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    Two dynamin-like proteins stabilize FtsZ rings during sporulation. Schlimpert Susan,Wasserstrom Sebastian,Chandra Govind,Bibb Maureen J,Findlay Kim C,Flärdh Klas,Buttner Mark J Proceedings of the National Academy of Sciences of the United States of America During sporulation, the filamentous bacteria undergo a massive cell division event in which the synthesis of ladders of sporulation septa convert multigenomic hyphae into chains of unigenomic spores. This process requires cytokinetic Z-rings formed by the bacterial tubulin homolog FtsZ, and the stabilization of the newly formed Z-rings is crucial for completion of septum synthesis. Here we show that two dynamin-like proteins, DynA and DynB, play critical roles in this process. Dynamins are a family of large, multidomain GTPases involved in key cellular processes in eukaryotes, including vesicle trafficking and organelle division. Many bacterial genomes encode dynamin-like proteins, but the biological function of these proteins has remained largely enigmatic. Using a cell biological approach, we show that the two dynamins specifically localize to sporulation septa in an FtsZ-dependent manner. Moreover, dynamin mutants have a cell division defect due to the decreased stability of sporulation-specific Z-rings, as demonstrated by kymographs derived from time-lapse images of FtsZ ladder formation. This defect causes the premature disassembly of individual Z-rings, leading to the frequent abortion of septum synthesis, which in turn results in the production of long spore-like compartments with multiple chromosomes. Two-hybrid analysis revealed that the dynamins are part of the cell division machinery and that they mediate their effects on Z-ring stability during developmentally controlled cell division via a network of protein-protein interactions involving DynA, DynB, FtsZ, SepF, SepF2, and the FtsZ-positioning protein SsgB. 10.1073/pnas.1704612114
    Regulation of Streptomyces development: reach for the sky! Claessen Dennis,de Jong Wouter,Dijkhuizen Lubbert,Wösten Han A B Trends in microbiology Streptomyces coelicolor is characterized by a complex life cycle and serves as a model system for bacterial development. After a feeding substrate mycelium has been formed, this filamentous bacterium differentiates by forming aerial hyphae that septate into spores. The bld cascade regulates initiation of aerial growth, whereas the whi genes control spore formation. Recent findings indicate the existence of another regulatory pathway that operates after aerial hyphae have started to grow into the air, which we call the sky pathway. This pathway controls the expression of the chaplin and rodlin genes. These genes encode proteins that assemble into a rodlet layer that provides surface hydrophobicity to aerial hyphae and spores. 10.1016/j.tim.2006.05.008
    Rewiring of transcriptional networks as a major event leading to the diversity of asexual multicellularity in fungi. Etxebeste Oier,Otamendi Ainara,Garzia Aitor,Espeso Eduardo A,Cortese Marc S Critical reviews in microbiology Complex multicellularity (CM) is characterized by the generation of three-dimensional structures that follow a genetically controlled program. CM emerged at least five times in evolution, one of them in fungi. There are two types of CM programs in fungi, leading, respectively, to the formation of sexual or asexual spores. Asexual spores foment the spread of mycoses, as they are the main vehicle for dispersion. In spite of this key dependence, there is great morphological diversity of asexual multicellular structures in fungi. To advance the understanding of the mechanisms that control initiation and progression of asexual CM and how they can lead to such a remarkable morphological diversification, we studied 503 fungal proteomes, representing all phyla and subphyla, and most known classes. Conservation analyses of 33 regulators of asexual development suggest stepwise emergence of transcription factors. While velvet proteins constitute one of the most ancient systems, the central regulator BrlA emerged late in evolution (with the class Eurotiomycetes). Some factors, such as MoConX4, seem to be species-specific. These observations suggest that the emergence and evolution of transcriptional regulators rewire transcriptional networks. This process could reach the species level, resulting in a vast diversity of morphologies. 10.1080/1040841X.2019.1630359
    Evolution of asexual and sexual reproduction in the aspergilli. Ojeda-López M,Chen W,Eagle C E,Gutiérrez G,Jia W L,Swilaiman S S,Huang Z,Park H-S,Yu J-H,Cánovas D,Dyer P S Studies in mycology has long-been used as a model organism to gain insights into the genetic basis of asexual and sexual developmental processes both in other members of the genus , and filamentous fungi in general. Paradigms have been established concerning the regulatory mechanisms of conidial development. However, recent studies have shown considerable genome divergence in the fungal kingdom, questioning the general applicability of findings from , and certain longstanding evolutionary theories have been questioned. The phylogenetic distribution of key regulatory elements of asexual reproduction in was investigated in a broad taxonomic range of fungi. This revealed that some proteins were well conserved in the ( AbaA, FlbA, FluG, NsdD, MedA, and some velvet proteins), suggesting similar developmental roles. However, other elements ( BrlA) had a more restricted distribution solely in the , and it appears that the genetic control of sporulation seems to be more complex in the aspergilli than in some other taxonomic groups of the . The evolution of the velvet protein family is discussed based on the history of expansion and contraction events in the early divergent fungi. Heterologous expression of the gene in failed to induce development of complete conidiophores as seen in the aspergilli, but did result in increased conidial production. The absence of many components of the asexual developmental pathway from members of the supports the hypothesis that differences in the complexity of their spore formation is due in part to the increased diversity of the sporulation machinery evident in the Pezizomycotina. Investigations were also made into the evolution of sex and sexuality in the aspergilli. loci were identified from the heterothallic () and () and the homothallic (=). A consistent architecture of the locus was seen in these and other heterothallic aspergilli whereas much variation was seen in the arrangement of loci in homothallic aspergilli. This suggested that it is most likely that the common ancestor of the aspergilli exhibited a heterothallic breeding system. Finally, the supposed prevalence of asexuality in the aspergilli was examined. Investigations were made using as a representative 'asexual' species. It was possible to induce a sexual cycle in given the correct and partners and environmental conditions, with recombination confirmed utilising molecular markers. This indicated that sexual reproduction might be possible in many supposedly asexual aspergilli and beyond, providing general insights into the nature of asexuality in fungi. 10.1016/j.simyco.2018.10.002
    Membrane-bound methyltransferase complex VapA-VipC-VapB guides epigenetic control of fungal development. Sarikaya-Bayram Ozlem,Bayram Ozgür,Feussner Kirstin,Kim Jong-Hwa,Kim Hee-Seo,Kaever Alexander,Feussner Ivo,Chae Keon-Sang,Han Dong-Min,Han Kap-Hoon,Braus Gerhard H Developmental cell Epigenetic and transcriptional control of gene expression must be coordinated in response to external signals to promote alternative multicellular developmental programs. The membrane-associated trimeric complex VapA-VipC-VapB controls a signal transduction pathway for fungal differentiation. The VipC-VapB methyltransferases are tethered to the membrane by the FYVE-like zinc finger protein VapA, allowing the nuclear VelB-VeA-LaeA complex to activate transcription for sexual development. Once the release from VapA is triggered, VipC-VapB is transported into the nucleus. VipC-VapB physically interacts with VeA and reduces its nuclear import and protein stability, thereby reducing the nuclear VelB-VeA-LaeA complex. Nuclear VapB methyltransferase diminishes the establishment of facultative heterochromatin by decreasing histone 3 lysine 9 trimethylation (H3K9me3). This favors activation of the regulatory genes brlA and abaA, which promote the asexual program. The VapA-VipC-VapB methyltransferase pathway combines control of nuclear import and stability of transcription factors with histone modification to foster appropriate differentiation responses. 10.1016/j.devcel.2014.03.020
    Systematic Dissection of the Evolutionarily Conserved WetA Developmental Regulator across a Genus of Filamentous Fungi. Wu Ming-Yueh,Mead Matthew E,Lee Mi-Kyung,Ostrem Loss Erin M,Kim Sun-Chang,Rokas Antonis,Yu Jae-Hyuk mBio Asexual sporulation is fundamental to the ecology and lifestyle of filamentous fungi and can facilitate both plant and human infection. In , the production of asexual spores is primarily governed by the BrlA→AbaA→WetA regulatory cascade. The final step in this cascade is controlled by the WetA protein and governs not only the morphological differentiation of spores but also the production and deposition of diverse metabolites into spores. While WetA is conserved across the genus , the structure and degree of conservation of the gene regulatory network (GRN) remain largely unknown. We carried out comparative transcriptome analyses of comparisons between null mutant and wild-type asexual spores in three representative species spanning the diversity of the genus : , , and We discovered that WetA regulates asexual sporulation in all three species via a negative-feedback loop that represses BrlA, the cascade's first step. Furthermore, data from chromatin immunoprecipitation sequencing (ChIP-seq) experiments in asexual spores suggest that WetA is a DNA-binding protein that interacts with a novel regulatory motif. Several global regulators known to bridge spore production and the production of secondary metabolites show species-specific regulatory patterns in our data. These results suggest that the BrlA→AbaA→WetA cascade's regulatory role in cellular and chemical asexual spore development is functionally conserved but that the -associated GRN has diverged during evolution. The formation of resilient spores is a key factor contributing to the survival and fitness of many microorganisms, including fungi. In the fungal genus , spore formation is controlled by a complex gene regulatory network that also impacts a variety of other processes, including secondary metabolism. To gain mechanistic insights into how fungal spore formation is controlled across , we dissected the gene regulatory network downstream of a major regulator of spore maturation (WetA) in three species that span the diversity of the genus: the genetic model , the human pathogen , and the aflatoxin producer Our data show that WetA regulates asexual sporulation in all three species via a negative-feedback loop and likely binds a novel regulatory element that we term the WetA response element (WRE). These results shed light on how gene regulatory networks in microorganisms control important biological processes and evolve across diverse species. 10.1128/mBio.01130-18
    HapX, an Indispensable bZIP Transcription Factor for Iron Acquisition, Regulates Infection Initiation by Orchestrating Conidial Oleic Acid Homeostasis and Cytomembrane Functionality in Mycopathogen Beauveria bassiana. Peng Yue-Jin,Wang Jia-Jia,Lin Hai-Yan,Ding Jin-Li,Feng Ming-Guang,Ying Sheng-Hua mSystems In pathogenic filamentous fungi, conidial germination not only is fundamental for propagation in the environment but is also a critical step of infection. In the insect mycopathogen , we genetically characterized the role of the basic leucine zipper (bZIP) transcription factor HapX () in conidial nutrient reserves and pathogen-host interaction. Ablation of resulted in an almost complete loss of virulence in the topical inoculation and intrahemocoel injection assays. Comparative transcriptomic analysis revealed that is required for fatty acid (FA)/lipid metabolism, and biochemical analyses indicated that loss caused a significant reduction in conidial FA contents. Exogenous oleic acid could partially or completely restore the impaired phenotypes of the Δ mutant, including germination rate, membrane integrity, vegetative growth, and virulence. mediates fungal iron acquisition which is not required for desaturation of stearic acid. Additionally, inactivation of the Δ9-fatty acid desaturase gene () generated defects similar to those of the Δ mutant; oleic acid also had significant restorative effects on the defective phenotypes of the Δ mutant. A gel retarding assay revealed that BbHapX directly regulated the expression of Lipidomic analyses indicated that both and contributed to the homeostasis of phospholipids with nonpolar tails derived from oleic acid; therefore, exogenous phospholipids could significantly restore membrane integrity. These data reveal that the HapX-Ole1 pathway contributes to conidial fatty acid/lipid reserves and that there are important links between the lipid biology and membrane functionality involved in the early stages of infection caused by Conidial maturation and germination are highly coupled physiological processes in filamentous fungi that are critical for the pathogenicity of mycopathogens. Compared to the mechanisms involved in conidial germination, those of conidial reserves during maturation are less understood. The insect-pathogenic fungus , as a representative species of filamentous fungi, is important for applied and fundamental research. In addition to its conserved roles in fungal adaptation to iron status, the bZIP transcription factor HapX acts as a master regulator involved in conidial virulence and regulates fatty acid/lipid metabolism. Further investigation revealed that the Δ9-fatty acid desaturase gene () is a direct downstream target of HapX. This study reveals the HapX-Ole1 pathway involved in the fatty acid/lipid accumulation associated with conidial maturation and provides new insights into the startup mechanism of infection caused by spores from pathogenic fungi. 10.1128/mSystems.00695-20
    Nuclear Ssr4 Is Required for the and Asexual Cycles and Global Gene Activity of Beauveria bassiana. Shao Wei,Cai Qing,Tong Sen-Miao,Ying Sheng-Hua,Feng Ming-Guang mSystems Ssr4 serves as a cosubunit of chromatin-remodeling SWI/SNF and RSC complexes in yeasts but remains functionally uncharacterized due to its essentiality for yeast viability. Here, we report pleiotropic effects of the deletion of the ortholog nonessential for cell viability in , an asexual insect mycopathogen. The deletion of resulted in severe growth defects on different carbon/nitrogen sources, increased hyphal hydrophilicity, blocked hyphal differentiation, and 98% reduced conidiation capacity compared to a wild-type standard. The limited Δ conidia featured an impaired coat with disordered or obscure hydrophobin rodlet bundles, decreased hydrophobicity, increased size, and lost insect pathogenicity via normal cuticle infection and 90% of virulence via intrahemocoel injection. The expression of genes required for hydrophobin biosynthesis and assembly of the rodlet layer was drastically repressed in more hydrophilic Δ cells. Transcriptomic analysis revealed 2,517 genes differentially expressed in the Δ mutant, including 1,505 downregulated genes and 1,012 upregulated genes. The proteins encoded by hundreds of repressed genes were involved in metabolism and/or transport of carbohydrates, amino acids, and lipids, inorganic ion transport and energy production or conversion, including dozens involved in DNA replication, transcription, translation, and posttranslational modifications. However, purified Ssr4 samples showed no DNA-binding activity, implying that the role of Ssr4 in genome-wide gene regulation could rely upon its acting as a cosubunit of the two complexes. These findings provide the first insight into an essential role of Ssr4 in the asexual cycle and of and highlights its importance for the filamentous fungal lifestyle. Ssr4 is known to serve as a cosubunit of chromatin-remodeling SWI/SNF and RSC complexes in yeasts but has not been functionally characterized in fungi. This study unveils for the first time the pleiotropic effects caused by deletion of and its role in mediating global gene expression in a fungal insect pathogen. Our findings confirm an essential role of Ssr4 in hydrophobin biosynthesis and assembly required for growth, differentiation, and development of aerial hyphae for conidiation and conidial adhesion to insect surface and its essentiality for insect pathogenicity and virulence-related cellular events. Importantly, Ssr4 can regulate nearly one-fourth of all genes in the fungal genome in direct and indirect manners, including dozens involved in gene activity and hundreds involved in metabolism and/or transport of carbohydrates, amino acids, lipids, and/or inorganic ions. These findings highlight a significance of Ssr4 for filamentous fungal lifestyle. 10.1128/mSystems.00677-19
    StuAp is a sequence-specific transcription factor that regulates developmental complexity in Aspergillus nidulans. Dutton J R,Johns S,Miller B L The EMBO journal The Aspergillus nidulans Stunted protein (StuAp) regulates multicellular complexity during asexual reproduction by moderating the core developmental program that directs differentiation of uninucleate, terminally differentiated spores from multinucleate, vegetative hyphae. StuAp is also required for ascosporogenesis and multicellular development during sexual reproduction. StuAp is a member of a family of fungal transcription factors that regulate development or cell cycle progression. Further, StuAp characterizes a sub-family possessing the conserved APSES domain. We demonstrate for the first time that the APSES domain is a sequence-specific DNA-binding domain that can be modeled as a basic helix-loop-helix (bHLH)-like structure. We have found that StuAp response elements (A/TCGCGT/ANA/C) are located upstream of both critical developmental regulatory genes and cell cycle genes in A.nidulans. StuAp is shown to act as a transcriptional repressor in A.nidulans, but as a weak activator in budding yeast. Our data suggest that the differentiation of pseudohyphal-like sterigmatal cells during multicellular conidiophore development requires correct StuAp-regulated expression of both developmental and cell cycle genes in A.nidulans. The budding pattern of sterigmata may involve processes related to those controlling pseudohyphal growth in budding yeast. 10.1093/emboj/16.18.5710
    HpoR, a novel c-di-GMP effective transcription factor, links the second messenger's regulatory function to the mycobacterial antioxidant defense. Li Weihui,Li Meng,Hu Lihua,Zhu Jingpeng,Xie Zhiwei,Chen Jiarui,He Zheng-Guo Nucleic acids research Cyclic di-GMP (c-di-GMP) is a global signaling molecule that widely modulates diverse cellular processes. However, whether or not the c-di-GMP signal participates in regulation of bacterial antioxidant defense is unclear, and the involved regulators remain to be explored. In this study, we characterized HpoR as a novel c-di-GMP effective transcription factor and found a link between the c-di-GMP signal and the antioxidant regulation in Mycobacterium smegmatis. H2O2 stress induces c-di-GMP accumulation in M. smegmatis. High level of c-di-GMP triggers expression of a redox gene cluster, designated as hpoR operon, which is required for the mycobacterial H2O2 resistance. HpoR acts as an inhibitor of the hpoR operon and recognizes a 12-bp motif sequence within the upstream regulatory region of the operon. c-di-GMP specifically binds with HpoR at a ratio of 1:1. Low concentrations of c-di-GMP stimulate the DNA-binding activity of HpoR, whereas high concentrations of the signal molecule inhibit the activity. Strikingly, high level of c-di-GMP de-represses the intracellular association of HpoR with the regulatory region of the hpoR operon in M. smegmatis and enhances the mycobacterial H2O2 resistance. Therefore, we report a novel c-di-GMP effective regulator in mycobacteria, which extends the second messenger's function to bacterial antioxidant defense. 10.1093/nar/gky146
    Multiple Roles of c-di-GMP Signaling in Bacterial Pathogenesis. Valentini Martina,Filloux Alain Annual review of microbiology The intracellular signaling molecule cyclic di-GMP (c-di-GMP) regulates the lifestyle of bacteria and controls many key functions and mechanisms. In the case of bacterial pathogens, a wide variety of virulence lifestyle factors have been shown to be regulated by c-di-GMP. Evidence of the importance of this molecule for bacterial pathogenesis has become so great that new antimicrobial agents are tested for their capacity of targeting c-di-GMP signaling. This review summarizes the current knowledge on this topic and reveals its application for the development of new antivirulence intervention strategies. 10.1146/annurev-micro-020518-115555
    Cyclic di-GMP: second messenger extraordinaire. Jenal Urs,Reinders Alberto,Lori Christian Nature reviews. Microbiology Cyclic dinucleotides (CDNs) are highly versatile signalling molecules that control various important biological processes in bacteria. The best-studied example is cyclic di-GMP (c-di-GMP). Known since the late 1980s, it is now recognized as a near-ubiquitous second messenger that coordinates diverse aspects of bacterial growth and behaviour, including motility, virulence, biofilm formation and cell cycle progression. In this Review, we discuss important new insights that have been gained into the molecular principles of c-di-GMP synthesis and degradation, which are mediated by diguanylate cyclases and c-di-GMP-specific phosphodiesterases, respectively, and the cellular functions that are exerted by c-di-GMP-binding effectors and their diverse targets. Finally, we provide a short overview of the signalling versatility of other CDNs, including c-di-AMP and cGMP-AMP (cGAMP). 10.1038/nrmicro.2016.190
    c-di-GMP Arms an Anti-σ to Control Progression of Multicellular Differentiation in Streptomyces. Gallagher Kelley A,Schumacher Maria A,Bush Matthew J,Bibb Maureen J,Chandra Govind,Holmes Neil A,Zeng Wenjie,Henderson Max,Zhang Hengshan,Findlay Kim C,Brennan Richard G,Buttner Mark J Molecular cell Streptomyces are our primary source of antibiotics, produced concomitantly with the transition from vegetative growth to sporulation in a complex developmental life cycle. We previously showed that the signaling molecule c-di-GMP binds BldD, a master repressor, to control initiation of development. Here we demonstrate that c-di-GMP also intervenes later in development to control differentiation of the reproductive hyphae into spores by arming a novel anti-σ (RsiG) to bind and sequester a sporulation-specific σ factor (σ). We present the structure of the RsiG-(c-di-GMP)-σ complex, revealing an unusual, partially intercalated c-di-GMP dimer bound at the RsiG-σ interface. RsiG binds c-di-GMP in the absence of σ, employing a novel E(X)S(X)R(X)Q(X)D motif repeated on each helix of a coiled coil. Further studies demonstrate that c-di-GMP is essential for RsiG to inhibit σ. These findings reveal a newly described control mechanism for σ-anti-σ complex formation and establish c-di-GMP as the central integrator of Streptomyces development. 10.1016/j.molcel.2019.11.006
    Fungal pathogens occurring on in Thailand. Thanakitpipattana D,Tasanathai K,Mongkolsamrit S,Khonsanit A,Lamlertthon S,Luangsa-Ard J J Persoonia Two new fungal genera and six species occurring on insects in the orders Orthoptera and Phasmatodea (superorder Orthopterida) were discovered that are distributed across three families in the . Sixty-seven sequences generated in this study were used in a multi-locus phylogenetic study comprising SSU, LSU, , and together with the nuclear intergenic region (IGR). These new taxa are introduced as , , , , and . shows resemblance to by infecting egg cases (ootheca) of praying mantis (Mantidae) and having obovoid perithecial heads but differs in the size of its perithecia and ascospore shape. Two new species in the cluster belonging to the complex are described that differ from known species with respect to phialide size, conidia and host. resembles in the absence of a stipe and can be distinguished by the production of whole ascospores, which are not commonly found in (except in , which produces multiseptate, whole ascospores) is pathogenic to mole crickets and shows resemblance to , and in having darkly pigmented stromata. occurs on small crickets, and is the phylogenetic sister species of taxa in the 'sphecocephala' clade. 10.3767/persoonia.2020.44.06
    From Root to Tips: Sporulation Evolution and Specialization in Bacillus subtilis and the Intestinal Pathogen Clostridioides difficile. Ramos-Silva Paula,Serrano Mónica,Henriques Adriano O Molecular biology and evolution Bacteria of the Firmicutes phylum are able to enter a developmental pathway that culminates with the formation of highly resistant, dormant endospores. Endospores allow environmental persistence, dissemination and for pathogens, are also infection vehicles. In both the model Bacillus subtilis, an aerobic organism, and in the intestinal pathogen Clostridioides difficile, an obligate anaerobe, sporulation mobilizes hundreds of genes. Their expression is coordinated between the forespore and the mother cell, the two cells that participate in the process, and is kept in close register with the course of morphogenesis. The evolutionary mechanisms by which sporulation emerged and evolved in these two species, and more broadly across Firmicutes, remain largely unknown. Here, we trace the origin and evolution of sporulation using the genes known to be involved in the process in B. subtilis and C. difficile, and estimating their gain-loss dynamics in a comprehensive bacterial macroevolutionary framework. We show that sporulation evolution was driven by two major gene gain events, the first at the base of the Firmicutes and the second at the base of the B. subtilis group and within the Peptostreptococcaceae family, which includes C. difficile. We also show that early and late sporulation regulons have been coevolving and that sporulation genes entail greater innovation in B. subtilis with many Bacilli lineage-restricted genes. In contrast, C. difficile more often recruits new sporulation genes by horizontal gene transfer, which reflects both its highly mobile genome, the complexity of the gut microbiota, and an adjustment of sporulation to the gut ecosystem. 10.1093/molbev/msz175
    Evolutionary Analysis of the Bacillus subtilis Genome Reveals New Genes Involved in Sporulation. Shi Lei,Derouiche Abderahmane,Pandit Santosh,Rahimi Shadi,Kalantari Aida,Futo Momir,Ravikumar Vaishnavi,Jers Carsten,Mokkapati Venkata R S S,Vlahoviček Kristian,Mijakovic Ivan Molecular biology and evolution Bacilli can form dormant, highly resistant, and metabolically inactive spores to cope with extreme environmental challenges. In this study, we examined the evolutionary age of Bacillus subtilis sporulation genes using the approach known as genomic phylostratigraphy. We found that B. subtilis sporulation genes cluster in several groups that emerged at distant evolutionary time-points, suggesting that the sporulation process underwent several stages of expansion. Next, we asked whether such evolutionary stratification of the genome could be used to predict involvement in sporulation of presently uncharacterized genes (y-genes). We individually inactivated a representative sample of uncharacterized genes that arose during the same evolutionary periods as the known sporulation genes and tested the resulting strains for sporulation phenotypes. Sporulation was significantly affected in 16 out of 37 (43%) tested strains. In addition to expanding the knowledge base on B. subtilis sporulation, our findings suggest that evolutionary age could be used to help with genome mining. 10.1093/molbev/msaa035