In many cases protein assemblies are stabilized by covalent bonds, one example of which is the formation of intra- or intermolecular ε-(γ-glutamyl)lysil cross-links catalyzed by transglutaminases (TGases). Because of the potential for unwanted cross-linking reactions, the activities of many TGases have been shown to be tightly controlled. Bacterial endospores are highly resilient cells in part because they are surrounded by a complex protein coat.
View Article and Find Full Text PDFAt a late stage in spore development in , the mother cell directs synthesis of a layer of peptidoglycan known as the cortex between the two forespore membranes, as well as the assembly of a protective protein coat at the surface of the forespore outer membrane. SafA, the key determinant of inner coat assembly, is first recruited to the surface of the developing spore and then encases the spore under the control of the morphogenetic protein SpoVID. SafA has a LysM peptidoglycan-binding domain, SafA, and localizes to the cortex-coat interface in mature spores.
View Article and Find Full Text PDFDuring sporulation in Bacillus subtilis, a group of mother cell-specific proteins guides the assembly of the coat, a multiprotein structure that protects the spore and influences many of its environmental interactions. SafA and CotE behave as party hubs, governing assembly of the inner and outer coat layers. Targeting of coat proteins to the developing spore is followed by encasement.
View Article and Find Full Text PDFThe coat of spores is a multiprotein protective structure that also arbitrates many of the environmental interactions of the spore. The coat assembles around the cortex peptidoglycan layer and is differentiated into an inner and an outer layer and a crust. SafA governs assembly of the inner coat, whereas CotE drives outer coat assembly.
View Article and Find Full Text PDFGene expression during spore development in Bacillus subtilis is controlled by cell type-specific RNA polymerase sigma factors. σFand σE control early stages of development in the forespore and the mother cell, respectively. When, at an intermediate stage in development, the mother cell engulfs the forespore, σF is replaced by σG and σE is replaced by σK.
View Article and Find Full Text PDFDuring sporulation in Bacillus subtilis, the onset of activity of the late forespore-specific sigma factor σG coincides with completion of forespore engulfment by the mother cell. At this stage, the forespore becomes a free protoplast, surrounded by the mother cell cytoplasm and separated from it by two membranes that derive from the asymmetric division septum. Continued gene expression in the forespore, isolated from the surrounding medium, relies on the SpoIIIA-SpoIIQ secretion system assembled from proteins synthesised both in the mother cell and in the forespore.
View Article and Find Full Text PDFProteins SpoIIQ and SpoIIIAH interact through two membranes to connect the forespore and the mother cell during endospore development in the bacterium Bacillus subtilis. SpoIIIAH consists of a transmembrane segment and an extracellular domain with similarity to YscJ proteins. YscJ proteins form large multimeric rings that are the structural scaffolds for the assembly of type III secretion systems in gram-negative bacteria.
View Article and Find Full Text PDFTwo highly similar RNA polymerase sigma subunits, σ(F) and σ(G), govern the early and late phases of forespore-specific gene expression during spore differentiation in Bacillus subtilis. σ(F) drives synthesis of σ(G) but the latter only becomes active once engulfment of the forespore by the mother cell is completed, its levels rising quickly due to a positive feedback loop. The mechanisms that prevent premature or ectopic activation of σ(G) while discriminating between σ(F) and σ(G) in the forespore are not fully comprehended.
View Article and Find Full Text PDFA large number of proteins are known to reside at specific subcellular locations in bacterial cells. However, the molecular mechanisms by which many of these proteins are anchored at these locations remains unclear. During endospore formation in Bacillus subtilis, several integral membrane proteins are located specifically at the interface of the two adjacent cells of the developing sporangium, the mother cell and forespore.
View Article and Find Full Text PDFSporulation in Bacillus subtilis involves two cells that follow separate but coordinately regulated developmental programs. Late in sporulation, the developing spore (the forespore) resides within a mother cell. The regulation of the forespore transcription factor sigma(G) that acts at this stage has remained enigmatic.
View Article and Find Full Text PDFActivation of the late prespore-specific RNA polymerase sigma factor sigma(G) during Bacillus subtilis sporulation coincides with completion of the engulfment process, when the prespore becomes a protoplast fully surrounded by the mother cell cytoplasm and separated from it by a double membrane system. Activation of sigma(G) also requires expression of spoIIIJ, coding for a membrane protein translocase of the YidC/Oxa1p/Alb3 family, and of the mother cell-specific spoIIIA operon. Here we present genetic and biochemical evidence indicating that SpoIIIAE, the product of one of the spoIIIA cistrons, and SpoIIIJ interact in the membrane, thereby linking the function of the spoIIIJ and spoIIIA loci in the activation of sigma(G).
View Article and Find Full Text PDFProc Natl Acad Sci U S A
September 2008
At an early stage during Bacillus subtilis endospore development the bacterium divides asymmetrically to produce two daughter cells. The smaller cell (forespore) differentiates into the endospore, while the larger cell (mother cell) becomes a terminally differentiated cell that nurtures the developing forespore. During development the mother cell engulfs the forespore to produce a protoplast, surrounded by two bilayer membranes, which separate it from the cytoplasm of the mother cell.
View Article and Find Full Text PDFSpo0A, a classical two-component-type response regulator in Bacillus subtilis, binds to a specific DNA sequence found in many promoters to repress or activate the transcription of over 100 genes. On the spoIIG promoter, one of the Spo0A binding sites, centered at position -40, overlaps a consensus -35 element that may also interact with region 4 of the sigma A (sigma(A)) subunit of RNA polymerase. Molecular modeling corroborated by genetic evidence led us to propose that the binding of Spo0A to this site repositions sigma(A) region 4 on the promoter.
View Article and Find Full Text PDFEndospores formed by Bacillus, Clostridia, and related genera are encased in a protein shell called the coat. In many species, including B. subtilis, the coat is the outermost spore structure, and in other species, such as the pathogenic organisms B.
View Article and Find Full Text PDFThe Bacillus subtilis spoIIIA locus encodes eight proteins, SpoIIIAA to SpoIIIAH, which are expressed in the mother cell during endospore formation and which are essential for the activation of sigma(G) in the forespore. Complementation studies indicated that this locus may be transcribed from two promoters, one promoter upstream from the first gene and possibly a second unidentified promoter within the locus. Fragments of the spoIIIA locus were expressed at an ectopic site to complement the sporulation-defective phenotype of a spoIIIAH deletion, and we determined that complementation required a fragment of DNA that extended into spoIIIAF.
View Article and Find Full Text PDFThe synthesis of structural components and morphogenetic factors required for the assembly of the Bacillus subtilis spore coat is governed by a mother cell-specific transcriptional cascade. The first two temporal classes of gene expression, which involve RNA polymerase sigma sigma(E) factor and the ancillary regulators GerR and SpoIIID, are deployed prior to engulfment of the prespore by the mother cell. The two last classes rely on sigma(K), whose activation follows engulfment completion, and GerE.
View Article and Find Full Text PDFAlthough not thought to become competent for DNA uptake, the bacterium Streptococcus pyogenes appears to encode within its genome the DNA uptake and recombination machinery required for competence. The promoters of these genes contain a conserved sequence, CIN-box, which is recognized by the S. pyogenes alternative RNA polymerase sigma factor X.
View Article and Find Full Text PDFMorphogenetic proteins such as SpoVID and SafA govern assembly of the Bacillus subtilis endospore coat by guiding the various protein structural components to the surface of the developing spore. Previously, a screen for peptides able to interact with SpoVID led to the identification of a PYYH motif present in the C-terminal half of the SafA protein and to the subsequent demonstration that SpoVID and SafA directly interact. spoVID and safA spores show deficiencies in coat assembly and are lysozyme susceptible.
View Article and Find Full Text PDFAt the onset of endospore formation in Bacillus subtilis the DNA-binding protein Spo0A directly activates transcription from promoters of about 40 genes. One of these promoters, Pskf, controls expression of an operon encoding a killing factor that acts on sibling cells. AbrB-mediated repression of Pskf provides one level of security ensuring that this promoter is not activated prematurely.
View Article and Find Full Text PDFThe assembly of a multiprotein coat around the Bacillus subtilis spore confers resistance to lytic enzymes and noxious chemicals and ensures normal germination. Part of the coat is cross-linked and resistant to solubilization. The coat contains epsilon-(gamma-glutamyl)lysyl cross-links, and the expression of the gene (tgl) for a spore-associated transglutaminase was shown before to be required for the cross-linking of coat protein GerQ.
View Article and Find Full Text PDFWe report here studies of expression and functional analysis of a Bacillus subtilis gene, ywcE, which codes for a product with features of a holin. Primer extension analysis of ywcE transcription revealed that a single transcript accumulated from the onset of sporulation onwards, produced from a sigma(A)-type promoter bearing the TG dinucleotide motif of "extended" -10 promoters. No primer extension product was detected in vivo during growth.
View Article and Find Full Text PDFProtein localization is crucial for cellular morphogenesis and intracellular signal transduction cascades. Here we describe an interaction between two membrane proteins expressed in different cells of the Bacillus subtilis sporangium, the mother cell protein SpoIIIAH and the forespore protein SpoIIQ. We used affinity chromatography, coimmunoprecipitation, and the yeast two-hybrid system to demonstrate that the extracellular domains of these proteins interact, tethering SpoIIIAH to the sporulation septum, and directing its assembly with SpoIIQ into helical arcs and foci around the forespore.
View Article and Find Full Text PDFRNA polymerase sigma factor sigma(F) initiates the prespore-specific program of gene expression during Bacillus subtilis sporulation. sigma(F) governs transcription of spoIIIG, encoding the late prespore-specific regulator sigma(G). However, transcription of spoIIIG is delayed relative to other genes under the control of sigma(F), and after synthesis, sigma(G) is initially kept in an inactive form.
View Article and Find Full Text PDFSpores formed by wild-type Bacillus subtilis are encased in a multilayered protein structure (called the coat) formed by the ordered assembly of over 30 polypeptides. One polypeptide (CotB) is a surface-exposed coat component that has been used as a vehicle for the display of heterologous antigens at the spore surface. The cotB gene was initially identified by reverse genetics as encoding an abundant coat component.
View Article and Find Full Text PDFAt the onset of endospore formation in Bacillus subtilis, the DNA binding protein Spo0A activates transcription from two types of promoters. The first type includes the spoIIG and spoIIE promoters, which are used by sigma(A)-RNA polymerase, whereas the second type includes the spoIIA promoter, which is used by RNA polymerase containing the secondary sigma factor sigma(H). Previous genetic analyses have identified specific amino acids in alpha-helix E of Spo0A that are important for activation of Spo0A-dependent, sigma(A)-dependent promoters.
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