An adhesome comprising laminin, dystroglycan and myosin IIA is required during notochord development in Xenopus laevis.

Dystroglycan (Dg) is a transmembrane receptor for laminin that must be expressed at the right time and place in order to be involved in notochord morphogenesis. The function of Dg was examined in Xenopus laevis embryos by knockdown of Dg and overexpression and replacement of the endogenous Dg with a mutated form of the protein. This analysis revealed that Dg is required for correct laminin assembly, for cell polarization during mediolateral intercalation and for proper differentiation of vacuoles. Using mutations in the cytoplasmic domain, we identified two sites that are involved in cell polarization and are required for mediolateral cell intercalation, and a site that is required for vacuolation. Furthermore, using a proteomic analysis, the cytoskeletal non-muscle myosin IIA has been identified for the first time as a molecular link between the Dg-cytoplasmic domain and cortical actin. The data allowed us to identify the adhesome laminin-Dg-myosin IIA as being required to maintain the cortical actin cytoskeleton network during vacuolation, which is crucial to maintain the shape of notochordal cells.

The dystroglycan: nestled in an adhesome during embryonic development.

Invertebrate and vertebrate development relies on complex processes that require many coordinated cell functions including cell adhesion, migration, proliferation and polarization. These processes depend on tissues and are spatio-temporally regulated by specific interactions between cells and between cells and the extracellular matrices. The dystroglycan, a transmembrane receptor that binds multiple extracellular matrix proteins, is expressed from oogenesis to organogenesis. There are increasing data suggesting that the axis, consisting of extracellular component-dystroglycan-cytoplasmic proteins, controls both the adhesion of cells to matrices as well as the transduction of signals coming from or directed to matrices. In this article, we review current advances leading to consider that the dystroglycan is a key protein nestled in an adhesome involved in mechanisms of cell adhesion during embryonic development.

A function for dystroglycan in pronephros development in Xenopus laevis.

Dystroglycan (Dg) is a laminin receptor that is expressed at the interface between the basement membrane and the cell membrane. Dg has been reported to play a role in skeletal muscle cell stability, morphogenesis of neuroepithelial tissues, and in regulating cytoskeletal organization, cell polarization, and cell signalling. In this study, we have focused our analysis on the expression of Dg-mRNA and protein at different developmental stages in the pronephros of Xenopus laevis. In order to study its role, we performed loss-of-function experiments mediated by Dg antisense morpholinos and dominant negative mutant. We show that Dg expression is first detectable when epithelialization begins in the pronephric anlage and persists later during tubulogenesis. Loss-of-function experiments induced a disorganization of the basement membrane, a drastic reduction of pronephric tubules and duct that can lead to a renal agenesis. A diminished proliferation of pronephric cell progenitors was also observed in Dg depleted embryos. Together, these data indicate that Dg plays a key role for laminin-1 assembly and pronephric cell anchoring to the basement membrane during early development of the pronephros. They also indicate that Dg may induce a signal transduction pathway controlling cell proliferation needed for the formation of tubules and their growth.

Synthesis and antibiotic activity of a small molecules library of 1,2,3-triazole derivatives.

A small library of simple 1,4-disubstituted 1,2,3-triazoles was prepared using a known one-pot procedure starting from organic halides and terminal alkynes. The compounds were then tested for their antibacterial activity against normal and resistant species of Staphylococcus aureus.

Anti-bacterial activity of some Brazilian medicinal plants.

Extracts from various organs of 25 plants of Brazilian traditional medicine were assayed with respect to their anti-bacterial activities against Escherichia coli, a susceptible strain of Staphylococcus aureus and two resistant strains of Staphylococcus aureus harbouring the efflux pumps NorA and MsrA. Amongst the 49 extracts studied, 14 presented anti-bacterial activity against Staphylococcus aureus, including the ethanolic extracts from the rhizome of Jatropha elliptica, from the stem barks of Schinus terebinthifolius and Erythrina mulungu, from the stems and leaves of Caesalpinia pyramidalis and Serjania lethalis, and from the stem bark and leaves of Lafoensia pacari. The classes of compounds present in the active extracts were determined as a preliminary step towards their bioactivity-guided separation. No extracts were active against Escherichia coli.

Multidrug resistance reversal agent from Jatropha elliptica.

As part of an ongoing project to identify plant natural products as resistance-modifying agents, bioassay-guided fractionation of an extract of Jatropha elliptica (Pohl) Muell Arg. led to the isolation of a penta-substituted pyridine, namely 2,6-dimethyl-4-phenyl-pyridine-3,5-dicarboxylic acid diethyl ester (8). The structure was established by spectroscopic methods. This known compound was assayed for in vitro antibacterial and resistance-modifying activities against strains of Staphylococcus aureus possessing the MsrA and NorA resistance efflux mechanisms. Antibiotic efflux studies indicated that (8) acts as an inhibitor of the NorA efflux pump and restores the level of intracellular drug concentration.

Dystroglycan is involved in skin morphogenesis downstream of the Notch signaling pathway.

Dystroglycan (Dg) is a transmembrane protein involved both in the assembly and maintenance of basement membrane structures essential for tissue morphogenesis, and the transmission of signals across the plasma membrane. We used a morpholino knockdown approach to investigate the function of Dg during Xenopus laevis skin morphogenesis. The loss of Dg disrupts epidermal differentiation by affecting the intercalation of multiciliated cells, deposition of laminin, and organization of fibronectin in the extracellular matrix (ECM). Depletion of Dg also affects cell-cell adhesion, as shown by the reduction of E-cadherin expression at the intercellular contacts, without affecting the distribution of ß(1) integrins. This was associated with a decrease of cell proliferation, a disruption of multiciliated-cell intercalation, and the down-regulation of the transcription factor P63, a marker of differentiated epidermis. In addition, we demonstrated that inhibition or activation of the Notch pathway prevents and promotes transcription of X-dg. Our study showed for the first time in vivo that Dg, in addition to organizing laminin in the ECM, also acts as a key signaling component in the Notch pathway.

In vivo analyzes of dystroglycan function during somitogenesis in Xenopus laevis.

Dystroglycan (Dg) is a cell adhesion receptor for laminin that has been reported to play a role in skeletal muscle cell stability, cytoskeletal organization, cell polarity, and signaling. Here we show that Dg is expressed at both the notochord/somite and the intersomitic boundaries, where laminin and fibronectin are accumulated during somitogenesis. Inhibition of Dg function with morpholino antisense oligonucleotides or a dominant negative mutant results in the normal segmentation of the presomitic mesoderm but affects the number, the size, and the integrity of somites. Depletion of Dg disrupts proliferation and alignment of myoblasts without affecting XMyoD and XMRF4 expression. It also leads to defects in laminin deposition at the intersomitic junctions, whereas expression of integrin beta1 subunits and fibronectin assembly occur normally. Our results show that Dg is critical for both proliferation and elongation of somitic cells and that the Dg-cytoplasmic domain is required for the laminin assembly at the intersomitic boundaries. Developmental Dynamics 238:1332-1345, 2009. (c) 2008 Wiley-Liss, Inc.

Synthesis and evaluation of new arylbenzo[b]thiophene and diarylthiophene derivatives as inhibitors of the NorA multidrug transporter of Staphylococcus aureus.

The synthesis based on palladium catalytic coupling of 38 new-arylated benzo[b]thiophenes or thiophenes is described in a few steps. We also report the direct arylation of the position 3 of the benzo[b]thiophenic structure, a 'one pot' 2,5-heterodiarylation of thiophenes as well as the synthesis of precursors of amino-acids with a 2-arylated benzo[b]thiophene core. These compounds were evaluated on bacteria strains: most of them did not exhibit any antibiotic activity but were found to selectively inhibit the NorA multidrug transporter of Staphylococcus aureus. As such, they restored the activity of the NorA substrates ciprofloxacin against a resistant S. aureus strain in which this efflux pump is over-expressed.

Developmental dysplasia of the hip: quality of reporting of diagnostic accuracy for US.

PURPOSE: To systematically review the quality of diagnostic accuracy reporting in studies on the use of ultrasonography (US) for the diagnosis of developmental dysplasia of the hip (DDH). MATERIALS AND METHODS: A systematic review of the MEDLINE, EMBASE, DARE, and Cochrane Library databases was performed by using a validated search strategy. Two independent reviewers evaluated articles by using the Standards for Reporting of Diagnostic Accuracy (STARD) and Quality Assessment of Studies of Diagnostic Accuracy included in Systematic Reviews (QUADAS) statements. Items were reported individually for STARD and QUADAS because these instruments do not incorporate a summary score. A simple kappa statistic with 95% confidence intervals was used to measure the level of agreement between the two reviewers. RESULTS: Ten studies were included. In three studies, reliability was investigated, and in seven studies elements of both validity and reliability were investigated. In no study did the authors adequately report more than 40% of the STARD items. The quality of methods that were used in the studies was poor. Only one (14%) of seven studies provided information on more than 50% of the QUADAS items. All studies included a good description of image acquisition, but data analysis was imperfect and lacked estimates of diagnostic accuracy and precision. Authors tended to overinterpret their results. CONCLUSION: Overall, there was imperfect reporting of diagnostic accuracy in studies on the use of US for diagnosis of DDH.

Relocalization of an 82-kDa protein from lampbrush loops into the nucleoskeleton during amphibian oogenesis.

Affinity-purified antibodies directed against an 82-kDa oocyte nuclear protein ofPleurodeles waltl (Amphibia, Urodela) were prepared using antigen bound to nitrocellulose paper. The specificity of the antibody was controlled on two-dimensional electrophoretic gels of nuclear proteins. The intranuclear distribution of the 82-kDa protein was analyzed by the indirect immunofluorescence method on spreads of oocyte nuclear content. Localization of the protein appeared to be extremely variable. The antibody recognized the protein (a) on normal and landmark loop matrices (but not simultaneously), (b) on ribonucleoprotein particles associated (or not) with the nucleoskeleton and (c) on nucleoli. This suggests the intervention of the protein at a certain physiological moment in the transcriptional or posttranscriptional process.

Cloning and expression patterns of dystroglycan during the early development of Xenopus laevis.

Dystroglycan is a cell surface receptor involved in the pathogenesis of muscular dystrophy, and plays a critical role in the assembly and homeostasis of basement membranes. Since data about the amphibian homologue are limited, we have cloned the full-length dystroglycan cDNAs from the frog Xenopus laevis. Using in situ hybridization, we show that mRNA expression is dynamic, particularly in the notochord at the end of gastrulation and during neurulation, suggesting that the protein might play unexplored roles in the specification and/or formation of this tissue. Subsequently, the transcripts are detected in the otic vesicle, the developing brain, and in mesenchymal cells of the visceral arches, as well as in pharyngeal endoderm, the pronephros, pronephric ducts, proctodaeum and the somites.

Purification and inhibition by quinolones of DNA gyrases from Mycobacterium avium, Mycobacterium smegmatis and Mycobacterium fortuitum bv. peregrinum.

The DNA gyrases from Mycobacterium avium, Mycobacterium smegmatis and Mycobacterium fortuitum bv. peregrinum, which are species naturally resistant, moderately susceptible and susceptible to fluoroquinolones, respectively, were purified by affinity chromatography on novobiocin-Sepharose columns. The DNA gyrase inhibiting activities (IC50 values) of classical quinolones and fluoroquinolones were determined from the purified enzymes and were compared to the corresponding antibacterial activities (MICs). Regarding M. fortuitum bv. peregrinum, which is nearly as susceptible as Escherichia coli, the corresponding MIC and IC50 values of quinolones were significantly lower than those found for M. avium and M. smegmatis (e.g. for ofloxacin, MICs of 0.25 versus 32 and 1 microg ml(-1), and IC50 values of 1 versus 8 and 6 microg ml(-1), respectively). Such a result could be related to the presence of Ser-83 in the quinolone-resistance-determining region of the gyrase A subunit of M. fortuitum bv. peregrinum, as found in wild-type E. coli, instead of Ala-83 in M. avium and M. smegmatis, as found in fluoroquinolone-resistant E. coli mutants. The IC50 values of quinolones against the M. avium and M. smegmatis DNA gyrases were similar, while the corresponding MICs were 32-fold higher for M. avium when compared to M. smegmatis, suggesting that an additional mechanism, such as a low cell wall permeability or a drug efflux, could contribute to the low antibacterial potency of quinolones against M. avium.