Re-structuring the host cell: up close with Salmonella's molecular machinery

A common theme in bacterial disease is the manipulation of the eukaryotic cytoskeleton by pathogenic factors. Understanding how virulence factors operate to achieve these alterations is a growing and important research effort. This review focuses on several virulence factors from the pathogen Salmonella, and considers the contributions of structural biology to our appreciation of bacterial modulation of cytoskeletal dynamics.     

Highly reactive cysteine residues are part of the substrate binding site of mammalian dipeptidyl peptidases III

Dipeptidyl peptidase III (DPP III) is a cytosolic zinc-exopeptidase involved in the intracellular protein catabolism of eukaryotes. Although inhibition by thiol reagents is a general feature of DPP III originating from various species, the function of activity important sulfhydryl groups is still inadequately understood. The present study of the reactivity of these groups was undertaken in order to clarify their biological significance.The inactivation kinetics of human and rat DPP III by sulfhydryl reagent p-hydroxy-mercuribenzoate (pHMB) was monitored by determination of the enzyme's residual activity with fluorimetric detection.Inactivation of this human enzyme exhibited pseudo-first-order kinetics, suggesting that all reactive SH-groups have equivalent reactivity, and the second-order rate constant was calculated to be 3523+/-567M(-1)min(-1). Rat DPP III was hyperreactive to pHMB and showed biphasic kinetics indicating two classes of reactive SH-groups. The second-order rate constants of 3540M(-1)s(-1) for slower reacting sulfhydryl, and 21,855M(-1)s(-1) for faster reacting sulfhydryl were obtained from slopes of linear plots of pseudo-first-order constants versus reagent concentration. Peptide substrates protected both mammalian DPPs III from inactivation by pHMB. Physiological concentrations of biological thiols and H(2)O(2) inactivated the rat DPP III. Human enzyme was resistant to H(2)O(2) attack and less affected by reduced glutathione (GSH) than the rat homologue. A significantly lower DPP III level, determined by activity measurement and Western blotting, was found in the cytosols of highly oxygenated rat tissues.These results provide kinetic evidence that cysteine residues are involved in substrate binding of mammalian DPPs III.     

Compartmentalization of the splicing machinery in plant cell nuclei

The cell nucleus is a membrane-surrounded organelle that contains numerous compartments in addition to chromatin. Compartmentalization of the nucleus is now accepted as an important feature for the organization of nuclear processes and for gene expression. Recent studies on nuclear organization of splicing factors in plant cells provide insights into the compartmentalization of the plant cell nuclei and conservation of nuclear compartments between plants and metazoans.     

Possible apoptotic mechanism in epidermal cell acantholysis induced by pemphigus vulgaris autoimmunoglobulins

Through a still unclear mechanism, pemphigus vulgaris autoantibodies (PV-IgG) induce intra-epidermal acantholytic lesions responsible for severe to fatal skin wounding. We present evidence that PV lesions contain apoptotic keratinocytes, and that cell death is induced in the lesional tissue apparently before cell separation. These data suggest that apoptosis could be the cause of the acantholytic phenomenon. We show that PV-IgG and an antibody against Fas receptor (anti-FasR) induce lesions in vitro in a similar way, causing: (1) secretion of soluble FasL; (2) elevated cellular amounts of FasR, FasL (soluble and membranal), Bax and p53 proteins; (3) reduction in levels of cellular Bcl-2; (4) enrichment in caspase 8, and activation of caspases 1 and 3; (5) co-aggregation of FasL and FasR with caspase 8 in membranal death-inducing signaling complex (DISC). Hence, the Fas-mediated death signaling pathway seems to be involved in lesion formation. Moreover, we have shown that in skin organ cultures and in keratinocyte cultures, PV-IgG can induce caspase activation and DNA fragmentation, and caspase inhibitors can prevent the formation of PV-IgG-induced epidermal lesions. Altogether, these results suggest that PV-IgG-induced acantholysis may proceed through the death-signaling pathway. They highlight new perspectives on mechanisms of tissue damage in autoimmune diseases.     

Structure of the integrin binding fragment from fibrillin-1 gives new insights into microfibril organization

Human fibrillin-1, the major structural protein of extracellular matrix (ECM) 10-12 nm microfibrils, is dominated by 43 calcium binding epidermal growth factor-like (cbEGF) and 7 transforming growth factor beta binding protein-like (TB) domains. Crystal structures reveal the integrin binding cbEGF22-TB4-cbEGF23 fragment of human fibrillin-1 to be a Ca(2+)-rigidified tetragonal pyramid. We suggest that other cbEGF-TB pairs within the fibrillins may adopt a similar orientation to cbEGF22-TB4. In addition, we have located a flexible RGD integrin binding loop within TB4. Modeling, cell attachment and spreading assays, immunocytochemistry, and surface plasmon resonance indicate that cbEGF22 bound to TB4 is a requirement for integrin activation and provide insight into the molecular basis of the fibrillin-1 interaction with alphaVbeta3. In light of our data, we propose a novel model for the assembly of the fibrillin microfibril and a mechanism to explain its extensibility.     

Testing the phylogenetic position of a parasitic plant (Cuscuta, Convolvulaceae, asteridae): Bayesian inference and the parametric bootstrap on data drawn from three genomes

Previous findings on structural rearrangements in the chloroplast genome of Cuscuta (dodder), the only parasitic genus in the morning-glory family, Convolvulaceae, were attributed to its parasitic life style, but without proper comparison to related nonparasitic members of the family. Before molecular evolutionary questions regarding genome evolution can be answered, the phylogenetic problems within the family need to be resolved. However, the phylogenetic position of parasitic angiosperms and their precise relationship to nonparasitic relatives are difficult to infer. Problems are encountered with both morphological and molecular evidence. Molecular data have been used in numerous studies to elucidate relationships of parasitic taxa, despite accelerated rates of sequence evolution. To address the question of the position of the genus Cuscuta within Convolvulaceae, we generated a new molecular data set consisting of mitochondrial (atpA) and nuclear (RPB2) genes, and analyzed these data together with an existing chloroplast data matrix (rbcL, atpB, trnL-F, and psbE-J), to which an additional chloroplast gene (rpl2) was added. This data set was analyzed with an array of phylogenetic methods, including Bayesian analysis, maximum likelihood, and maximum parsimony. Further exploration of data was done by using methods of phylogeny hypothesis testing. At least two nonparasitic lineages are shown to diverge within the Convolvulaceae before Cuscuta. However, the exact sister group of Cuscuta could not be ascertained, even though many alternatives were rejected with confidence. Caution is therefore warranted when interpreting the causes of molecular evolution in Cuscuta. Detailed comparisons with nonparasitic Convolvulaceae are necessary before firm conclusions can be reached regarding the effects of the parasitic mode of life on patterns of molecular evolution in Cuscuta.     

Influence of the soil on certain dermatophytes and their evolutional trend

Conclusions From our experiments it could be seen that a quite definite relationship exists between various dermatophytes and the soil which is expressed on the one side by antagonistic action of the microflora in the soil inhibiting the growth of dermatophytes and on the other, by ability of some dermatophytes to persist in the soil or to use the soil as habitat in their saprophytic life. This condition appears to be the reflection of the evolutive tendency of dermatophytes to develop from its primary habitat, the soil, to the parasitic life on man and animal. It is persumed that this evolution of dermatophytes is based on mutations of gens as one of the fundamental phenomenons in nature. It is obvious that this evolution of dermatophytes is not completed. Our experiments regardingT. mentagrophytes indicate that this species is at present still to a certain extent in a transitional evolutive stage with retained but limited ability to the life in the soil and with an evident tendency for adaption on small rodents and human beings. In this course of development on a lower level isK. ajelloi andM. gypseum with full ability of saprophytic life in the soil and with slowly but increasing pathogenicity for men and animals. E. I. Grin, Prof. of Dermatology, Med. Faculty, Univ. Sarajevo, Director Institute of Dermatovenerology. L. Oegovi, Prof. of Intern. Dis., Veter. Faculty, Univ. Sarajevo, Chief of Dept. of Mycology Inst. of Dermatovenerology.     

Identification of biomotor structures as a precondition for programming kinesiologic education in children aged seven to nine years

A sample consisting of 487 children (249 male and 236 female) aged 7-9 years (+/- 2 months) underwent programmed kinesiologic transformation procedures for 18 months. The morphological and motor development was followed up by use of 14 morphological and 12 motor variables at 9-month intervals. Three taxonomic analyses for each measurement time point were calculated for either sex in order to determine the initial and transitive position for each individual study subject, and to identify most homogeneous groups within the sample as a whole. Three taxonomic variables were isolated on each measurement for either sex. Study results revealed female children to undergo faster development with earlier formation of the three morphological-motor structures ranked according to their predominance: mass, i.e. ectomesomorphy, motor, and endomorphy. Entity projections upon taxonomic variables at particular measurement points clearly identified the morphological-motor variables to be addressed by general and differentiated programs of kinesiologic education in order to achieve optimal effects during the development of the child's body as a whole.     

Analysis of indicators of load during the game in activity of the second line attacker in water polo

Water polo, as an activity, belongs in the category of polystructural complex move sports. The activities of a player's in role of second line attacker is observed on the sample of competitive games in the First national league. The study is aimed to define a set of new measurement variables for the objective recording of amount, intensity and duration of player's activities, and its evaluation by means of factor validity criteria. On the sample of 87 players, 29 variables were applied. Competent, trained officials made measurements. Basic statistics of all measured variables is presented as referent values of various player activities. In the factor analysis, three factors are found to be significant, explaining 84.6% of source variability, which is a subset of multivariate normally, distributed variables. Factors are interpreted as: "quantity of actions", "intensity of activity in the vertical body posture", and "intensity and extensity of activity in horizontal body posture". Of the last two, body posture is found to be specific in water polo, due to specifics of the game that is played in water. It is concluded that the proposed variables and measurement procedures are very well suited and objectively instrument for the purpose of measurement of the energetic aspect of kinesiological activity analysis.