in distinct environments and, in this manner, expand its colonization capacities.</p>","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":null,"pages":null},"PeriodicalIF":2.6,"publicationDate":"2024-09-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142133258","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}