記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：CELL PRESS  

Condensins load onto DNA to organize chromosomes. Smc-ScpAB clearly loads onto the parS sites bound by Spo0J, but other loading site(s) must operate independently of parS. In this study, we asked where and how Smc-ScpAB normally selects its loading site. Our results suggest that rDNA is also a loading site. A pull-down assay revealed that Smc-ScpAB preferentially loads onto rDNA in the wildtype cell and even in a Delta spo0J mutant but not in a Delta smc mutant. Moreover, we showed that deletion mutants of rDNAs cause a defect in nucleoid separation, and at least two rDNAs near oriC are essential for separation. Full-length rDNA, including promoters, is required for loading and nucleoid separation. A synthetic defect by deletions of both rDNA and spo0J resulted in more aberrant nucleoid separation. We propose that a single-stranded segment of DNA that is exposed at highly transcribed rRNA operons would become a target for Smc-ScpAB loading.

DOI： 10.1016/j.celrep.2017.10.014

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：NATURE PUBLISHING GROUP  

Condensin is the major driving force in the segregation of daughter chromosomes in prokaryotes. Core subunits of condensin belong to the SMC protein family, whose members are characterized by a unique ATPase activity and dimers with a V-shaped structure. The V-shaped dimers might close between head domains, forming a ring structure that can encircle DNA. Indeed, cohesin, which is a subfamily of SMC proteins, encircles double-stranded DNA to hold sister chromatids in eukaryotes. However, the question of whether or not condensin encircles the chromosomal DNA remains highly controversial. Here we report that MukB binds topologically to DNA in vitro, and this binding is preferentially single-stranded DNA (ssDNA) rather than double-stranded DNA. The binding of MukB to ssDNA does not require ATP. In fact, thermal energy enhances the binding. The non-SMC subunits MukF and MukE did stimulate the topological binding of MukB, although they hindered DNA-binding of MukB. Recent reports on the distribution of condensin in genomes reveal that actively transcribed genes in yeast and humans are enriched in condensin. In consideration of all these results, we propose that the binding specificity of condensin to chromosome is provided not by the DNA sequence but by the DNA structure, which is ssDNA.

DOI： 10.1038/srep29469

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：MICROBIOLOGY SOC  

Ribosome dimers are a translationally inactive form of ribosomes found in Escherichia coli and many other bacterial cells. In this study, we found that the 70S ribosomes of Bacillus subtilis dimerized during the early stationary phase and these dimers remained in the cytoplasm until regrowth was initiated. Ribosome dimerization during the stationary phase required the hpf gene, which encodes a homologue of the E. coli hibernation-promoting factor (Hpf). The expression of hpf was induced at an early stationary phase and its expression was observed throughout the rest of the experimental period, including the entire 6 h of the stationary phase. Ribosome dimerization followed the induction of hpf in WT cells, but the dimerization was impaired in cells harbouring a deletion in the hpf gene. Although the absence of ribosome dimerization in these Hpf-deficient cells did not affect their viability in the stationary phase, their ability to regrow from the stationary phase decreased. Thus, following the transfer of stationary-phase cells to fresh LB medium, Delta hpf mutant cells grew slower than WT cells. This observed lag in growth of Delta hpf cells was probably due to a delay in restoring their translational activity. During regrowth, the abundance of ribosome dimers in WT cells decreased with a concomitant increase in the abundance of 70S ribosomes and growth rate. These results suggest that the ribosome dimers, by providing 70S ribosomes to the cells, play an important role in facilitating rapid and efficient regrowth of cells under nutrient-rich conditions.

DOI： 10.1099/mic.0.000234

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：SOC GENERAL MICROBIOLOGY  

The genome of Bacillus subtilis strain 168 encodes ten rRNA (rrn) operons. We previously reported that strains with only a single rrn operon had a decreased growth and sporulation frequency. We report here the isolation and characterization of suppressor mutants from seven strains that each have a single rrn operon (rrnO, A, J, 1, E, D or B). The suppressor mutants for strain RIK656 with a single rrnO operon had a higher frequency of larger colonies. These suppressor mutants had not only increased growth rates, but also increased sporulation frequencies and ribosome levels compared to the parental mutant strain RIK656. Quantitative PCR analyses showed that all these suppressor mutants had an increased number of copies of the rrnO operon. Suppressor mutants were also isolated from the six other strains with single rrn operons (rrnA, J, 1, E, D or B). Next generation and capillary sequencing showed that all of the suppressor mutants had tandem repeats of the chromosomal locus containing the remaining rrn operon (amplicon). These amplicons varied in size from approximately 9 to 179 kb. The amplifications were likely to be initiated by illegitimate recombination between non- or micro-homologous sequences, followed by unequal crossing-over during DNA replication. These results are consistent with our previous report that rrn operon copy number has a major role in cellular processes such as cell growth and sporulation.

DOI： 10.1099/mic.0.000207

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：SOC GENERAL MICROBIOLOGY  

The number of copies of rRNA (rrn) operons in a bacterial genome differs greatly among bacterial species. Here we examined the phenotypic effects of variations in the number of copies of rRNA genes in the genome of Bacillus subtilis by analysis of eight mutant strains constructed to carry from two to nine copies of the rrn operon. We found that a decrease in the number of copies from ten to one increased the doubling time, and decreased the sporulation frequency and motility. The maximum levels for transformation activity were similar among the strains, although the competence development was significantly delayed in the strain with a single rrn operon. Normal sporulation only occurred if more than four copies of the rrn operon were present, although ten copies were needed for vegetative growth after germination of the spores. This behaviour was seen even though the intracellular level of ribosomes was similar among strains with four to ten copies of the rrn operon. Furthermore, ten copies of the rrn operon were needed for the highest swarming activity. We also constructed 21 strains that carried all possible combinations of two copies of the rrn operons, and found that these showed a range of growth rates and sporulation frequencies that all fell between those recorded for strains with one or three copies of the rrn operon. The results suggested that the copy number of the rrn operon has a major influence on cellular processes such as growth rate and sporulation frequency.

DOI： 10.1099/mic.0.067025-0

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：4  

Bacillus subtilis contains 10 rRNA (rrn) operons. We found that rRNA operon-engineered B. subtilis strain RIK543, with only the rrnO operon, is specifically hypersensitive to RNA polymerase inhibitors such as rifamycin SV and rifampin (80-fold and 20-fold, respectively). In pilot screening experiments, we found actinomycete isolates successfully at an incidence of 1.9% (18/945) that produced antibacterials that were detectable only with RIK543 as the test organism. Strain RIK543 may be a feasible test organism for the discovery of novel RNA polymerase inhibitors. Copyright © 2013, American Society for Microbiology. All Rights Reserved.

DOI： 10.1128/AAC.02604-12

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：MICROBIOL RES FOUNDATION  

We introduced single mutations into the rplC and rpsJ genes, which encode the essential ribosomal proteins L3 (RplC) and S10 (RpsJ), respectively, and are located in the S10 gene cluster of the gram-positive, endospore-forming bacterium Bacillus subtilis, and examined whether these mutations affected their growth rate, sporulation, competence development and 70S ribosome formation. Mutant cells harboring the G52D mutation in the L3 ribosomal protein, which is located at the peptidyl transferase center of 50S, accumulated 30S subunit at 45 degrees C, probably due to a defect in 50S formation, and exhibited a reduction in the sporulation frequency at high temperature. On the other hand, mutant cells harboring the H56R mutation in the S10 protein, which is located near the aminoacyl-tRNA site of 30S, showed severe growth defect and deficiency in spore formation, and also exhibited significant delay in competence development.

DOI： 10.2323/jgam.59.105

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：AMER SOC MICROBIOLOGY  

Among the 57 genes that encode ribosomal proteins in the genome of Bacillus subtilis, a Gram-positive bacterium, 50 genes were targeted by systematic inactivation. Individual deletion mutants of 16 ribosomal proteins (L1, L9, L15, L22, L23, L28, L29, L32, L33.1, L33.2, L34, L35, L36, S6, S20, and S21) were obtained successfully. In conjunction with previous reports, 22 ribosomal proteins have been shown to be nonessential in B. subtilis, at least for cell proliferation. Although several mutants that harbored a deletion of a ribosomal protein gene did not show any significant differences in any of the phenotypes that were tested, various mutants showed a reduced growth rate and reduced levels of 70S ribosomes compared with the wild type. In addition, severe defects in the sporulation frequency of the Delta rplA (L1) mutant and the motility of the Delta rpsU (S21) mutant were observed. These data provide the first evidence in B. subtilis that L1 and S21 are required for the progression of cellular differentiation.

DOI： 10.1128/JB.01544-12

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：MICROBIOL RES FOUNDATION  

DOI： 10.2323/jgam.57.153

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：TAYLOR & FRANCIS LTD  

A nucleotide sequence of the whole genome of Bacillus subtilis phage SP10 was determined. It was composed of 143,986 bp with 236 putative open reading frames (ORFs). Sixty-five of 236 predicted ORFs showed high similarity to that of SPO1, and the genome organizations of the two phages were similar to each other. SP10 belongs to the Myoviridae family, for which the well-studied phage SPO1 is the representative phage. Hence, we compared SP10 to SPO1. The SP10 genome DNA showed different sensitivity to restriction enzymes than SPO1, due to differences in base modification. According to transcriptional analysis, the gene expression of regulatory network of SP10 was similar to SPO1. It was observed that RNA polymerase containing sigma-A was inactive in directing the host genes but active in directing the phage genes. It appeared that the association of sigma-A with the core enzyme complex of RNA polymerase was strengthened during development.

DOI： 10.1271/bbb.100921

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：TAYLOR & FRANCIS LTD  

Pairs of the ECF sigma factor and its anti-sigma factor, SigW and RsiW, of Bacillus-related species that inhabit extreme environments were heterologously expressed in B. subtilis. All the RsiWs, membrane proteins, failed to fill their function of repressing cognate SigW activity, despite their close structural similarities. Particularly, uncontrolled expression of Oceanobacillus iheyensis OISigW due to abortive OIRsiW was harmful to B. subtilis. Analysis of revertants of this growth defect and site-directed mutagenesis indicated that the insertion of six and a minimum of three hydrophobic amino acid residues occurring in the transmembrane region allowed OIRsiW to function as anti-OISigW. Subcellular localization of OIRsiW was detected by immunoblot analysis, suggesting that both the wild-type and the mutant form of OIRsiW were localized to the membrane. An appropriate length of a transmembrane region required for proper integration into the membrane after translocation might vary among these Bacillus-related species.

DOI： 10.1271/bbb.110035

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記述言語：英語   掲載種別：研究論文（学術雑誌）   出版者・発行元：1  

DOI： 10.1266/ggs.86.7

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記述言語：英語   掲載種別：研究発表ペーパー・要旨（国際会議）   出版者・発行元：GENETICS SOC JAPAN  

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記述言語：英語   掲載種別：研究発表ペーパー・要旨（国際会議）   出版者・発行元：GENETICS SOC JAPAN  

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記述言語：英語   掲載種別：研究発表ペーパー・要旨（国際会議）   出版者・発行元：GENETICS SOC JAPAN  

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記述言語：英語   掲載種別：研究発表ペーパー・要旨（国際会議）   出版者・発行元：GENETICS SOC JAPAN  

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記述言語：英語   掲載種別：研究発表ペーパー・要旨（国際会議）   出版者・発行元：GENETICS SOC JAPAN  

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開催年月日： 2017年5月21日 - 2017年5月26日

記述言語：英語   会議種別：口頭発表（一般）  

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