Bacillus subtilis degSU operon is regulated by the ClpXP-Spx regulated proteolysis system.

The DegS-DegU two-component regulatory system regulates many cellular events in Bacillus subtilis. Genes for DegSU constitutes an operon directed by the P1 promoter and downstream degU is autoregulated via the P3 promoter activated by phosphorylated DegU. In the Gram-positive bacteria, Spx plays a major role in the protection system against oxidative stresses as a transcriptional regulator.

The Bacillus subtilis response regulator gene degU is positively regulated by CcpA and by catabolite-repressed synthesis of ClpC.

In Bacillus subtilis, the response regulator DegU and its cognate kinase, DegS, constitute a two-component system that regulates many cellular processes, including exoprotease production and genetic competence. Phosphorylated DegU (DegU-P) activates its own promoter and is degraded by the ClpCP protease. We observed induction of degU by glucose in sporulation medium.

Effect of Bacillus subtilis BsuM restriction-modification on plasmid transfer by polyethylene glycol-induced protoplast fusion.

Polyethylene glycol (PEG)-induced cell fusion is a promising method to transfer larger DNA from one cell to another than conventional genetic DNA transfer systems. The laboratory strain Bacillus subtilis 168 contains a restriction (R) and modification (M) system, BsuM, which recognizes the sequence 5'-CTCGAG-3'. To study whether the BsuM system affects DNA transfer by the PEG-induced cell fusion between R(+)M(+) and R(-)M(-) strains, we examined transfer of plasmids pHV33 and pLS32neo carrying no and eight BsuM sites, respectively.

Identification of the sequences recognized by the Bacillus subtilis response regulator YclJ.

The Bacillus subtilis yclJ gene encodes an OmpR-type response regulator of a two-component regulatory system with unknown function. A previous DNA microarray experiment suggested that multicopy yclJ greatly enhances the expression of several operons in a cognate kinase (YclK)-deficient strain. To confirm this, lacZ fusion analysis was performed in the yclK background with overexpressed yclJ.

Functional analysis of the stability determinant AlfB of pBET131, a miniplasmid derivative of bacillus subtilis (natto) plasmid pLS32.

Bacillus subtilis plasmid pBET131 is a derivative of pLS32, which was isolated from a natto strain of Bacillus subtilis. The DNA region in pBET131 that confers segregational stability contains an operon consisting of three genes, of which alfA, encoding an actin-like ATPase, and alfB are essential for plasmid stability. In this work, the alfB gene product and its target DNA region were studied in detail.

TRPV2 expression in rat oral mucosa.

The oral mucosa is a highly specialised, stratified epithelium that confers protection from infection and physical, chemical and thermal stimuli. The non-keratinised junctional epithelium surrounds each tooth like a collar and is easily attacked by foreign substances from the oral sulcus. We found that TRPV2, a temperature-gated channel, is highly expressed in junctional epithelial cells, but not in oral sulcular epithelial cells or oral epithelial cells.

Regulation of Bacillus subtilis aprE expression by glnA through inhibition of scoC and sigma(D)-dependent degR expression.

Expression of the gene for the extracellular alkaline protease (aprE) of Bacillus subtilis is subject to regulation by many positive and negative regulators. We have found that aprE expression was increased by disruption of the glutamine synthetase gene glnA. The increase in aprE expression was attributed to a decreased in expression of scoC, which encodes a negative regulator of aprE expression.

The Bacillus subtilis late competence operon comE is transcriptionally regulated by yutB and under post-transcription initiation control by comN (yrzD).

The Bacillus subtilis genome has been sequenced, and disruptants with disruptions in genes that were not characterized previously were systematically generated. We screened these gene disruptants for decreased transformation frequency and identified two genes, yrzD and yutB, whose disruption resulted in severely reduced transformation frequency and modestly reduced transformation frequency, respectively. In the regulation of competence development, various signals affect the expression of comK, which encodes a master regulator of genetic competence that drives late competence gene transcription.

Distribution of substance P and neurokinin-1 receptors in the peri-implant epithelium around titanium dental implants in rats.

We examined the distribution of substance P and neurokinin-1 (NK1) receptors and substance-P-containing nerve fibers in the peri-implant mucosa around titanium dental implants in rats. Immunohistochemistry and immunocytochemistry revealed that substance-P-immunoreactive nerve fibers abundantly innervated the peri-implant epithelium (PIE) compared with other epithelia of the peri-implant mucosa. NK1 receptor mRNA and protein expression in the peri-implant mucosa were confirmed by reverse transcription with the polymerase chain reaction and immunoblotting.

Participation of intercellular adhesion molecule-2 (CD102) in B lymphopoiesis.

The survival and fate of blood cell precursors is dependent on their communication with stromal cells of various types within bone marrow. Monoclonal antibodies have proven to be powerful tools for identifying molecules responsible for such interactions and we now describe one that selectively blocks B lymphopoiesis. The BF/32 antibody inhibited the establishment, but not the maintenance of long-term bone marrow cultures capable of lymphocyte production.

Involvement of nitrogen regulation in Bacillus subtilis degU expression.

Bacillus subtilis DegS-DegU belongs to a bacterial two-component system that controls many processes, including the production of exocellular proteases and competence development. It was found that when the glutamine synthetase gene glnA, which is involved in nitrogen regulation, was disrupted, the expression of the response regulator degU gene was increased. Deletion analysis and 5'-end mapping of the degU transcripts showed that the increase was caused by induction of a promoter (P2) located before the degU gene.

Intracellular ClC-3 chloride channels promote bone resorption in vitro through organelle acidification in mouse osteoclasts.

ClC-7 Cl(-) channels expressed in osteoclasts are important for bone resorption since it has been shown that disruption of the ClCN7 gene in mice leads to severe osteopetrosis. We have previously reported that Cl(-) currents recorded from mouse osteoclasts resemble those of ClC-3 Cl(-) channels. The aim of the present study was to determine the expression of ClC-3 channels in mouse osteoclasts and their functional role during bone resorption.

Identification of the sequences recognized by the Bacillus subtilis response regulator YrkP.

The Bacillus subtilis yrkP gene encodes a response regulator of a two-component regulatory system of unknown function. A previous DNA microarray experiment suggested that multicopy yrkP greatly enhanced the expression of yrkN, the ykcBC operon, and yrkO, which encodes a putative transporter. Here, lacZ fusion analysis confirmed these results and also revealed that YrkP autoregulates the putative yrkPQR operon, indicating that yrkPQR and yrkO form a divergon structure.

Controlled release of simvastatin acid using cyclodextrin inclusion system.

Simvastatin acid (SVA) has been reported to stimulate bone formation by increasing expression of BMP-2 in osteoblasts. Due to their multi-functional characteristics and bioadaptability, cyclodextrins (CDs) are capable of forming inclusion complexes with many drugs by including a whole drug molecule inside their cavity. In the present study, we prepared SVA/CD inclusion complex solutions with different pH values.

Cystatin C stimulates the differentiation of mouse osteoblastic cells and bone formation.

Cystatin C (CysC) is a natural cysteine proteinase inhibitor that suppresses the differentiation and bone-resorptive function of osteoclasts. By contrast, the effect of CysC on the differentiation and bone-formative function of osteoblasts has not been elucidated thoroughly. We examined the effects of CysC on mouse osteoblastic cells using in vitro cultures from bone marrow and calvaria and ex vivo calvarial cultures.

The Bacillus subtilis NatK-NatR two-component system regulates expression of the natAB operon encoding an ABC transporter for sodium ion extrusion.

A previous microarray analysis suggested that multicopy yccH, encoding a function-unknown response regulator, enhances expression of natAB, which encodes a two-gene ATP-binding cassette transporter involved in the extrusion of sodium ions. The two-component regulatory system YccG-YccH was therefore renamed NatK-NatR. Here, this observation was confirmed by a lacZ fusion analysis using a strain carrying natA-lacZ.

Sequential expression of endothelial nitric oxide synthase, inducible nitric oxide synthase, and nitrotyrosine in odontoblasts and pulp cells during dentin repair after tooth preparation in rat molars.

Nitric oxide (NO) stimulates osteoblast differentiation, but whether NO contributes to odontoblast differentiation during dentin repair is unknown. By using reverse transcription/polymerase chain reaction and immunostaining, we investigated the gene expression and/or immunolocalization of endothelial NO synthase (eNOS), inducible NOS (iNOS), and nitrotyrosine (a biomarker for NO-derived peroxinitrite), and alkaline phosphatase (ALP) and osteocalcin (early and terminal differentiation markers of odontoblasts, respectively) in dental pulp tissue after rat tooth preparation. At the early stage (1-3 days) post-preparation, markedly increased expression of iNOS and nitrotyrosine was found in odontoblasts and pulp cells beneath the cavity, whereas eNOS expression was significantly decreased.

Oxygen plasma surface modification enhances immobilization of simvastatin acid.

Simvastatin acid (SVA) has been reported to stimulate bone formation with increased expression of BMP-2. Therefore, immobilization of SVA onto dental implants is expected to promote osteogenesis at the bone tissue/implant interface. The aim of this study was to evaluate the immobilization behavior of SVA onto titanium (Ti), O(2)-plasma treated titanium (Ti + O(2)), thin-film coatings of hexamethyldisiloxane (HMDSO), and O(2)-plasma treated HMDSO (HMDSO + O(2)) by using the quartz crystal microbalance-dissipation (QCM-D) technique.

Capsaicin receptor expression in the rat temporomandibular joint.

Experimentally, temporomandibular joint (TMJ) nerve units respond to capsaicin, which is used clinically to treat TMJ pain. However, the existence of capsaicin receptors in the TMJ has not previously been clearly demonstrated. Immunohistochemical analysis has revealed the presence of transient receptor potential vanilloid subtype 1 (TRPV1) expression in the nerves and synovial lining cells of the TMJ.

Immunocytochemical localization of the neurokinin 1 receptor in rat dental pulp.

The dentin-pulp complex is a peripheral end-organ supplied by dense sensory nerve fibers. Substance P, a representative neuropeptide widely distributed in the dental pulp, has been reported to play roles in pain transmission and the amplification of inflammation. We analyzed here the expression of the neurokinin 1 (NK1) receptor, preferentially activated by substance P, using immunocytochemistry in rat dental pulp at both the light and electron microscopic levels.

Inhibition of Bacillus subtilis scoC expression by multicopy senS.

The Bacillus subtilis aprE gene, which encodes the extracellular alkaline protease, is regulated by many positive and negative transcriptional regulators. SenS is one such positive regulator consisting of 65 amino acids. We found that the senS gene on a multicopy plasmid, pSEN24, caused an increase in aprE expression in strains carrying the upstream region of aprE up to -340 with respect to the transcription initiation site but not in a strain carrying the region up to -299, which is within the binding site of the negative regulator ScoC (Hpr).

Characterization of the replication region of plasmid pLS32 from the Natto strain of Bacillus subtilis.

Plasmid pL32 from the Natto strain of Bacillus subtilis belongs to a group of low-copy-number plasmids in gram-positive bacteria that replicate via a theta mechanism of replication. We studied the DNA region encoding the replication protein, RepN, of pLS32, and obtained the following results. Transcription of the repN gene starts 167 nucleotides upstream from the translational start site of repN.

Cytoskeleton and surface structures of cells directly attached to the tooth in the rat junctional epithelium.

Objective: It is still an open question whether cells directly attached to the tooth (DAT) cells are migratory or non-migratory cells. The purpose of this study was to examine cytoskeletal and surface structures of DAT cells that might be involved in migration.

Methods: We investigated the distribution of stress fibers composed of actin filaments in DAT cells using phallacidin fluorescent dye methods in a confocal laser scanning microscope.

Ultrastructural localization of laminin-5 (gamma2 chain) in the rat peri-implant oral mucosa around a titanium-dental implant by immuno-electron microscopy.

Laminin-5 (Ln-5) is an important molecule associated with epithelial cell adhesion and migration. In the gingiva around the tooth, Ln-5 localizes within basement membranes between the junctional epithelium (JE) and the tooth or connective tissue. Recently, we reported that in the oral mucosa around a dental implant, Ln-5 is expressed within the basement membranes at the implant-peri-implant epithelium (PIE) interface, and at the PIE-connective tissue interface.

Genes involved in SkfA killing factor production protect a Bacillus subtilis lipase against proteolysis.

Small lipases of Bacillus species, such as LipA from Bacillus subtilis, have a high potential for industrial applications. Recent studies showed that deletion of six AT-rich islands from the B. subtilis genome results in reduced amounts of extracellular LipA.