Anshu Chauhan, Hans Carolus, Dimitrios Sofras, Mohit Kumar, Praveen Kumar, Remya Nair, Aswathy Narayanan, Kusum Yadav, Basharat Ali, Vladislav Biriukov, Amandeep Saini, Ian Leaves, Rudy Vergauwen, Celia Lobo Romero, Dhara Malavia-Jones, Ashutosh Singh, Atanu Banerjee, Shivaprakash M. Rudramurthy, Arunaloke Chakrabarti, Alok K. Mondal, Naseem A. Gaur, Kaustuv Sanyal, Jeffrey M. Rybak, Toni Gabaldón, Patrick Van Dijck, Neil A. R. Gow, Rajendra Prasad
Clinical isolates of Candida auris show a high prevalence of resistance to Amphotericin B (AmB)—an uncommon trait in most Candida species. Alterations in ergosterol biosynthesis can contribute to acquired AmB resistance in C. auris laboratory strains but are rarely seen in clinical isolates. In this study, we experimentally evolved two drug-susceptible Clade II isolates of C. auris to develop AmB resistance. The evolved strains displayed a four to eight fold increase in MIC50 compared to the parental cells. We analyzed changes in their karyotype, genome, lipidome, and transcriptome associated with this acquired resistance. In one lineage, AOX2 was upregulated, and its deletion reversed the AmB resistance phenotype. The aox2Δ mutant also failed to evolve AmB resistance under experimental conditions. In the same lineage, restoring the UPC2S332Rand RTG3S101T mutations to the wild-type allele restored AmB susceptibility. In another lineage, the ergosterol and sphingolipid pathways were observed to play a critical role, and upregulation of the ERG genes elevated the total sterol content, while significant downregulation of HSX11 (glucosylceramide synthase) resulted in lower levels of glucosylceramides. To our knowledge, this study is the first to show that AmB resistance in C. auris can be acquired through mechanisms both dependent on or independent of sterol content modulation, highlighting Aox2 and Upc2 as key regulators of amphotericin resistance.
{"title":"Multi-Omics Analysis of Experimentally Evolved Candida auris Isolates Reveals Modulation of Sterols, Sphingolipids, and Oxidative Stress in Acquired Amphotericin B Resistance","authors":"Anshu Chauhan, Hans Carolus, Dimitrios Sofras, Mohit Kumar, Praveen Kumar, Remya Nair, Aswathy Narayanan, Kusum Yadav, Basharat Ali, Vladislav Biriukov, Amandeep Saini, Ian Leaves, Rudy Vergauwen, Celia Lobo Romero, Dhara Malavia-Jones, Ashutosh Singh, Atanu Banerjee, Shivaprakash M. Rudramurthy, Arunaloke Chakrabarti, Alok K. Mondal, Naseem A. Gaur, Kaustuv Sanyal, Jeffrey M. Rybak, Toni Gabaldón, Patrick Van Dijck, Neil A. R. Gow, Rajendra Prasad","doi":"10.1111/mmi.15379","DOIUrl":"https://doi.org/10.1111/mmi.15379","url":null,"abstract":"Clinical isolates of <i>Candida auris</i> show a high prevalence of resistance to Amphotericin B (AmB)—an uncommon trait in most <i>Candida</i> species. Alterations in ergosterol biosynthesis can contribute to acquired AmB resistance in <i>C. auris</i> laboratory strains but are rarely seen in clinical isolates. In this study, we experimentally evolved two drug-susceptible Clade II isolates of <i>C. auris</i> to develop AmB resistance. The evolved strains displayed a four to eight fold increase in MIC<sub>50</sub> compared to the parental cells. We analyzed changes in their karyotype, genome, lipidome, and transcriptome associated with this acquired resistance. In one lineage, <i>AOX2</i> was upregulated, and its deletion reversed the AmB resistance phenotype. The <i>aox2Δ</i> mutant also failed to evolve AmB resistance under experimental conditions. In the same lineage, restoring the <i>UPC2</i><sup><i>S332R</i></sup> <i>and RTG3</i><sup><i>S101T</i></sup> mutations to the wild-type allele restored AmB susceptibility. In another lineage, the ergosterol and sphingolipid pathways were observed to play a critical role, and upregulation of the ERG genes elevated the total sterol content, while significant downregulation of <i>HSX11</i> (glucosylceramide synthase) resulted in lower levels of glucosylceramides. To our knowledge, this study is the first to show that AmB resistance in <i>C. auris</i> can be acquired through mechanisms both dependent on or independent of sterol content modulation, highlighting Aox2 and Upc2 as key regulators of amphotericin resistance.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":"102 1","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-06-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144219516","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}
Number and arrangement of flagella, the bacterial locomotion organelles, are species-specific and serve as key taxonomic markers. The FlhG ATPase (also: YlxH, FleN), along with FlhF, plays pivotal roles in determining flagellation patterns. In Bacillus subtilis, FlhG and FlhF govern the spatial arrangement of peritrichous flagella. FlhG aids in flagellar assembly by interacting with the flagellar C-ring protein FliY, yet the molecular implications of this interaction have been unclear. Our study reveals that the ATP-dependent FlhG homodimer interacts with the C-terminal domain of GpsB, a cell cycle regulator, which recruits the peptidoglycan synthase PBP1 (also: ponA) to sites of cell wall elongation. A deletion of gpsB leads to dysregulation of the flagellation pattern mimicking the effects of a flhG deletion strain. The finding that GpsB can interact simultaneously with FlhG and PBP1, combined with the observation that GpsB and FliY can simultaneously interact with FlhG, strongly argues for a model in which FlhG confines flagella biosynthesis to regions of active cell wall biosynthesis. Thus, the FlhG-GpsB interaction appears to enable the locally restrained stimulation of the GTPase FlhF, known for its role to localize flagella in various bacterial species.
{"title":"FlhG Cooperates With the Cell Cycle Regulator GpsB to Confine Peritrichous Flagella in B. subtilis.","authors":"Anita Dornes,Patrica Bedrunka,Benjamin Pillet,Dieter Kressler,Thomas Heimerl,Jan Pané-Farré,Gert Bange","doi":"10.1111/mmi.15375","DOIUrl":"https://doi.org/10.1111/mmi.15375","url":null,"abstract":"Number and arrangement of flagella, the bacterial locomotion organelles, are species-specific and serve as key taxonomic markers. The FlhG ATPase (also: YlxH, FleN), along with FlhF, plays pivotal roles in determining flagellation patterns. In Bacillus subtilis, FlhG and FlhF govern the spatial arrangement of peritrichous flagella. FlhG aids in flagellar assembly by interacting with the flagellar C-ring protein FliY, yet the molecular implications of this interaction have been unclear. Our study reveals that the ATP-dependent FlhG homodimer interacts with the C-terminal domain of GpsB, a cell cycle regulator, which recruits the peptidoglycan synthase PBP1 (also: ponA) to sites of cell wall elongation. A deletion of gpsB leads to dysregulation of the flagellation pattern mimicking the effects of a flhG deletion strain. The finding that GpsB can interact simultaneously with FlhG and PBP1, combined with the observation that GpsB and FliY can simultaneously interact with FlhG, strongly argues for a model in which FlhG confines flagella biosynthesis to regions of active cell wall biosynthesis. Thus, the FlhG-GpsB interaction appears to enable the locally restrained stimulation of the GTPase FlhF, known for its role to localize flagella in various bacterial species.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":"83 1","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-05-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144087646","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}
Social interactions among bacteria can induce behaviors that affect their fitness and influence how complex communities assemble. Here we report a new socially induced motility behavior that we refer to as baited expansion in Pseudomonas syringae pv. tomato DC3000 (Pst DC3000), a plant pathogenic bacterium. We found Pst DC3000 displayed strongly induced swimming motility towards nearby colonies of Dickeya dianthicola or Escherichia coli. We developed a controlled system to visualize and characterize the development of baited expansion. Our results provide evidence that baited expansion behavior occurs in response to a chemical gradient established and maintained by the bait colony. We also found this behavior correlated with distinct transcriptional profiles and identified molybdenum cofactor (Moco) and a Moco‐utilizing oxidoreductase as crucial factors facilitating the baited expansion behavior.
{"title":"Pseudomonas syringae Socially Induced Swimming Motility Requires the Molybdenum Cofactor","authors":"Zichu Yang, Bryan Swingle","doi":"10.1111/mmi.15378","DOIUrl":"https://doi.org/10.1111/mmi.15378","url":null,"abstract":"Social interactions among bacteria can induce behaviors that affect their fitness and influence how complex communities assemble. Here we report a new socially induced motility behavior that we refer to as baited expansion in <jats:styled-content style=\"fixed-case\"><jats:italic>Pseudomonas syringae</jats:italic></jats:styled-content> pv. tomato DC3000 (<jats:italic>Pst</jats:italic> DC3000), a plant pathogenic bacterium. We found <jats:italic>Pst</jats:italic> DC3000 displayed strongly induced swimming motility towards nearby colonies of <jats:styled-content style=\"fixed-case\"><jats:italic>Dickeya dianthicola</jats:italic></jats:styled-content> or <jats:styled-content style=\"fixed-case\"><jats:italic>Escherichia coli</jats:italic></jats:styled-content>. We developed a controlled system to visualize and characterize the development of baited expansion. Our results provide evidence that baited expansion behavior occurs in response to a chemical gradient established and maintained by the bait colony. We also found this behavior correlated with distinct transcriptional profiles and identified molybdenum cofactor (Moco) and a Moco‐utilizing oxidoreductase as crucial factors facilitating the baited expansion behavior.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":"1 1","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-05-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144097134","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}
Iron is vital for most organisms, serving as a cofactor in enzymes, regulatory proteins, and respiratory cytochromes. In Corynebacterium glutamicum, iron and heme homeostasis are tightly interconnected and controlled by the global regulators DtxR and HrrA. While DtxR senses intracellular Fe2+, HrrSA is activated by heme. This study provides the first genome-wide analysis of DtxR and HrrA binding dynamics under varying iron and heme conditions using chromatin affinity purification and sequencing (ChAP-Seq). We revealed 25 novel DtxR targets and 210 previously unrecognized HrrA targets. Among these, metH, encoding homocysteine methyltransferase, and xerC, encoding a tyrosine recombinase, were bound by DtxR exclusively under heme conditions, underscoring condition-dependent variation. Activation of metH by DtxR links iron metabolism to methionine synthesis, potentially relevant for the mitigation of oxidative stress. Beyond novel targets, 16 shared targets between DtxR and HrrA, some with overlapping operator sequences, highlight their interconnected regulons. Strikingly, we demonstrate the significance of weak ChAP-Seq peaks that are often disregarded in global approaches, but feature an impact of the regulator on differential gene expression. These findings emphasize the importance of genome-wide profiling under different conditions to uncover novel targets and shed light on the complexity and dynamic nature of bacterial regulatory networks.
{"title":"Genome-Wide Analysis of DtxR and HrrA Regulons Reveals Novel Targets and a High Level of Interconnectivity Between Iron and Heme Regulatory Networks in Corynebacterium glutamicum","authors":"Aileen Krüger, Ulrike Weber, Julia Frunzke","doi":"10.1111/mmi.15376","DOIUrl":"https://doi.org/10.1111/mmi.15376","url":null,"abstract":"Iron is vital for most organisms, serving as a cofactor in enzymes, regulatory proteins, and respiratory cytochromes. In <i>Corynebacterium glutamicum</i>, iron and heme homeostasis are tightly interconnected and controlled by the global regulators DtxR and HrrA. While DtxR senses intracellular Fe<sup>2+</sup>, HrrSA is activated by heme. This study provides the first genome-wide analysis of DtxR and HrrA binding dynamics under varying iron and heme conditions using chromatin affinity purification and sequencing (ChAP-Seq). We revealed 25 novel DtxR targets and 210 previously unrecognized HrrA targets. Among these, <i>metH,</i> encoding homocysteine methyltransferase, and <i>xerC,</i> encoding a tyrosine recombinase, were bound by DtxR exclusively under heme conditions, underscoring condition-dependent variation. Activation of <i>metH</i> by DtxR links iron metabolism to methionine synthesis, potentially relevant for the mitigation of oxidative stress. Beyond novel targets, 16 shared targets between DtxR and HrrA, some with overlapping operator sequences, highlight their interconnected regulons. Strikingly, we demonstrate the significance of weak ChAP-Seq peaks that are often disregarded in global approaches, but feature an impact of the regulator on differential gene expression. These findings emphasize the importance of genome-wide profiling under different conditions to uncover novel targets and shed light on the complexity and dynamic nature of bacterial regulatory networks.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":"43 1","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-05-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144066349","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}
Nathan Hill, Lara M. Matulina, Cameron MacIntyre, M. Amine Hassani, Sheila Thomas, Matteo Luban, Isabelle Ward, Amina Abdalla, John M. Leong, Brandon L. Garcia, Jacob E. Lemieux
Lyme disease is a tick-borne spirochetosis with diverse clinical manifestations. Genotypic and phenotypic variation among Borrelia burgdorferi strains correlates with variable manifestations of Lyme disease in humans; this diversity is attributed in part to variation in surface-exposed lipoproteins, which are targets of the human antibody response and contribute to tissue adhesion, immune evasion, and other host interactions. Many B. burgdorferi lipoproteins are encoded as multi-copy gene families, such as the OspE/F-like leader peptide (Elp) protein family, which inhibits classical complement activation by binding complement C1s. To characterize Elp allelic variants, we adapted the Pseudomonas syringae ice nucleation protein (INP) system to present B. burgdorferi lipoproteins on the surface of Escherichia coli. Using this system, we identified interactions with classical complement proteins and mapped binding regions, then validated interactions using recombinant proteins and B. burgdorferi surface display. We also discovered a novel potential interaction between Elp proteins and the mammalian basement membrane protein perlecan, thus revealing a bifunctional nature of Elps. Our findings indicate that Elps have undergone functional diversification while maintaining classical complement inhibition mediated by potent and conserved C1s binding and demonstrate that E. coli surface display offers an efficient, cost-effective, and relatively high-throughput approach to characterize B. burgdorferi lipoproteins.
{"title":"Heterologous Surface Display Reveals Conserved Complement Inhibition and Functional Diversification of Borrelia burgdorferi Elp Proteins","authors":"Nathan Hill, Lara M. Matulina, Cameron MacIntyre, M. Amine Hassani, Sheila Thomas, Matteo Luban, Isabelle Ward, Amina Abdalla, John M. Leong, Brandon L. Garcia, Jacob E. Lemieux","doi":"10.1111/mmi.15369","DOIUrl":"https://doi.org/10.1111/mmi.15369","url":null,"abstract":"Lyme disease is a tick-borne spirochetosis with diverse clinical manifestations. Genotypic and phenotypic variation among <i>Borrelia burgdorferi</i> strains correlates with variable manifestations of Lyme disease in humans; this diversity is attributed in part to variation in surface-exposed lipoproteins, which are targets of the human antibody response and contribute to tissue adhesion, immune evasion, and other host interactions. Many <i>B. burgdorferi</i> lipoproteins are encoded as multi-copy gene families, such as the OspE/F-like leader peptide (Elp) protein family, which inhibits classical complement activation by binding complement C1s. To characterize Elp allelic variants, we adapted the <i>Pseudomonas syringae</i> ice nucleation protein (INP) system to present <i>B. burgdorferi</i> lipoproteins on the surface of <i>Escherichia coli</i>. Using this system, we identified interactions with classical complement proteins and mapped binding regions, then validated interactions using recombinant proteins and <i>B. burgdorferi</i> surface display. We also discovered a novel potential interaction between Elp proteins and the mammalian basement membrane protein perlecan, thus revealing a bifunctional nature of Elps. Our findings indicate that Elps have undergone functional diversification while maintaining classical complement inhibition mediated by potent and conserved C1s binding and demonstrate that <i>E. coli</i> surface display offers an efficient, cost-effective, and relatively high-throughput approach to characterize <i>B. burgdorferi</i> lipoproteins.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":"11 1","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-05-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144066647","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}
All methanogens that can fix nitrogen use molybdenum (Mo) nitrogenase. Some methanogens, including Methanosarcina acetivorans, also contain alternative vanadium- and iron-nitrogenases, encoded by the vnf and anf operons, respectively. These nitrogenases are produced when there is insufficient Mo to support Mo-nitrogenase activity. The factors that control the expression of the alternative nitrogenases in response to Mo availability are unknown in methanogens. Here we show that ModE is the regulator that represses transcription of the vnf and anf operons in M. acetivorans when cells are grown with Mo. CRISPRi repression of modE results in a significant increase in the transcription of the vnf and anf operons as well as the detection of Fe-nitrogenase during nitrogen fixation in the presence of Mo. Gel shift assays with recombinant ModE demonstrated that ModE binds a specific sequence motif upstream of the vnf and anf operons, as well as other genes and operons related to nitrogen fixation and Mo transport. However, purified ModE does not contain Mo, and the addition of Mo does not alter the affinity of ModE for DNA, indicating M. acetivorans ModE may not directly bind Mo. This study shows that ModE is the primary Mo-responsive regulator of alternative nitrogenase expression in M. acetivorans, but other factor(s) are likely involved in directly sensing Mo.
{"title":"ModE Regulates Alternative Nitrogenase Expression in the Methanogen Methanosarcina acetivorans.","authors":"Melissa Chanderban,Daniel J Lessner","doi":"10.1111/mmi.15377","DOIUrl":"https://doi.org/10.1111/mmi.15377","url":null,"abstract":"All methanogens that can fix nitrogen use molybdenum (Mo) nitrogenase. Some methanogens, including Methanosarcina acetivorans, also contain alternative vanadium- and iron-nitrogenases, encoded by the vnf and anf operons, respectively. These nitrogenases are produced when there is insufficient Mo to support Mo-nitrogenase activity. The factors that control the expression of the alternative nitrogenases in response to Mo availability are unknown in methanogens. Here we show that ModE is the regulator that represses transcription of the vnf and anf operons in M. acetivorans when cells are grown with Mo. CRISPRi repression of modE results in a significant increase in the transcription of the vnf and anf operons as well as the detection of Fe-nitrogenase during nitrogen fixation in the presence of Mo. Gel shift assays with recombinant ModE demonstrated that ModE binds a specific sequence motif upstream of the vnf and anf operons, as well as other genes and operons related to nitrogen fixation and Mo transport. However, purified ModE does not contain Mo, and the addition of Mo does not alter the affinity of ModE for DNA, indicating M. acetivorans ModE may not directly bind Mo. This study shows that ModE is the primary Mo-responsive regulator of alternative nitrogenase expression in M. acetivorans, but other factor(s) are likely involved in directly sensing Mo.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":"137 1","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-05-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143932897","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}
Cheyenne D. Lee, Arshad Rizvi, Zavier A. Carter, Adrianne N. Edwards, Shonna M. McBride
Clostridioides difficile is an anaerobic enteric pathogen that disseminates in the environment as a dormant spore. For C. difficile and other sporulating bacteria, the initiation of sporulation is a regulated process that prevents spore formation under favorable growth conditions. In Bacillus subtilis, one such mechanism for preventing sporulation is the prokaryotic 5-oxoprolinase, PxpB (KipI), which impedes the activation of the main sporulation kinase. In addition, PxpB functions as part of a complex that detoxifies the intermediate metabolite, 5-oxoproline (OP), a harmful by-product of glutamic acid and its derivatives. In this study, we investigate the orthologous Pxp proteins in C. difficile to determine their roles in the regulation of sporulation and metabolism. Through deletion of the pxpAGBC operon, we show that, unlike in B. subtilis, the Pxp (Kip) proteins have no significant impact on sporulation. However, we found that the pxp operon encodes a functional oxoprolinase that facilitates detoxification of OP. Furthermore, our data demonstrate that PxpAGBC not only detoxifies OP but also allows OP to be used as a nutrient source that supports the growth of C. difficile, thereby facilitating the conversion of a toxic by-product of metabolism into an energy source.
{"title":"The Pxp Complex Detoxifies 5-Oxoproline and Promotes the Growth of Clostridioides difficile","authors":"Cheyenne D. Lee, Arshad Rizvi, Zavier A. Carter, Adrianne N. Edwards, Shonna M. McBride","doi":"10.1111/mmi.15373","DOIUrl":"https://doi.org/10.1111/mmi.15373","url":null,"abstract":"<i>Clostridioides difficile</i> is an anaerobic enteric pathogen that disseminates in the environment as a dormant spore. For <i>C. difficile</i> and other sporulating bacteria, the initiation of sporulation is a regulated process that prevents spore formation under favorable growth conditions. In <i>Bacillus subtilis</i>, one such mechanism for preventing sporulation is the prokaryotic 5-oxoprolinase, PxpB (KipI), which impedes the activation of the main sporulation kinase. In addition, PxpB functions as part of a complex that detoxifies the intermediate metabolite, 5-oxoproline (OP), a harmful by-product of glutamic acid and its derivatives. In this study, we investigate the orthologous Pxp proteins in <i>C. difficile</i> to determine their roles in the regulation of sporulation and metabolism. Through deletion of the <i>pxpAGBC</i> operon, we show that, unlike in <i>B. subtilis,</i> the Pxp (Kip) proteins have no significant impact on sporulation. However, we found that the <i>pxp</i> operon encodes a functional oxoprolinase that facilitates detoxification of OP. Furthermore, our data demonstrate that PxpAGBC not only detoxifies OP but also allows OP to be used as a nutrient source that supports the growth of <i>C. difficile</i>, thereby facilitating the conversion of a toxic by-product of metabolism into an energy source.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":"37 1","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-05-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143920960","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}
Otmane Lamrabet, Raphael Munoz-Ruiz, Imen Ayadi, Alixia Bourbon, Erwann Pain, Joseph Oddy, Pierre Cosson
Phagocytic cells ingest bacteria and kill them in phagosomes. A variety of molecular mechanisms allow the killing and destruction of bacteria in phagosomes, but their complete list and relative importance remain poorly defined. Here we have used Dictyostelium discoideum amoebae as model phagocytic cells. Our results reveal that PldX, a luminal phospholipase D, plays an important role in the phagosomal destruction of ingested bacteria. Analysis of bacterial destruction in wild-type and pldX KO living cells suggests that PldX participates in the permeabilization of the bacterial membrane. The bacteriolytic activity of D. discoideum extracts was also measured in vitro: extracts from pldX KO cells exhibit significantly less bacteriolytic activity than wild-type cells, confirming the role of PldX in the lysis of bacterial membranes. These results identify luminal phospholipase D as a major player in the permeabilization of bacterial membranes in phagosomes.
{"title":"Luminal Phospholipase D Attacks Bacterial Membranes in Dictyostelium discoideum Phagosomes","authors":"Otmane Lamrabet, Raphael Munoz-Ruiz, Imen Ayadi, Alixia Bourbon, Erwann Pain, Joseph Oddy, Pierre Cosson","doi":"10.1111/mmi.15367","DOIUrl":"https://doi.org/10.1111/mmi.15367","url":null,"abstract":"Phagocytic cells ingest bacteria and kill them in phagosomes. A variety of molecular mechanisms allow the killing and destruction of bacteria in phagosomes, but their complete list and relative importance remain poorly defined. Here we have used <i>Dictyostelium discoideum</i> amoebae as model phagocytic cells. Our results reveal that PldX, a luminal phospholipase D, plays an important role in the phagosomal destruction of ingested bacteria. Analysis of bacterial destruction in wild-type and <i>pldX</i> KO living cells suggests that PldX participates in the permeabilization of the bacterial membrane. The bacteriolytic activity of <i>D. discoideum</i> extracts was also measured in vitro: extracts from <i>pldX</i> KO cells exhibit significantly less bacteriolytic activity than wild-type cells, confirming the role of PldX in the lysis of bacterial membranes. These results identify luminal phospholipase D as a major player in the permeabilization of bacterial membranes in phagosomes.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":"79 1","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-05-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143905431","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}
Actinobacillus pleuropneumoniae is the causative agent of pleuropneumonia in swine, a highly contagious and economically significant disease. The genetic variability of A. pleuropneumoniae complicates disease control efforts, as it enables rapid adaptation to various stressors, including antimicrobial treatments. To better understand the molecular mechanisms underlying this adaptability, we investigated the role of the bacterial defensome and its relationship with mobile genetic elements (MGEs), such as prophages, plasmids, and integrative conjugative elements (ICEs). Using bioinformatic tools, we identified a diverse and rich defensome in A. pleuropneumoniae, with an average of 16 different defense systems per strain. We found that CRISPR-Cas systems, along with other defense mechanisms, are actively involved in restricting the entry of foreign genetic material, playing a crucial role in bacterial adaptation. Additionally, we characterized several novel prophages and examined their distribution across different strains, revealing their potential contribution to the bacterium's evolutionary success. Our findings underscore the complex interplay between the bacterium's defense systems and MGEs, shedding light on how A. pleuropneumoniae maintains genetic diversity while also safeguarding itself against external threats. These insights provide a better understanding of the genetic factors that influence the pathogen's adaptability and highlight potential avenues for more effective disease control strategies.
{"title":"The Arms Race Between Actinobacillus pleuropneumoniae and Its Genetic Environment: A Comprehensive Analysis of Its Defensome and Mobile Genetic Elements","authors":"Giarlã Cunha da Silva, Ciro César Rossi","doi":"10.1111/mmi.15374","DOIUrl":"https://doi.org/10.1111/mmi.15374","url":null,"abstract":"<i>Actinobacillus pleuropneumoniae</i> is the causative agent of pleuropneumonia in swine, a highly contagious and economically significant disease. The genetic variability of <i>A. pleuropneumoniae</i> complicates disease control efforts, as it enables rapid adaptation to various stressors, including antimicrobial treatments. To better understand the molecular mechanisms underlying this adaptability, we investigated the role of the bacterial defensome and its relationship with mobile genetic elements (MGEs), such as prophages, plasmids, and integrative conjugative elements (ICEs). Using bioinformatic tools, we identified a diverse and rich defensome in <i>A. pleuropneumoniae</i>, with an average of 16 different defense systems per strain. We found that CRISPR-Cas systems, along with other defense mechanisms, are actively involved in restricting the entry of foreign genetic material, playing a crucial role in bacterial adaptation. Additionally, we characterized several novel prophages and examined their distribution across different strains, revealing their potential contribution to the bacterium's evolutionary success. Our findings underscore the complex interplay between the bacterium's defense systems and MGEs, shedding light on how <i>A. pleuropneumoniae</i> maintains genetic diversity while also safeguarding itself against external threats. These insights provide a better understanding of the genetic factors that influence the pathogen's adaptability and highlight potential avenues for more effective disease control strategies.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":"109 1","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-05-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143901813","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}
Brian Nguyen, Carly Ching, Ashley MacGuire, Pranav Casula, Connor Newman, Faith Finley, Veronica G. Godoy
Acinetobacter baumannii is an opportunistic pathogen causing several infections that are increasingly difficult to treat due to its ability to rapidly gain antibiotic resistances. These resistances can arise due to mutations through the activity of error-prone DNA polymerases, such as DNA polymerase V (DNA Pol V) in response to DNA damage. The regulation of the DNA damage response (DDR) in A. baumannii is not completely understood; the regulation of genes encoding multiple copies of DNA Pol V is not fully characterized. Through genome-wide mutagenesis, we have identified a novel TetR-like family regulator of the umuDC and umuC genes, which we have named Error-prone polymerase regulator (EppR). We have found that EppR represses the expression of the genes encoding DNA Pol V and itself through direct binding to an EppR motif in their promoters. Lastly, we show that EppR also regulates UmuDAb, previously identified as a regulator of genes encoding DNA Pol V. These two gene products are functionally required to ensure regulation of the expression of the two umuDC, the two umuC genes as well as the regulators umuDAb and eppR genes. With these results, we propose a model in which multiple transcription factors regulate the expression of all these genes.
鲍曼不动杆菌是一种机会致病菌，引起多种感染，由于其迅速获得抗生素耐药性的能力，这种感染越来越难以治疗。这些抗性可能是由于易出错的DNA聚合酶（如DNA聚合酶V (DNA Pol V)）响应DNA损伤而产生的突变而产生的。鲍曼不动杆菌DNA损伤反应（DDR）的调控机制尚不完全清楚；编码DNA多拷贝Pol V的基因调控尚未完全确定。通过全基因组诱变，我们鉴定出了umuDC和umuC基因的一种新的类似于rt的家族调节剂，我们将其命名为易出错聚合酶调节剂（Error-prone polymerase regulator, EppR）。我们发现EppR通过直接结合启动子中的EppR基序来抑制编码DNA Pol V及其自身的基因的表达。最后，我们发现EppR还能调控UmuDAb，而UmuDAb先前被认为是编码DNA Pol v的基因的调节因子。这两个基因产物在功能上是必需的，以确保调控两个umuDC、两个umuC基因以及调节因子UmuDAb和EppR基因的表达。根据这些结果，我们提出了一个模型，其中多个转录因子调节所有这些基因的表达。
{"title":"Identification of EppR, a Second Repressor of Error-Prone DNA Polymerase Genes in Acinetobacter baumannii","authors":"Brian Nguyen, Carly Ching, Ashley MacGuire, Pranav Casula, Connor Newman, Faith Finley, Veronica G. Godoy","doi":"10.1111/mmi.15368","DOIUrl":"https://doi.org/10.1111/mmi.15368","url":null,"abstract":"<i>Acinetobacter baumannii</i> is an opportunistic pathogen causing several infections that are increasingly difficult to treat due to its ability to rapidly gain antibiotic resistances. These resistances can arise due to mutations through the activity of error-prone DNA polymerases, such as DNA polymerase V (DNA Pol V) in response to DNA damage. The regulation of the DNA damage response (DDR) in <i>A. baumannii</i> is not completely understood; the regulation of genes encoding multiple copies of DNA Pol V is not fully characterized. Through genome-wide mutagenesis, we have identified a novel TetR-like family regulator of the <i>umuDC</i> and <i>umuC</i> genes, which we have named Error-prone polymerase regulator (EppR). We have found that EppR represses the expression of the genes encoding DNA Pol V and itself through direct binding to an EppR motif in their promoters. Lastly, we show that EppR also regulates UmuDAb, previously identified as a regulator of genes encoding DNA Pol V. These two gene products are functionally required to ensure regulation of the expression of the two <i>umuDC</i>, the two <i>umuC</i> genes as well as the regulators <i>umuDAb</i> and <i>eppR</i> genes. With these results, we propose a model in which multiple transcription factors regulate the expression of all these genes.","PeriodicalId":19006,"journal":{"name":"Molecular Microbiology","volume":"41 1","pages":""},"PeriodicalIF":3.6,"publicationDate":"2025-04-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143849527","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}
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