Functional conservation of the lipid II biosynthesis pathway in the cell wall‐less bacteria Chlamydia and Wolbachia: why is lipid II needed?

Cell division and cell wall biosynthesis in prokaryotes are driven by partially overlapping multiprotein machineries whose activities are tightly controlled and co-ordinated. So far, a number of protein components have been identified and acknowledged as essential for both fundamental cellular processes. Genes for enzymes of both machineries have been found in the genomes of the cell wall-less genera Chlamydia and Wolbachia , raising questions as to the functionality of the lipid II biosynthesis pathway and reasons for its conservation. We provide evidence on three levels that the lipid II biosynthesis pathway is indeed functional and essential in both genera: (i) fosfomycin, an inhibitor of MurA, catalysing the initial reaction in lipid II biosynthesis, has a detrimental effect on growth of Wolbachia cells; (ii) isolated cytoplasmic membranes from Wolbachia synthesize lipid II ex vivo ; and (iii) recombinant MraY and MurG from Chlamydia and Wolbachia exhibit in vitro activity, synthesizing lipid I and lipid II respectively. We discuss the hypothesis that the necessity for maintaining lipid II biosynthesis in cell wall-lacking bacteria reflects an essential role of the precursor in prokaryotic cell division. Our results also indicate that the lipid II pathway may be exploited as an antibacterial target for chlamydial and filarial infections.     

A Role for the Karyopherin Kap123p in Microtubule Stability

Several components of the nuclear transport machinery play a role in mitotic spindle assembly in higher eukaryotes. To further investigate the role of this family of proteins in microtubule function, we screened for mutations in Saccharomyces cerevisiae that confer sensitivity to microtubule-destabilizing drugs. One mutant exhibiting this phenotype lacked the gene encoding the karyopherin Kap123p. Analysis of kap123 Δ cells revealed that the drug sensitivity was caused by a defect in microtubule stability and/or assembly. In support of this idea, we demonstrated genetic interactions between the kap123 Δ mutation and mutated alleles of genes encoding α-tubulins and factors controlling microtubule dynamics. Moreover, kap123 Δ cells exhibit defects in spindle structure and dynamics as well as nuclear positioning defects during mitosis. Cultures of kap123 Δ strains are enriched for mononucleated large-budded cells often containing short spindles and nuclei positioned away from the budneck, phenotypes indicative of defects in both cytoplasmic and nuclear microtubules. Finally, we identified a gene, CAJ1 , which when deleted in combination with KAP123 exacerbated the microtubule-related defects of the kap123 Δ mutants. We propose that Kap123p and Caj1p, a member of the Hsp40 family of proteins, together play an essential role in normal microtubule function.     

Benefits of a root fungal endophyte on physiological processes and growth of the vulnerable legume tree Prosopis chilensis (Fabaceae)

Aims Desertification is a major concern in arid and semi-arid regions globally. Understanding interactions between vulnerable plant species and associated microbial symbionts may have important applications for conservation and restoration strategies in affected areas.     

Protein kinase C increase in rat brain cortical membranes may be promoted by cognition enhancing drugs.

Protein Kinase C (PKC) activity was measured in soluble and particulate fractions of rat individual brain cortices after in vivo treatment with two cognition enhancers: oxiracetam and alpha-glicerylphosphorylcholine. Both drugs induced an increase (+40-50%) of PKC particulate activity at 1 hr after the treatment. The effect was transient; at 5 hours PKC activity was lower than in controls. The dose response curve to oxiracetam was bell shaped, the increase of PKC being significant at 100 mg/kg. At higher doses the drug induced a decrease in enzyme activity. The increased PKC activity may be related to the cortical effects of these compounds.     

Maturation and secretion of a serine proteinase is associated with events of late microsporogenesis

An antiserum against meiotic proteins which bind to DNA cellulose was generated as a tool to assist the identification and purification of microsporogenesis-specific proteins. In immunoblotting experiments, this antiserum identified three meiotic proteins which are differentially expressed in anthers during microsporogenesis. One of these proteins was purified and characterized by biochemical and immunological techniques. This 82 kDa protein is synthesized as a preproprotein, acquires glycans as it moves through the endoplasmic reticulum and Golgi body, and is secreted into the anther locule. Immunocytochemical experiments demonstrate that the protein is expressed primarily in tapetal cells, and reaches peak concentrations as the microsporocytes reach the tetrad stage. Zymogram analyses and protein sequence comparisons indicate that the protein is a member of the serine proteinase family. The possible roles of the proteinase in microsporogenesis and pollen development are discussed.     

In vitro differentiation of human retinoblastoma cells into neuronal phenotypes

In order to characterize the cell type(s) of origin of human retinoblastoma cells by immunophenotyping, primary cells from seven retinoblastomas and of the corresponding cell lines (RBL lines), as well as four retinoblastoma (RB) lines established by other groups, were compared with rat and human retina cells, and with the adenovirus E1A-transformed human retinoblast cell line HER-Xho1-CC2. Analyses using monoclonal antibodies (Mabs) RB13-2 and RB21-7, originally raised against prenatal rat brain cells and recognizing neural cell surface antigens expressed in a developmental-stage-dependent manner, and three cell-type-specific Mabs (Q211, M501, Mab directed against vimentin) developed by other groups, gave the following results: (i) Retinoblastomas consist of cells expressing differentiated neuronal phenotypes during cultivation in vitro; (ii) All of the newly established RBL lines express neuronal phenotypes; and (iii) Cell lines such as Y79, which have been propagated in vitro for extended periods, do not express antigens specific for the neuronal pathway and cannot, therefore, be considered phenotypically representative of retinoblastoma cells.     

Pulsed-field electrophoresis in contour-clamped homogenous electric fields (CHEF) for fingerprinting of Rhizobium spp

Abstract: A pulsed-field gel electrophoretic method based on contour-clamped homogeneous electric field (CHEF) was developed for the analysis of natural isolates of Rhizobium leguminosarum biovar viciae . The procedure involves the use of 'rare-cutting' endonucleases. The separation of genomic DNA fragments with split runs ( τ = 5 s for 15 h followed by τ = 12 s for 9 h) allows a clear definition of profiles with bands ranging from 20–300-kb pairs. Two and a half days are sufficient to reproducibly accomplish the procedure from cell lysis to gel picture. The method can be used for different fast-growing rhizobia.