Bacteria often use multiple transcription factors to regulate specific biological processes.Biosynthesis of heat-stable antifungal factor(HSAF)is regulated by multiple factors in Lysobacter enzymogenes.However,the mec...Bacteria often use multiple transcription factors to regulate specific biological processes.Biosynthesis of heat-stable antifungal factor(HSAF)is regulated by multiple factors in Lysobacter enzymogenes.However,the mechanism of HSAF biosynthesis regulation remains largely unknown.In this study,we screened a potential HSAF biosynthesis regulator,RecX,by a DNA pull-down assay.Deletion of recX resulted in a significant increase in the production of HSAF,and overexpression of recX significantly suppressed HSAF production.Importantly,our results showe that RecX directly binds to the promoter region of the lafB gene to inhibit its transcription and thus decreases HSAF production in L.enzymogenes.These findings reveal the novel mechanism of RecX regulation of antifungal antibiotic production in L.enzymogenes.展开更多
Bacterial species of the genus Lysobacter are environmentally ubiquitous with strong antifungal biocontrol potential.Heat-stable antifungal factor(HSAF)secreted by the biocontrol bacterium Lysobacter enzymogenes OH11 ...Bacterial species of the genus Lysobacter are environmentally ubiquitous with strong antifungal biocontrol potential.Heat-stable antifungal factor(HSAF)secreted by the biocontrol bacterium Lysobacter enzymogenes OH11 has broad-spectrum and highly efficient antifungal activity.Studying the biosynthetic regulations of HSAF would lay an important foundation for strain engineering toward improved HSAF production.In this work,we demonstrate that Le0752,an orotidine-5´-phosphate decarboxylase enzyme(ODCase)catalyzing a pivotal step of the UMP de novo biosynthesis pathway,is vital for HSAF-mediated antimicrobial activities and growth of L.enzymogenes OH11,but not for twitching motility.This gene regulates the production of HSAF by affecting the expression of lafB,a key gene in the HSAF biosynthesis operon,through the transcription factor Clp.Interestingly,bioinformatics analysis revealed that Le0752 belongs to the Group III ODCases,whereas its homologs in the closely related genera Xanthomonas and Stenotrophomonas belong to Group I,which contains most ODCases from Gram-positive bacteria,Gram-negative bacteria and cyanobacteria.Moreover,the Group I ODCase PXO_3614 from the Xanthomonas oryzae pv.oryzae PXO99A strain complemented the Le0752 mutant in regulating HSAF-mediated antagonistic activity.Together,these results highlight the important requirement of de novo pyrimidine biosynthetic enzymes for antibiotic HSAF production in L.enzymogenes,which lays an important foundation for improving HSAF production via metabolic flow design and for dissecting the regulatory functions of bacterial ODCases.展开更多
Small molecules are able to regulate numerous cellular processes through binding to various bacterial receptor proteins,but the mechanisms and functions by which these chemicals coordinate and execute remain poorly un...Small molecules are able to regulate numerous cellular processes through binding to various bacterial receptor proteins,but the mechanisms and functions by which these chemicals coordinate and execute remain poorly understood.4-hydroxybenzoic acid(4-HBA)and cyclic di-GMP(c-di-GMP)are two such molecules with distinct structures that are produced in Lysobacter enzymogenes to synergistically affect the secretion of an antifungal antibiotic,known as heat-stable antifungal factor(HSAF).In our earlier studies,we showed that CdgL,a YajQ-like protein without DNA-binding domain,was able to physically interact with LysR,a transcription factor,to enhance its binding affinity toward the upstream region of the HSAF biosynthesis operon promoter,hence increasing the HSAF biosynthesis.Interestingly,4-HBA or c-di-GMP can bind to its cognate receptor of LysR or CdgL,respectively,to regulate the HSAF biosynthesis.Further,c-di-GMP acts by binding to CdgL to induce the dissociation of the CdgL-LysR complex,leading to decreased downstream expression.We now showed that CdgL controlled the transcription of lenB2,which encodes an oxygenase to convert chorismate to 4-HBA.Notably,overexpression of cdgL was found to stimulate lenB2 transcription,which likely increased the intracellular 4-HBA content.Also,4-HBA could bind to LysR to interrupt the LysR-CdgL complex formation and release of CdgL,which caused a lower affinity of LysR toward DNA and hence decreased HSAF operon expression.These findings,along with our earlier report,allow us to propose a coordination mechanism demonstrating how the HSAF biosynthesis is co-regulated by 4-HBA and c-di-GMP through interactions with their cognate receptors.This new mechanism shall shed light on improving the HSAF yield for practical usage.展开更多
文摘HSAF(heat stable antifungal factor)是产酶溶杆菌Lysobacter enzymogenes OH11中分离鉴定的一种小分子广谱抗真菌和卵菌物质,为进一步明确HSAF防治梨树腐烂病的潜力,分别采用生长速率测定法、显微镜观察法和刮治涂抹法测定了HSAF对梨树腐烂病菌的室内毒力以及对梨树腐烂病的田间防治效果。结果表明,HSAF对梨树腐烂病菌有较强的生长抑制作用,EC50值为0.5μg/m L;HSAF可导致梨树腐烂病菌菌丝体出现扭曲、近顶端处分枝和顶端肿胀等畸形现象;40μg/m L HSAF凝胶剂对梨树腐烂病的涂治愈合率可达81.3%,与2.12%腐植酸铜水剂的涂治愈合率(85.3%)无显著差异;对于分枝裂口或冻伤处、向阴、主杆或中心枝上的病疤,经40μg/m L HSAF凝胶剂涂抹后的愈合率高于2.12%腐植酸铜水剂。对于剪锯口处和向阳或分枝上的梨树腐烂病病疤,经2.12%腐植酸铜水剂涂抹后的愈合率高于40μg/m L HSAF凝胶剂;两种药剂对不同位置病疤的涂治效果不同,可作互补使用。本研究结果为梨树腐烂病的田间防治提供了一种新的生防制剂,同时也可望为其他果树腐烂病害的防治提供生防药剂资源。
基金supported by the grants from the Natural Science Special Research Fund of Guizhou University,Special Post([2022]50)the National Natural Science Foundation of China(32260653).
文摘Bacteria often use multiple transcription factors to regulate specific biological processes.Biosynthesis of heat-stable antifungal factor(HSAF)is regulated by multiple factors in Lysobacter enzymogenes.However,the mechanism of HSAF biosynthesis regulation remains largely unknown.In this study,we screened a potential HSAF biosynthesis regulator,RecX,by a DNA pull-down assay.Deletion of recX resulted in a significant increase in the production of HSAF,and overexpression of recX significantly suppressed HSAF production.Importantly,our results showe that RecX directly binds to the promoter region of the lafB gene to inhibit its transcription and thus decreases HSAF production in L.enzymogenes.These findings reveal the novel mechanism of RecX regulation of antifungal antibiotic production in L.enzymogenes.
基金supported by the National Natural Science Foundation of China(32102283 to Mingming Yang)the Science and Technology Major Project of China National Tobacco Corporation(110202101056(LS-16))the Science and Technology Project of Shaanxi Branch of China National Tobacco Corporation(KJ-2021-02 and KJ-2022-04).
文摘Bacterial species of the genus Lysobacter are environmentally ubiquitous with strong antifungal biocontrol potential.Heat-stable antifungal factor(HSAF)secreted by the biocontrol bacterium Lysobacter enzymogenes OH11 has broad-spectrum and highly efficient antifungal activity.Studying the biosynthetic regulations of HSAF would lay an important foundation for strain engineering toward improved HSAF production.In this work,we demonstrate that Le0752,an orotidine-5´-phosphate decarboxylase enzyme(ODCase)catalyzing a pivotal step of the UMP de novo biosynthesis pathway,is vital for HSAF-mediated antimicrobial activities and growth of L.enzymogenes OH11,but not for twitching motility.This gene regulates the production of HSAF by affecting the expression of lafB,a key gene in the HSAF biosynthesis operon,through the transcription factor Clp.Interestingly,bioinformatics analysis revealed that Le0752 belongs to the Group III ODCases,whereas its homologs in the closely related genera Xanthomonas and Stenotrophomonas belong to Group I,which contains most ODCases from Gram-positive bacteria,Gram-negative bacteria and cyanobacteria.Moreover,the Group I ODCase PXO_3614 from the Xanthomonas oryzae pv.oryzae PXO99A strain complemented the Le0752 mutant in regulating HSAF-mediated antagonistic activity.Together,these results highlight the important requirement of de novo pyrimidine biosynthetic enzymes for antibiotic HSAF production in L.enzymogenes,which lays an important foundation for improving HSAF production via metabolic flow design and for dissecting the regulatory functions of bacterial ODCases.
基金supported by the Natural Science Foundation of Jiangsu Province(BK20190026,BK20181325)the National Natural Science Foundation of China(31872016)+2 种基金the Fundamental Research Funds for the Central Universities(KJJQ202001,KYYJ201904,KYT201805 and KYTZ201403)Jiangsu Agricultural Sciences and Technology Innovation Fund[CX(18)1003]Innovation Team Program for Jiangsu Universities(2017).
文摘Small molecules are able to regulate numerous cellular processes through binding to various bacterial receptor proteins,but the mechanisms and functions by which these chemicals coordinate and execute remain poorly understood.4-hydroxybenzoic acid(4-HBA)and cyclic di-GMP(c-di-GMP)are two such molecules with distinct structures that are produced in Lysobacter enzymogenes to synergistically affect the secretion of an antifungal antibiotic,known as heat-stable antifungal factor(HSAF).In our earlier studies,we showed that CdgL,a YajQ-like protein without DNA-binding domain,was able to physically interact with LysR,a transcription factor,to enhance its binding affinity toward the upstream region of the HSAF biosynthesis operon promoter,hence increasing the HSAF biosynthesis.Interestingly,4-HBA or c-di-GMP can bind to its cognate receptor of LysR or CdgL,respectively,to regulate the HSAF biosynthesis.Further,c-di-GMP acts by binding to CdgL to induce the dissociation of the CdgL-LysR complex,leading to decreased downstream expression.We now showed that CdgL controlled the transcription of lenB2,which encodes an oxygenase to convert chorismate to 4-HBA.Notably,overexpression of cdgL was found to stimulate lenB2 transcription,which likely increased the intracellular 4-HBA content.Also,4-HBA could bind to LysR to interrupt the LysR-CdgL complex formation and release of CdgL,which caused a lower affinity of LysR toward DNA and hence decreased HSAF operon expression.These findings,along with our earlier report,allow us to propose a coordination mechanism demonstrating how the HSAF biosynthesis is co-regulated by 4-HBA and c-di-GMP through interactions with their cognate receptors.This new mechanism shall shed light on improving the HSAF yield for practical usage.
