Cell-cell communication regulates the effects of protein aspartate phosphatases on the phosphorelay controlling development in Bacillus subtilis.

Rap phosphatases are a recently discovered family of protein aspartate phosphatases that dephosphorylate the Spo0F--P intermediate of the phosphorelay, thus preventing sporulation of Bacillus subtilis. They are regulators induced by physiological processes that are antithetical to sporulation. The RapA phosphatase is induced by the ComP-ComA two-component signal transduction system responsible for initiating competence. RapA phosphatase activity was found to be controlled by a small protein, PhrA, encoded on the same transcript as RapA. PhrA resembles secreted proteins and the evidence suggests that it is cleaved by signal peptidase I and a 19-residue C-terminal domain is secreted from the cell. The sporulation deficiency caused by the uncontrolled RapA activity of a phrA mutant can be complemented by synthetic peptides comprising the last six or more of the C-terminal residues of PhrA. Whether the peptide controls RapA activity directly or by regulating its synthesis remains to be determined. Complementation of the phrA mutant can also be obtained in mixed cultures with a wild-type strain, suggesting the peptide may serve as a means of communication between cells. Importation of the secreted peptide required the oligopeptide transport system. The sporulation deficiency of oligopeptide transport mutants can be suppressed by mutating the rapA and rapB genes or by introduction of a spo0F mutation Y13S that renders the protein insensitive to Rap phosphatases. The data indicate that the sporulation deficiency of oligopeptide transport mutants is due to their inability to import the peptides controlling Rap phosphatases.

Coordinate regulation of Bacillus subtilis peroxide stress genes by hydrogen peroxide and metal ions.

The Bacillus subtilis mrgA gene encodes an abundant DNA-binding protein that protects cells against the lethal effects of H2O2. Transcription of mrgA is induced by H2O2 or by entry into stationary phase when manganese and iron levels are low. We have selected for strains derepressed for transcription of mrgA in the presence of Mn(II). The resulting cis-acting mutants define an operator site just upstream of the mrgA promoter. Similar sequences flank the promoters for the catalase gene, katA, and the heme biosynthesis operon, hemAXCDBL. Like mrgA, transcription of the katA and hem genes is repressed by Mn(II), which thereby potentiates the killing action of H2O2. We identified two classes of trans-acting mutants derepressed for mrgA transcription in the presence of Mn(II): some exhibit a coordinate derepression of MrgA, catalase, heme biosynthesis, and alkyl hydroperoxide reductase and are H2O2 resistant, while others have reduced catalase activity and are H2O2 sensitive. These data indicate that the peroxide stress response of B. subtilis is regulated by a repressor that senses both metal ion levels and H2O2.

TRAP, the trp RNA-binding attenuation protein of Bacillus subtilis, is a toroid-shaped molecule that binds transcripts containing GAG or UAG repeats separated by two nucleotides.

The trp RNA-binding attenuation protein of Bacillus subtilis, TRAP, regulates both transcription and translation by binding to specific transcript sequences. The optimal transcript sequences required for TRAP binding were determined by measuring complex formation between purified TRAP protein and synthetic RNAs. RNAs were tested that contained repeats of different trinucleotide sequences, with differing spacing between the repeats. A transcript containing GAG repeats separated by two-nucleotide spacers was bound most tightly. In addition, transmission electron microscopy was used to examine the structure of TRAP and the TRAP-transcript complex. TRAP was observed to be a toroid-shaped oligomer when free or when bound to either a natural or a synthetic RNA.

Identification of the Bacillus subtilis pur operon repressor.

Transcription of the Bacillus subtilis pur operon is repressed in response to a signal of excess adenine. We have purified the repressor protein and have identified, cloned, and overexpressed the purR regulatory gene that controls transcription initiation of the operon. B. subtilis purR encodes a 62-kDa homodimer that binds to the pur operon control region. The PurR binding site which overlaps the promoter encompasses approximately 110 bp. The protein-DNA interaction is inhibited by 5-phosphoribosyl 1-pyrophosphate. A mutation that deletes the repressor binding site or one that disrupts purR abolishes binding activity in vitro and repression of transcription in vivo in response to the excess adenine signal. These results lead to a model in which an excess-adenine signal is transmitted to PurR via the 5-phosphoribosyl 1-pyrophosphate pool. In addition, purR is autoregulated. There is no structural or mechanistic similarity between the B. subtilis and Escherichia coli purine repressors.

Promoter architecture in the flagellar regulon of Bacillus subtilis: high-level expression of flagellin by the sigma D RNA polymerase requires an upstream promoter element.

Flagellin is one of the most abundant proteins in motile bacteria, yet its expression requires a low abundance sigma factor (sigma 28). We show that transcription from the Bacillus subtilis flagellin promoter is stimulated 20-fold by an upstream A+T-rich region [upstream promoter (UP) element] both in vivo and in vitro. This UP element is contacted by sigma 28 holoenzyme bound at the flagellin promoter and binds the isolated alpha 2 subassembly of RNA polymerase. The UP element increases the affinity of RNA polymerase for the flagellin promoter and stimulates transcription when initiation is limited by the rate of RNA polymerase binding. Comparison with other promoters in the flagellar regulon reveals a bipartite architecture: the -35 and -10 elements confer specificity for sigma 28, while promoter strength is determined largely by upstream DNA sequences.

Desempenho de bezerros leiteiros recebendo probiótico contendo Bacillus subtilis e Bacillus licheniformis

O objetivo deste estudo foi avaliar a suplementação de um probiótico composto por cepas de Bacillus subtilis e Bacillus licheniformis, fornecidos via sucedâneo lácteo, no que se refere ao escore e pH fecal, contagem de microrganismos intestinais, parâmetros sanguíneos e desempenho geral dos animais. Foram utilizados 24 animais da raça Holandês que receberam 4L/dia de sucedâneo comercial (15PB:15EE), além de livre acesso a água e concentrado inicial. O desaleitamento ocorreu na 8ª semana de vida. Os animais foram distribuídos em delineamento de blocos casualizados, em dois tratamentos: 1) Controle - sem a suplementação de probiótico; 2) Suplementação de 2g/d (1,6 x 109 UFC) de Bacillus subtilis e Bacillus licheniformis via sucedâneo lácteo. Semanalmente foram realizadas pesagens e aferições de medidas corporais (altura de cernelha, perímetro torácico e largura de garupa); e colheitas de sangue para determinação de glicose, proteína total, ureia e albumina, além de determinação de hematócrito. Foram colhidas amostras semanalmente para contagem de bactérias ácido láticas e enterobactérias e determinação de pH fecal. O monitoramento do consumo de concentrado e do escore fecal foi realizado diariamente. O peso corporal, o ganho de peso médio diário e as medidas corporais não foram alteradas (P>0,05) pela suplementação do probiótico contendo Bacillus subtilis e Bacillus licheniformis; muito embora tenham apresentado efeito significativo de idade dos animais (P<0,001). O escore fecal, pH fecal e consumo de concentrado diário também não foram afetados pela suplementação com probiótico. No entanto, o consumo de concentrado e o pH fecal sofreram influência da idade em resposta ao crescimento natural dos bezerros. A contagem de bactérias ácido láticas foi maior que número de enterobactérias durante todo o período (P<0,05). Apenas as enterobactérias sofreram efeito da idade (P<0,05), enquanto as bactérias ácido láticas permaneceram variando, porém dentro de um padrão constante. Os parâmetros sanguíneos também não foram afetados pela suplementação com probiótico (P>0,05) mas todos, com a exceção da albumina, tiveram influência da idade (P<0,001). A suplementação com o probiótico contendo Bacillus subtilis e Bacillus licheniformis via sucedâneo não apresentou benefícios no desempenho ou no metabolismo de bezerros leiteiros, bem como não reduziu a ocorrência de casos de diarreia.

Eximij doctoris magistri Nicholai de Orbellis Ordinis Minoru[m] sup[er] se[n]te[n]tias co[m]pe[n]diu[m] singulare elega[n]tiora doctoris subtilis dicta su[m]matim co[m]plecte[n]s

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Scotus academicus seu universa Doctoris Subtilis theologica dogmata /

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Theologia scotica in primum sententiarum magistri ... ad mentem subtilis doctoris Joannis Duns Scoti /

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Theologia scotica in secundum sententiarum magistri, ad mentem subtilis doctoris Joannis Duns Scoti /

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Theologia scotica in secundum sententiarum magistri, ad mentem subtilis doctoris Joannis Duns Scoti /

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Theologia scotica in tertium sententiarum magistri, ad mentem subtilis doctoris Joannis Duns Scoti /

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Theologia scotica in tertium sententiarum magistri, ad mentem subtilis doctoris Joannis Duns Scoti /

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Philosophia rationalis seu Logica magna Scotistica ad mentem venerabilis subtilis et Mariani doctoris Scoti /

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Logica magna ad mentem subtilis scoti seu Cursus philosophici Scotistici primae partis : tomus secundus /

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