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Modulation of Fibronectin Adhesins and Other Virulence Factors in a Teicoplanin-Resistant Derivative of Methicillin-Resistant Staphylococcus aureus. Adriana Renzoni, 2004.The impact of glycopeptide resistance on the molecular regulation of Staphylococcus aureus virulence and attachment to host tissues is poorly documented . We compared stable teicoplanin-resistant methicillin-resistant S . aureus (MRSA) strain 14-4 with its teicoplanin-susceptible MRSA parent, strain MRGR3, which exhibits a high degree of virulence in a rat model of chronic foreign body MRSA infection . The levels of fibronectin-mediated adhesion and surface display of fibronectin-binding proteins were higher in teicoplanin-resistant strain 14-4 than in its teicoplanin-susceptible parent or a teicoplanin-susceptible revertant (strain 14-4rev) that spontaneously emerged during tissue cage infection . Quantitative reverse transcription-PCR (qRT-PCR) showed four- and twofold higher steady-state levels of fnbA and fnbB transcripts, respectively, in strain 14-4 than in its teicoplanin-susceptible counterparts . Analysis of global regulatory activities by qRT-PCR revealed a strong reduction in the steady-state levels of RNAIII and RNAII in the teicoplanin-resistant strain compared to in its teicoplanin-susceptible counterparts . In contrast, sarA mRNA levels were more than fivefold higher in strain 14-4 than in MRGR3 and 14-4rev . Furthermore, the alternative transcription factor sigma B had a higher level of functional activity in the teicoplanin-resistant strain than in its teicoplanin-susceptible counterparts, as evidenced by significant increases in both the sigma B-dependent asp23 mRNA levels and the sarA P3 promoter-derived transcript levels, as assayed by qRT-PCR and Northern blotting, respectively . These data provide further evidence that the emergence of glycopeptide resistance is linked by still poorly understood molecular pathways with significant pleiotropic changes in the expression and regulation of some major virulence genes . These molecular and phenotypic changes may have a profound impact on the bacterial adhesion and colonization properties of such multiresistant organisms . CTP Limitation Increases Expression of CTP Synthase in Lactococcus lactis. Casper Møller Jørgensen, 2003.CTP synthase is encoded by the pyrG gene and catalyzes the conversion of UTP to CTP . A Lactococcus lactis pyrG mutant with a cytidine requirement was constructed, in which ß-galactosidase activity in a pyrG-lacLM transcriptional fusion was used to monitor gene expression of pyrG . A 10-fold decrease in the CTP pool induced by cytidine limitation was found to immediately increase expression of the L . lactis pyrG gene . The final level of expression of pyrG is 37-fold higher than the uninduced level . CTP limitation has pronounced effects on central cellular metabolism, and both RNA and protein syntheses are inhibited . Expression of pyrG responds only to the cellular level of CTP, since expression of pyrG has no correlation to alterations in UTP, GTP, and ATP pool sizes . In the untranslated pyrG leader sequence a potential terminator structure can be identified, and this structure is required for regulation of the pyrG gene . It is possible to fold the pyrG leader in an alternative structure that would prevent the formation of the terminator . We suggest a model for pyrG regulation in L . lactis, and probably in other gram-positive bacteria as well, in which pyrG expression is directly dependent on the CTP concentration through an attenuator mechanism . At normal CTP concentrations a terminator is preferentially formed in the pyrG leader, thereby reducing expression of CTP synthase . At low CTP concentrations the RNA polymerase pauses at a stretch of C residues in the pyrG leader, thereby allowing an antiterminator to form and transcription to proceed . This model therefore does not include any trans-acting protein for sensing the CTP concentration as previously proposed for Bacillus subtilis . Complexity of Gas Vesicle Biogenesis in Halobacterium sp . Strain NRC-1: Identification of Five New Proteins. Hem Dutt Shukla, 2004.The genome of Halobacterium sp . strain NRC-1 contains a large gene cluster, gvpMLKJIHGFEDACNO, that is both necessary and sufficient for the production of buoyant gas-filled vesicles . Due to the resistance of gas vesicles to solubilization, only the major gas vesicle protein GvpA and a single minor protein, GvpC, were previously detected . Here, we used immunoblotting analysis to probe for the presence of gas vesicle proteins corresponding to five additional gvp gene products . Polyclonal antisera were raised in rabbits against LacZ-GvpF, -GvpJ, and -GvpM fusion proteins and against synthetic 15-amino-acid peptides from GvpG and -L . Immunoblotting analysis was performed on cell lysates of wild-type Halobacterium sp . strain NRC-1, gas vesicle-deficient mutants, and purified gas vesicles, after purification of LacZ fusion antibodies on protein A and ß-galactosidase affinity columns . Our results show the presence of five new gas vesicle proteins (GvpF, GvpG, GvpJ, GvpL, and GvpM), bringing the total number of proteins identified in the organelles to seven . Two of the new gas vesicle proteins are similar to GvpA (GvpJ and GvpM), and two proteins contain predicted coiled-coil domains (GvpF and GvpL) . GvpL exhibited a multiplet ladder on sodium dodecyl sulfate-polyacrylamide gels indicative of oligomerization and self-assembly . We discuss the possible functions of the newly discovered gas vesicle proteins in biogenesis of these unique prokaryotic flotation organelles . Hydroxycinnamate (hca) Catabolic Genes from Acinetobacter sp . Strain ADP1 Are Repressed by HcaR and Are Induced by Hydroxycinnamoyl-Coenzyme A Thioesters. Donna Parke, 2003.
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