Constitutive septal murein synthesis in Escherichia coli with impaired activity of the morphogenetic proteins RodA and penicillin-binding protein 2.

The pattern of peptidoglycan (murein) segregation in cells of Escherichia coli with impaired activity of the morphogenetic proteins penicillin-binding protein 2 and RodA has been investigated by the D-cysteine-biotin immunolabeling technique (M. A. de Pedro, J. C. Quintela, J.-V. H?ltje, and H. Schwarz, J. Bacteriol. 179:2823-2834, 1997). Inactivation of these proteins either by amdinocillin treatment or by mutations in the corresponding genes, pbpA and rodA, respectively, leads to the generation of round, osmotically stable cells. In normal rod-shaped cells, new murein precursors are incorporated all over the lateral wall in a diffuse manner, being mixed up homogeneously with preexisting material, except during septation, when strictly localized murein synthesis occurs. In contrast, in rounded cells, incorporation of new precursors is apparently a zonal process, localized at positions at which division had previously taken place. Consequently, there is no mixing of new and old murein. Old murein is preserved for long periods of time in large, well-defined areas. We propose that the observed patterns are the result of a failure to switch off septal murein synthesis at the end of septation events. Furthermore, the segregation results confirm that round cells of rodA mutants do divide in alternate, perpendicular planes as previously proposed (K. J. Begg and W. D. Donachie, J. Bacteriol. 180:2564-2567, 1998).     

Differentiation between human and ovine isolates of Bordetellaparapertussis using pulsed-field gel electrophoresis

Abstract The genetic relatedness of 18 human and 29 ovine isolates of Bordetella parapertussis was examined by macrorestriction digestion of DNA with the rarely cutting enzyme Xba I and resolution by pulsed-field gel electrophoresis. There was clear separation of human and ovine isolates and variation within host types. The human isolates were separated into three types as were the 24 Scottish ovine isolates. Species-specific bands were observed with the human isolates at 114, 134, 166, 213, 346 and 372 kb. No species-specific bands were found in the B. parapertussis ovine isolates. Isolates of B. parapertussis recovered from sheep in New Zealand gave a further two DNA banding patterns which were clearly different from the Scottish ovine and the human isolates. These results indicate that human and ovine isolates of B. parapertussis are genetically distinct and that variation exists within isolates from the same host species. Pulsed-field gel electrbphoresis therefore appears to be a powerful discriminatory tool for the classification of B. parapertussis .     

Roles of FtsA and FtsZ in Activation of Division Sites

Increasing FtsZ induces the formation of minicells at cell poles but does not increase the frequency or timing of central divisions. A coordinate increase in both FtsZ and FtsA, however, increases the frequency of both polar and central divisions.     

Interaction of Sirt3 with OGG1 contributes to repair of mitochondrial DNA and protects from apoptotic cell death under oxidative stress

Sirtuin 3 (Sirt3), a major mitochondrial NAD⁺-dependent deacetylase, targets various mitochondrial proteins for lysine deacetylation and regulates important cellular functions such as energy metabolism, aging, and stress response. In this study, we identified the human 8-oxoguanine-DNA glycosylase 1 (OGG1), a DNA repair enzyme that excises 7,8-dihydro-8-oxoguanine (8-oxoG) from damaged genome, as a new target protein for Sirt3. We found that Sirt3 physically associated with OGG1 and deacetylated this DNA glycosylase and that deacetylation by Sirt3 prevented the degradation of the OGG1 protein and controlled its incision activity. We further showed that regulation of the acetylation and turnover of OGG1 by Sirt3 played a critical role in repairing mitochondrial DNA (mtDNA) damage, protecting mitochondrial integrity, and preventing apoptotic cell death under oxidative stress. We observed that following ionizing radiation, human tumor cells with silencing of Sirt3 expression exhibited deteriorated oxidative damage of mtDNA, as measured by the accumulation of 8-oxoG and 4977 common deletion, and showed more severe mitochondrial dysfunction and underwent greater apoptosis in comparison with the cells without silencing of Sirt3 expression. The results reported here not only reveal a new function and mechanism for Sirt3 in defending the mitochondrial genome against oxidative damage and protecting from the genotoxic stress-induced apoptotic cell death but also provide evidence supporting a new mtDNA repair pathway.     

Intra-articular injection of micronized dehydrated human amnion/chorion membrane attenuates osteoarthritis development

Introduction

Micronized dehydrated human amnion/chorion membrane (μ-dHACM) is derived from donated human placentae and has anti-inflammatory, low immunogenic and anti-fibrotic properties. The objective of this study was to quantitatively assess the efficacy of μ-dHACM as a disease modifying intervention in a rat model of osteoarthritis (OA). It was hypothesized that intra-articular injection of μ-dHACM would attenuate OA progression.

Methods

Lewis rats underwent medial meniscal transection (MMT) surgery to induce OA. Twenty four hours post-surgery, μ-dHACM or saline was injected intra-articularly into the rat joint. Naïve rats also received μ-dHACM injections. Microstructural changes in the tibial articular cartilage were assessed using equilibrium partitioning of an ionic contrast agent (EPIC-μCT) at 21 days post-surgery. The joint was also evaluated histologically and synovial fluid was analyzed for inflammatory markers at 3 and 21 days post-surgery.

Results

There was no measured baseline effect of μ-dHACM on cartilage in naïve animals. Histological staining of treated joints showed presence of μ-dHACM in the synovium along with local hypercellularity at 3 and 21 days post-surgery. In MMT animals, development of cartilage lesions at 21 days was prevented and number of partial erosions was significantly reduced by treatment with μ-dHACM. EPIC-μCT analysis quantitatively showed that μ-dHACM reduced proteoglycan loss in MMT animals.

Conclusions

μ-dHACM is rapidly sequestered in the synovial membrane following intra-articular injection and attenuates cartilage degradation in a rat OA model. These data suggest that intra-articular delivery of μ-dHACM may have a therapeutic effect on OA development.     

Influence of lipids on the hydrophobic barrier within the pore of the TWIK-1 K2P channel

Several recent ion channel structures have revealed large side portals, or ‘fenestrations’ at the interface between their transmembrane helices that potentially expose the ion conduction pathway to the lipid core of the bilayer. In a recent study we demonstrated that functional activity of the TWIK-1 K2P channel is influenced by the presence of hydrophobic residues deep within the inner pore. These residues are located near the fenestrations in the TWIK-1 structure and promote dewetting of the pore by forming a hydrophobic barrier to ion conduction. During our previous MD simulations, lipid tails were observed to enter these fenestrations. In this addendum to that study, we investigate lipid contribution to the dewetting process. Our results demonstrate that lipid tails from both the upper and lower leaflets can occupy the fenestrations and partially penetrate into the pore. The lipid tails do not sterically occlude the pore, but there is an inverse correlation between the presence of water within the hydrophobic barrier and the number of lipids tails within the lining of the pore. However, dewetting still occurs in the absence of lipids tails, and pore hydration appears to be determined primarily by those side-chains lining the narrowest part of the pore cavity.