Complete Genome Sequences of Bacillus subtilis subsp.
            <i>subtilis</i>
            Laboratory Strains JH642 (AG174) and AG1839

Smith, Janet L.; Goldberg, Jonathan M.; Grossman, Alan D.

doi:10.1128/genomea.00663-14

Cited by 46 publications

(74 citation statements)

References 15 publications

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“…Although the sporulation efficiencies of the 11 independently isolated suppressor strains were only moderately (threefold to eightfold) increased (Tables and ), we mapped the mutations in two of them by whole genome resequencing. Strikingly, in each of these two suppressors only a single nucleotide polymorphism was observed in comparison to the JH642 reference genome (Smith et al ., ). In strain NWB6, we identified a point mutation in the promoter region of kinA (Table ), and in strain NWB7 a point mutation in the coding region of the sigH gene was present that resulted in an amino acid substitution (E146A) in the σ H protein (Table ).…”

Section: Resultsmentioning

confidence: 97%

“…To investigate whether the inhibition of sporulation by high salinity was specific to the wild‐type strain JH642 (Smith et al ., ), we examined the sporulation efficiency of both domesticated and undomesticated B. subtilis isolates. Although sporulation efficiency in the presence of salt was somewhat variable, all strains tested displayed salt sensitivity (Fig.…”

Section: Resultsmentioning

confidence: 99%

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Salt‐sensitivity of σ^H and Spo0A prevents sporulation of Bacillus subtilis at high osmolarity avoiding death during cellular differentiation

Widderich

Rodrigues

Commichau

et al. 2016

Molecular Microbiology

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Summary The spore-forming bacterium Bacillus subtilis frequently experiences high osmolarity as a result of desiccation in the soil. The formation of a highly desiccation-resistant endospore might serve as a logical osmostress escape route when vegetative growth is no longer possible. However, sporulation efficiency drastically decreases concomitant with an increase in the external salinity. Fluorescence microscopy of sporulation-specific promoter fusions to gfp revealed that high salinity blocks entry into the sporulation pathway at a very early stage. Specifically, we show that both Spo0A- and SigH-dependent transcription are impaired. Furthermore, we demonstrate that the association of SigH with core RNA polymerase is reduced under these conditions. Suppressors that modestly increase sporulation efficiency at high salinity map to the coding region of sigH and in the regulatory region of kinA, encoding one the sensor kinases that activates Spo0A. These findings led us to discover that B. subtilis cells that overproduce KinA can bypass the salt-imposed block in sporulation. Importantly, these cells are impaired in the morphological process of engulfment and late forespore gene expression and frequently undergo lysis. Altogether our data indicate that B. subtilis blocks entry into sporulation in high-salinity environments preventing commitment to a developmental program that it cannot complete.

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Section: Resultsmentioning

confidence: 97%

Section: Resultsmentioning

confidence: 99%

Salt‐sensitivity of σ^H and Spo0A prevents sporulation of Bacillus subtilis at high osmolarity avoiding death during cellular differentiation

Widderich

Rodrigues

Commichau

et al. 2016

Molecular Microbiology

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“…Amino acids, the ninhydrin reagent for the quantification of proline by a colorimetric assay (35), and the antibiotics chloramphenicol, kanamycin, tetracycline, and spectinomycin were all purchased from Sigma-Aldrich (Steinheim, Germany) or from Carl Roth GmbH (Karlsruhe, Germany Bacterial strains, media, and growth conditions. The genetic properties of the B. subtilis strains used in this study are summarized in Table 1; they are all derived from the laboratory strain JH642 (36,37). B. subtilis strains were routinely cultivated in Spizizen=s minimal medium (SMM) (38) with 0.5% (wt/vol) glucose as the carbon source and L-tryptophan (20 mg liter Ϫ1 ) and L-phenylalanine (18 mg liter Ϫ1 ) to satisfy the auxotrophic requirements of strain JH642 (trpC2 pheA1) and its mutant derivatives (Table 1).…”

Section: Methodsmentioning

confidence: 99%

Uptake of Amino Acids and Their Metabolic Conversion into the Compatible Solute Proline Confers Osmoprotection to Bacillus subtilis

Zaprasis

Bleisteiner

Kerres

et al. 2015

Appl Environ Microbiol

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bThe data presented here reveal a new facet of the physiological adjustment processes through which Bacillus subtilis can derive osmostress protection. We found that the import of proteogenic (Glu, Gln, Asp, Asn, and Arg) and of nonproteogenic (Orn and Cit) amino acids and their metabolic conversion into proline enhances growth under otherwise osmotically unfavorable conditions. Osmoprotection by amino acids depends on the functioning of the ProJ-ProA-ProH enzymes, but different entry points into this biosynthetic route are used by different amino acids to finally yield the compatible solute proline. Glu, Gln, Asp, and Asn are used to replenish the cellular pool of glutamate, the precursor for proline production, whereas Arg, Orn, and Cit are converted into ␥-glutamic semialdehyde/⌬ 1 -pyrroline-5-carboxylate, an intermediate in proline biosynthesis. The import of Glu, Gln, Asp, Asn, Arg, Orn, and Cit did not lead to a further increase in the size of the proline pool that is already present in osmotically stressed cells. Hence, our data suggest that osmoprotection of B. subtilis by this group of amino acids rests on the savings in biosynthetic building blocks and energy that would otherwise have to be devoted either to the synthesis of the proline precursor glutamate or of proline itself. Since glutamate is the direct biosynthetic precursor for proline, we studied its uptake and found that GltT, an Na ؉ -coupled symporter, is the main uptake system for both glutamate and aspartate in B. subtilis. Collectively, our data show how effectively B. subtilis can exploit environmental resources to derive osmotic-stress protection through physiological means. Bacillus subtilis is a resident of the upper layers of the soil and of the rhizosphere, and it can also efficiently colonize root surfaces (1-3). The blueprint of its genome (4) bears the hallmarks of a bacterium that can exploit many plant-produced compounds for its growth. Accordingly, a considerable portion of the coding capacity of the B. subtilis chromosome (5) is devoted to highaffinity import systems (6) that allow the scavenging of a wide spectrum of nutrients. Reoccurring and persisting high osmolarity in the soil ecosystem (7) is a situation in which B. subtilis can take advantage of the import of plant-produced compounds (8-10) for its physiological adjustment to these unfavorable environmental conditions (7, 11).As in many bacterial species (11-13), cellular adaptation of B. subtilis to both sudden and sustained increases in the external osmolarity involves a two-stage process (7,14). It initially encompasses the uptake of large quantities of potassium as an emergency stress reaction to curb water efflux (14, 15) and, subsequently, the replacement of part of this ion by organic osmolytes, such as proline (Pro) and glycine betaine (GB), to decrease the ionic strength of the cytoplasm and to optimize its solvent properties (14,(16)(17)(18)(19). These organic osmolytes, commonly referred to as compatible solutes, are highly compliant with cellular physiolog...

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“…The Tn916 genomic integration site was mapped by inverse PCR essentially as described previously (42). As in the parental strain (43), Tn916 is integrated between chromosomal genes yufK and yufL at coordinate 3209748 (36). Tn916 is oriented such that transcription of orf24 and int of Tn916 is codirectional with that of yufL.…”

Section: Methodsmentioning

confidence: 99%

“…subtilis strains were derived from JH642 (pheA1 trpC2) (35,36) and are listed in Table 1. Most strains were derived from JMA222, a derivative of JH642 that was cured of ICEBs1 (37).…”

Section: Methodsmentioning

confidence: 99%

Autonomous Replication of the Conjugative Transposon Tn916

2016

Self Cite

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Integrative and conjugative elements (ICEs), also known as conjugative transposons, are self-transferable elements that are widely distributed among bacterial phyla and are important drivers of horizontal gene transfer. Many ICEs carry genes that confer antibiotic resistances to their host cells and are involved in the dissemination of these resistance genes. ICEs reside in host chromosomes but under certain conditions can excise to form a plasmid that is typically the substrate for transfer. A few ICEs are known to undergo autonomous replication following activation. However, it is not clear if autonomous replication is a general property of many ICEs. We found that Tn916, the first conjugative transposon identified, replicates autonomously via a rolling-circle mechanism. Replication of Tn916 was dependent on the relaxase encoded by orf20 of Tn916. The origin of transfer of Tn916, oriT(916), also functioned as an origin of replication. Using immunoprecipitation and mass spectrometry, we found that the relaxase (Orf20) and the two putative helicase processivity factors (Orf22 and Orf23) encoded by Tn916 likely interact in a complex and that the Tn916 relaxase contains a previously unidentified conserved helix-turn-helix domain in its N-terminal region that is required for relaxase function and replication. Lastly, we identified a functional single-strand origin of replication (sso) in Tn916 that we predict primes second-strand synthesis during rolling-circle replication. Together these results add to the emerging data that show that several ICEs replicate via a conserved, rolling-circle mechanism. IMPORTANCE Integrative and conjugative elements (ICEs) drive horizontal gene transfer and the spread of antibiotic resistances in bacteria.ICEs reside integrated in a host genome but can excise to create a plasmid that is the substrate for transfer to other cells. Here we show that Tn916, an ICE with broad host range, undergoes autonomous rolling-circle replication when in the plasmid form. We found that the origin of transfer functions as a double-stranded origin of replication and identified a single-stranded origin of replication. It was long thought that ICEs do not undergo autonomous replication. Our work adds to the evidence that ICEs replicate autonomously as part of their normal life cycle and indicates that diverse ICEs use the same replicative mechanism. Integrative and conjugative elements (ICEs), also called conjugative transposons, are mobile genetic elements that encode proteins that mediate transfer of the element from the host cell (donor) to a recipient by conjugation. ICEs often contain additional (cargo) genes that can provide a selective advantage to the host cells (reviewed in references 1 and 2). Most ICEs have been identified based on their cargo genes and the phenotypes conferred. For example, many ICEs carry genes encoding antibiotic resistances. The horizontal dissemination of ICEs and their associated cargo genes is a major driver of bacterial genome plasticity and evolution and the s...

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Complete Genome Sequences of Bacillus subtilis subsp. subtilis Laboratory Strains JH642 (AG174) and AG1839

Cited by 46 publications

References 15 publications

Salt‐sensitivity of σ^H and Spo0A prevents sporulation of Bacillus subtilis at high osmolarity avoiding death during cellular differentiation

Salt‐sensitivity of σ^H and Spo0A prevents sporulation of Bacillus subtilis at high osmolarity avoiding death during cellular differentiation

Uptake of Amino Acids and Their Metabolic Conversion into the Compatible Solute Proline Confers Osmoprotection to Bacillus subtilis

Autonomous Replication of the Conjugative Transposon Tn916

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Complete Genome Sequences of Bacillus subtilis subsp. subtilis Laboratory Strains JH642 (AG174) and AG1839

Cited by 46 publications

References 15 publications

Salt‐sensitivity of σH and Spo0A prevents sporulation of Bacillus subtilis at high osmolarity avoiding death during cellular differentiation

Salt‐sensitivity of σH and Spo0A prevents sporulation of Bacillus subtilis at high osmolarity avoiding death during cellular differentiation

Uptake of Amino Acids and Their Metabolic Conversion into the Compatible Solute Proline Confers Osmoprotection to Bacillus subtilis

Autonomous Replication of the Conjugative Transposon Tn916

Contact Info

Product

Resources

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Salt‐sensitivity of σ^H and Spo0A prevents sporulation of Bacillus subtilis at high osmolarity avoiding death during cellular differentiation

Salt‐sensitivity of σ^H and Spo0A prevents sporulation of Bacillus subtilis at high osmolarity avoiding death during cellular differentiation