Response of a Rice Paddy Soil Methanogen to Syntrophic Growth as Revealed by Transcriptional Analyses

ABSTRACTMembers ofMethanocellalesare widespread in paddy field soils and play the key role in methane production. These methanogens feature largely in these organisms' adaptation to low H2and syntrophic growth with anaerobic fatty acid oxidizers. The adaptive mechanisms, however, remain unknown. In the present study, we determined the transcripts of 21 genes involved in the key steps of methanogenesis and acetate assimilation ofMethanocella conradiiHZ254, a strain recently isolated from paddy field soil.M. conradiiwas grown in monoculture and syntrophically withPelotomaculum thermopropionicum(a propionate syntroph) orSyntrophothermus lipocalidus(a butyrate syntroph). Comparison of the relative transcript abundances showed that three hydrogenase-encoding genes and all methanogenesis-related genes tested were upregulated in cocultures relative to monoculture. The genes encoding formylmethanofuran dehydrogenase (Fwd), heterodisulfide reductase (Hdr), and the membrane-bound energy-converting hydrogenase (Ech) were the most upregulated among the evaluated genes. The expression of the formate dehydrogenase (Fdh)-encoding gene also was significantly upregulated. In contrast, an acetate assimilation gene was downregulated in cocultures. The genes coding for Fwd, Hdr, and the D subunit of F420-nonreducing hydrogenase (Mvh) form a large predicted transcription unit; therefore, the Mvh/Hdr/Fwd complex, capable of mediating the electron bifurcation and connecting the first and last steps of methanogenesis, was predicted to be formed inM. conradii. We propose thatMethanocellamethanogens cope with low H2and syntrophic growth by (i) stabilizing the Mvh/Hdr/Fwd complex and (ii) activating formate-dependent methanogenesis.

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Competition between Metals for Binding to Methanobactin Enables Expression of Soluble Methane Monooxygenase in the Presence of Copper

Applied and Environmental Microbiology ◽

10.1128/aem.03151-14 ◽

2014 ◽

Vol 81 (3) ◽

pp. 1024-1031 ◽

Cited By ~ 16

Author(s):

Bhagyalakshmi Kalidass ◽

Muhammad Farhan Ul-Haque ◽

Bipin S. Baral ◽

Alan A. DiSpirito ◽

Jeremy D. Semrau

Keyword(s):

Methane Monooxygenase ◽

High Specificity ◽

Copper Uptake ◽

Content Type ◽

Soluble Methane Monooxygenase ◽

Sds Page ◽

Genes Encoding ◽

Key Factor ◽

Membrane Bound

ABSTRACTIt is well known that copper is a key factor regulating expression of the two forms of methane monooxygenase found in proteobacterial methanotrophs. Of these forms, the cytoplasmic, or soluble, methane monooxygenase (sMMO) is expressed only at low copper concentrations. The membrane-bound, or particulate, methane monooxygenase (pMMO) is constitutively expressed with respect to copper, and such expression increases with increasing copper. Recent findings have shown that copper uptake is mediated by a modified polypeptide, or chalkophore, termed methanobactin. Although methanobactin has high specificity for copper, it can bind other metals, e.g., gold. Here we show that inMethylosinus trichosporiumOB3b, sMMO is expressed and active in the presence of copper if gold is also simultaneously present. Such expression appears to be due to gold binding to methanobactin produced byM. trichosporiumOB3b, thereby limiting copper uptake. Such expression and activity, however, was significantly reduced if methanobactin preloaded with copper was also added. Further, quantitative reverse transcriptase PCR (RT-qPCR) of transcripts of genes encoding polypeptides of both forms of MMO and SDS-PAGE results indicate that both sMMO and pMMO can be expressed when copper and gold are present, as gold effectively competes with copper for binding to methanobactin. Such findings suggest that under certain geochemical conditions, both forms of MMO may be expressed and activein situ. Finally, these findings also suggest strategies whereby field sites can be manipulated to enhance sMMO expression, i.e., through the addition of a metal that can compete with copper for binding to methanobactin.

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Efficient Production of 2,5-Diketo-d-Gluconate via Heterologous Expression of 2-Ketogluconate Dehydrogenase in Gluconobacter japonicus

Applied and Environmental Microbiology ◽

10.1128/aem.04176-14 ◽

2015 ◽

Vol 81 (10) ◽

pp. 3552-3560 ◽

Cited By ~ 14

Author(s):

Naoya Kataoka ◽

Minenosuke Matsutani ◽

Toshiharu Yakushi ◽

Kazunobu Matsushita

Keyword(s):

Mutant Strain ◽

Gluconobacter Oxydans ◽

Efficient Production ◽

Broad Host Range ◽

Putative Promoter Region ◽

Content Type ◽

Biotransformation Process ◽

Genes Encoding ◽

Membrane Bound ◽

Almost All

ABSTRACT2,5-Diketo-d-gluconate (2,5DKG) is a compound that can be the intermediate ford-tartrate and also vitamin C production. AlthoughGluconobacter oxydansNBRC3293 produces 2,5DKG fromd-glucose viad-gluconate and 2-keto-d-gluconate (2KG), with accumulation of the product in the culture medium, the efficiency of 2,5DKG production is unsatisfactory because there is a large amount of residuald-gluconate at the end of the biotransformation process. Oxidation of 2KG to 2,5DKG is catalyzed by a membrane-bound flavoprotein-cytochromeccomplex: 2-keto-gluconate dehydrogenase (2KGDH). Here, we studied thekgdSLCgenes encoding 2KGDH inG. oxydansNBRC3293 to improve 2,5DKG production byGluconobacterspp. ThekgdS,kgdL, andkgdCgenes correspond to the small, large, and cytochrome subunits of 2KGDH, respectively. ThekgdSLCgenes were cloned into a broad-host-range vector carrying a DNA fragment of the putative promoter region of the membrane-bound alcohol dehydrogenase gene ofG. oxydansfor expression inGluconobacterspp. According to our results, 2KGDH that was purified from the recombinantGluconobactercells showed characteristics nearly the same as those reported previously. We also expressed thekgdSLCgenes in a mutant strain ofGluconobacter japonicusNBRC3271 (formerlyGluconobacter dioxyacetonicusIFO3271) engineered to produce 2KG efficiently from a mixture ofd-glucose andd-gluconate. This mutant strain consumed almost all of the starting materials (d-glucose andd-gluconate) to produce 2,5DKG quantitatively as a seemingly unique metabolite. To our knowledge, this is the first report of aGluconobacterstrain that produces 2,5DKG efficiently and homogeneously.

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Methylomonas koyamae sp. nov., a type I methane-oxidizing bacterium from floodwater of a rice paddy field

INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY ◽

10.1099/ijs.0.035261-0 ◽

2012 ◽

Vol 62 (Pt_8) ◽

pp. 1832-1837 ◽

Cited By ~ 41

Author(s):

Takuya Ogiso ◽

Chihoko Ueno ◽

Dayéri Dianou ◽

Tran Van Huy ◽

Arata Katayama ◽

...

Keyword(s):

Paddy Field ◽

Rice Paddy ◽

Rrna Gene ◽

Type I ◽

Intracytoplasmic Membrane ◽

Content Type ◽

Rice Paddy Field ◽

Link Type ◽

Gene Sequence Analysis ◽

Bacterium Strain

A novel methane-oxidizing bacterium, strain Fw12E-YT, was isolated from floodwater of a rice paddy field in Japan. Cells of strain Fw12E-YT were Gram-negative, motile rods with a single polar flagellum and type I intracytoplasmic membrane arrangement. The strain grew only on methane or methanol as sole carbon and energy source. It was able to grow at 10–40 °C (optimum 30 °C), at pH 5.5–7.0 (optimum 6.5) and with 0–0.1 % (w/w) NaCl (no growth above 0.5 % NaCl). 16S rRNA gene sequence analysis showed that strain Fw12E-YT is related most closely to members of the genus Methylomonas , but at low levels of similarity (95.0–95.4 %). Phylogenetic analysis of pmoA and mxaF genes indicated that the strain belongs to the genus Methylomonas (97 and 92 % deduced amino acid sequence identities to Methylomonas methanica S1T, respectively). The DNA G+C content of strain Fw12E-YT was 57.1 mol%. Chemotaxonomic data regarding the major quinone (MQ-8) and major fatty acids (C16 : 1 and C14 : 0) also supported its affiliation to the genus Methylomonas . Based on phenotypic, genomic and phylogenetic data, strain Fw12E-YT is considered to represent a novel species of the genus Methylomonas , for which the name Methylomonas koyamae sp. nov. is proposed. The type strain is Fw12E-YT ( = JCM 16701T = NBRC 105905T = NCIMB 14606T).

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Two-Component Flavin-Dependent Riboflavin Monooxygenase Degrades Riboflavin inDevosia riboflavina

Journal of Bacteriology ◽

10.1128/jb.00022-18 ◽

2018 ◽

Vol 200 (12) ◽

Cited By ~ 2

Author(s):

Hiroshi Kanazawa ◽

Ryosuke Shigemoto ◽

Yukie Kawasaki ◽

Ken-Ichi Oinuma ◽

Akira Nakamura ◽

...

Keyword(s):

Initial Step ◽

Amino Acid Identity ◽

Flavin Mononucleotide ◽

Flavin Reductase ◽

Environmental Niche ◽

Content Type ◽

Monooxygenase Gene ◽

Adaptive Mechanisms ◽

Two Component ◽

Genes Encoding

ABSTRACTThe actinobacteriumMicrobacterium maritypicumsplits riboflavin (vitamin B2) into lumichrome andd-ribose. However, such degradation by other bacteria and the involvement of a two-component flavin-dependent monooxygenase (FMO) in the reaction remain unknown. Here we investigated the mechanism of riboflavin degradation by the riboflavin-assimilating alphaproteobacteriumDevosia riboflavina(formerlyPseudomonas riboflavina). We found that adding riboflavin to bacterial cultures induced riboflavin-degrading activity and a protein of the FMO family that had 67% amino acid identity with the predicted riboflavin hydrolase (RcaE) ofM. maritypicumMF109. TheD. riboflavinagenome clustered genes encoding the predicted FMO, flavin reductase (FR), ribokinase, and flavokinase, and riboflavin induced their expression. This finding suggests that these genes constitute a mechanism for utilizing riboflavin as a carbon source. Recombinant FMO (rFMO) protein ofD. riboflavinaoxidized riboflavin in the presence of reduced flavin mononucleotide (FMN) provided by recombinant FR (rFR), oxidized FMN and NADH, and produced stoichiometric amounts of lumichrome andd-ribose. Further investigation of the enzymatic properties ofD. riboflavinarFMO indicated that rFMO-rFR coupling accompanied O2consumption and the generation of enzyme-bound hydroperoxy-FMN, which are characteristic of two-component FMOs. These results suggest thatD. riboflavinaFMO is involved in hydroperoxy-FMN-dependent mechanisms to oxygenize riboflavin and a riboflavin monooxygenase is necessary for the initial step of riboflavin degradation.IMPORTANCEWhether bacteria utilize either a monooxygenase or a hydrolase for riboflavin degradation has remained obscure. The present study found that a novel riboflavin monooxygenase, not riboflavin hydrolase, facilitated this process inD. riboflavina. The riboflavin monooxygenase gene was clustered with flavin reductase, flavokinase, and ribokinase genes, and riboflavin induced their expression and riboflavin-degrading activity. The gene cluster is uniquely distributed inDevosiaspecies and actinobacteria, which have exploited an environmental niche by developing adaptive mechanisms for riboflavin utilization.

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New Mode of Energy Metabolism in the Seventh Order of Methanogens as Revealed by Comparative Genome Analysis of “Candidatus Methanoplasma termitum”

Applied and Environmental Microbiology ◽

10.1128/aem.03389-14 ◽

2014 ◽

Vol 81 (4) ◽

pp. 1338-1352 ◽

Cited By ~ 128

Author(s):

Kristina Lang ◽

Jörg Schuldes ◽

Andreas Klingl ◽

Anja Poehlein ◽

Rolf Daniel ◽

...

Keyword(s):

Energy Metabolism ◽

Physiological Data ◽

Dual Function ◽

The Novel ◽

Heterodisulfide Reductase ◽

Content Type ◽

Methanosphaera Stadtmanae ◽

Seventh Order ◽

Membrane Bound ◽

Energy Converting

ABSTRACTThe recently discovered seventh order of methanogens, theMethanomassiliicoccales(previously referred to as “Methanoplasmatales”), so far consists exclusively of obligately hydrogen-dependent methylotrophs. We sequenced the complete genome of “CandidatusMethanoplasma termitum” from a highly enriched culture obtained from the intestinal tract of termites and compared it with the previously published genomes of three other strains from the human gut, including the first isolate of the order. Like all other strains, “Ca. Methanoplasma termitum” lacks the entire pathway for CO2reduction to methyl coenzyme M and produces methane by hydrogen-dependent reduction of methanol or methylamines, which is consistent with additional physiological data. However, the shared absence of cytochromes and an energy-converting hydrogenase for the reoxidation of the ferredoxin produced by the soluble heterodisulfide reductase indicates thatMethanomassiliicoccalesemploy a new mode of energy metabolism, which differs from that proposed for the obligately methylotrophicMethanosphaera stadtmanae. Instead, all strains possess a novel complex that is related to the F420:methanophenazine oxidoreductase (Fpo) ofMethanosarcinalesbut lacks an F420-oxidizing module, resembling the apparently ferredoxin-dependent Fpo-like homolog inMethanosaeta thermophila. Since allMethanomassiliicoccalesalso lack the subunit E of the membrane-bound heterodisulfide reductase (HdrDE), we propose that the Fpo-like complex interacts directly with subunit D, forming an energy-converting ferredoxin:heterodisulfide oxidoreductase. The dual function of heterodisulfide inMethanomassiliicoccales, which serves both in electron bifurcation and as terminal acceptor in a membrane-associated redox process, may be a unique characteristic of the novel order.

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Transcriptional Regulation of Genes Encoding the Selenium-Free [NiFe]-Hydrogenases in the Archaeon Methanococcus voltae Involves Positive and Negative Control Elements

Genetics ◽

10.1093/genetics/152.4.1335 ◽

1999 ◽

Vol 152 (4) ◽

pp. 1335-1341

Author(s):

Izabela Noll ◽

Steffen Müller ◽

Albrecht Klein

Keyword(s):

Intergenic Region ◽

Genetic Information ◽

Regulatory Element ◽

Transcription Unit ◽

Negative Control ◽

Negative Regulation ◽

Negative Regulatory Element ◽

Methanococcus Voltae ◽

Genes Encoding ◽

Made In

Abstract Methanococcus voltae harbors genetic information for two pairs of homologous [NiFe]-hydrogenases. Two of the enzymes contain selenocysteine, while the other two gene groups encode apparent isoenzymes that carry cysteinyl residues in the homologous positions. The genes coding for the selenium-free enzymes, frc and vhc, are expressed only under selenium limitation. They are transcribed out of a common intergenic region. A series of deletions made in the intergenic region localized a common negative regulatory element for the vhc and frc promoters as well as two activator elements that are specific for each of the two transcription units. Repeated sequences, partially overlapping the frc promoter, were also detected. Mutations in these repeated heptanucleotide sequences led to a weak induction of a reporter gene under the control of the frc promoters in the presence of selenium. This result suggests that the heptamer repeats contribute to the negative regulation of the frc transcription unit.

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Functional soil organic matter fractions in response to long-term fertilizer management in a double-cropping paddy field of southern China

The Journal of Agricultural Science ◽

10.1017/s0021859621000125 ◽

2021 ◽

pp. 1-9

Author(s):

Haiming Tang ◽

Chao Li ◽

Lihong Shi ◽

Li Wen ◽

Kaikai Cheng ◽

...

Keyword(s):

Organic Matter ◽

Soil Organic Matter ◽

Paddy Field ◽

Southern China ◽

Rice Paddy ◽

Chemical Fertilizer ◽

Protection Mechanism ◽

Rice Paddy Field ◽

Double Cropping

Abstract Soil organic matter (SOM) and its fractions play an important role in maintaining or improving soil quality and soil fertility. Therefore, the effects of a 34-year long-term fertilizer regime on six functional SOM fractions under a double-cropping rice paddy field of southern China were studied in the current paper. The field experiment included four different fertilizer treatments: chemical fertilizer alone (MF), rice straw residue and chemical fertilizer (RF), 30% organic manure and 70% chemical fertilizer (OM) and without fertilizer input as control (CK). The results showed that coarse unprotected particulate organic matter (cPOM), biochemically, physically–biochemically and chemically protected silt-sized fractions (NH-dSilt, NH-μSilt and H-dSilt) were the main carbon (C) storage fractions under long-term fertilization conditions, accounting for 16.7–26.5, 31.1–35.6, 16.2–17.3 and 7.5–8.2% of the total soil organic carbon (SOC) content in paddy soil, respectively. Compared with control, OM treatment increased the SOC content in the cPOM, fine unprotected POM fraction, pure physically protected fraction and physico-chemically protected fractions by 58.9, 106.7, 117.6 and 28.3%, respectively. The largest proportion of SOC to total SOC in the different fractions was biochemically protected, followed by chemically and unprotected, and physically protected were the smallest. These results suggested that a physical protection mechanism plays an important role in stabilizing C of paddy soil. In summary, the results showed that higher functional SOM fractions and physical protection mechanism play an important role in SOM cycling in terms of C sequestration under the double-cropping rice paddy field.

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Methanobactin from Methylocystis sp. Strain SB2 Affects Gene Expression and Methane Monooxygenase Activity in Methylosinus trichosporium OB3b

Applied and Environmental Microbiology ◽

10.1128/aem.03981-14 ◽

2015 ◽

Vol 81 (7) ◽

pp. 2466-2473 ◽

Cited By ~ 11

Author(s):

Muhammad Farhan Ul-Haque ◽

Bhagyalakshmi Kalidass ◽

Alexey Vorobev ◽

Bipin S. Baral ◽

Alan A. DiSpirito ◽

...

Keyword(s):

Gene Expression ◽

Methane Monooxygenase ◽

Particulate Methane Monooxygenase ◽

Methylosinus Trichosporium Ob3b ◽

Methylosinus Trichosporium ◽

Monooxygenase Activity ◽

Content Type ◽

Soluble Methane Monooxygenase ◽

A Subunit ◽

Membrane Bound

ABSTRACTMethanotrophs can express a cytoplasmic (soluble) methane monooxygenase (sMMO) or membrane-bound (particulate) methane monooxygenase (pMMO). Expression of these MMOs is strongly regulated by the availability of copper. Many methanotrophs have been found to synthesize a novel compound, methanobactin (Mb), that is responsible for the uptake of copper, and methanobactin produced byMethylosinus trichosporiumOB3b plays a key role in controlling expression of MMO genes in this strain. As all known forms of methanobactin are structurally similar, it was hypothesized that methanobactin from one methanotroph may alter gene expression in another. WhenMethylosinus trichosporiumOB3b was grown in the presence of 1 μM CuCl2, expression ofmmoX, encoding a subunit of the hydroxylase component of sMMO, was very low.mmoXexpression increased, however, when methanobactin fromMethylocystissp. strain SB2 (SB2-Mb) was added, as did whole-cell sMMO activity, but there was no significant change in the amount of copper associated withM. trichosporiumOB3b. IfM. trichosporiumOB3b was grown in the absence of CuCl2, themmoXexpression level was high but decreased by several orders of magnitude if copper prebound to SB2-Mb (Cu-SB2-Mb) was added, and biomass-associated copper was increased. Exposure ofMethylosinus trichosporiumOB3b to SB2-Mb had no effect on expression ofmbnA, encoding the polypeptide precursor of methanobactin in either the presence or absence of CuCl2.mbnAexpression, however, was reduced when Cu-SB2-Mb was added in both the absence and presence of CuCl2. These data suggest that methanobactin acts as a general signaling molecule in methanotrophs and that methanobactin “piracy” may be commonplace.

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Craniofacial Diseases Caused by Defects in Intracellular Trafficking

Genes ◽

10.3390/genes12050726 ◽

2021 ◽

Vol 12 (5) ◽

pp. 726

Author(s):

Chung-Ling Lu ◽

Jinoh Kim

Keyword(s):

Endoplasmic Reticulum ◽

Membrane Proteins ◽

Intracellular Trafficking ◽

Critical Role ◽

Treatment Strategies ◽

Soluble Proteins ◽

Genes Encoding ◽

Membrane Bound ◽

Signaling Receptors ◽

Craniofacial Morphogenesis

Cells use membrane-bound carriers to transport cargo molecules like membrane proteins and soluble proteins, to their destinations. Many signaling receptors and ligands are synthesized in the endoplasmic reticulum and are transported to their destinations through intracellular trafficking pathways. Some of the signaling molecules play a critical role in craniofacial morphogenesis. Not surprisingly, variants in the genes encoding intracellular trafficking machinery can cause craniofacial diseases. Despite the fundamental importance of the trafficking pathways in craniofacial morphogenesis, relatively less emphasis is placed on this topic, thus far. Here, we describe craniofacial diseases caused by lesions in the intracellular trafficking machinery and possible treatment strategies for such diseases.

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Genes Required for Aerial Growth, Cell Division, and Chromosome Segregation Are Targets of WhiA before Sporulation in Streptomyces venezuelae

mBio ◽

10.1128/mbio.00684-13 ◽

2013 ◽

Vol 4 (5) ◽

Cited By ~ 61

Author(s):

Matthew J. Bush ◽

Maureen J. Bibb ◽

Govind Chandra ◽

Kim C. Findlay ◽

Mark J. Buttner

Keyword(s):

Cell Division ◽

Chromosome Segregation ◽

Regulatory Networks ◽

Liquid Culture ◽

Biological Processes ◽

Streptomyces Venezuelae ◽

Content Type ◽

Master Regulators ◽

Aerial Growth ◽

Genes Encoding

ABSTRACTWhiA is a highly unusual transcriptional regulator related to a family of eukaryotic homing endonucleases. WhiA is required for sporulation in the filamentous bacteriumStreptomyces, but WhiA homologues of unknown function are also found throughout the Gram-positive bacteria. To better understand the role of WhiA inStreptomycesdevelopment and its function as a transcription factor, we identified the WhiA regulon through a combination of chromatin immunoprecipitation-sequencing (ChIP-seq) and microarray transcriptional profiling, exploiting a new model organism for the genus,Streptomyces venezuelae, which sporulates in liquid culture. The regulon encompasses ~240 transcription units, and WhiA appears to function almost equally as an activator and as a repressor. Bioinformatic analysis of the upstream regions of the complete regulon, combined with DNase I footprinting, identified a short but highly conserved asymmetric sequence, GACAC, associated with the majority of WhiA targets. Construction of a null mutant showed thatwhiAis required for the initiation of sporulation septation and chromosome segregation inS. venezuelae, and several genes encoding key proteins of theStreptomycescell division machinery, such asftsZ,ftsW, andftsK, were found to be directly activated by WhiA during development. Several other genes encoding proteins with important roles in development were also identified as WhiA targets, including the sporulation-specific sigma factor σWhiGand the diguanylate cyclase CdgB. Cell division is tightly coordinated with the orderly arrest of apical growth in the sporogenic cell, andfilP, encoding a key component of the polarisome that directs apical growth, is a direct target for WhiA-mediated repression during sporulation.IMPORTANCESince the initial identification of the genetic loci required forStreptomycesdevelopment, all of thebldandwhidevelopmental master regulators have been cloned and characterized, and significant progress has been made toward understanding the cell biological processes that drive morphogenesis. A major challenge now is to connect the cell biological processes and the developmental master regulators by dissecting the regulatory networks that link the two. Studies of these regulatory networks have been greatly facilitated by the recent introduction ofStreptomyces venezuelaeas a new model system for the genus, a species that sporulates in liquid culture. Taking advantage ofS. venezuelae, we have characterized the regulon of genes directly under the control of one of these master regulators, WhiA. Our results implicate WhiA in the direct regulation of key steps in sporulation, including the cessation of aerial growth, the initiation of cell division, and chromosome segregation.

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