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Phase-variable expression of type 1 fimbriae in Escherichia coli K-12 involves inversion by site-specific recombination of a 314 bp sequence containing the promoter for fim structural gene expression. The invertible sequence is flanked by 9 bp inverted repeats, and each repeat is in turn flanked by non-identical recombinase-binding elements (RBEs) to which the FimB or FimE site-specific recombinases bind. These proteins have distinct DNA inversion preferences: FimB inverts the switch in the ON-to-OFF and OFF-to-ON directions with similar efficiencies, whereas FimE inverts it predominantly in the ON-to-OFF direction. We have found that FimB and FimE invert the switch through a common mechanism. A genetic investigation involving base-by-base substitution combined with a biochemical study shows that the same DNA cleavage and religation sites are used within the 9 bp inverted repeats, and that each recombination involves a common 3 bp spacer region. A comprehensive programme of RBE exchanges and replacements reveals that FimB is much more tolerant of RBE sequence variation than FimE. The asymmetric location of conserved 5'-CA motifs at either side of each spacer region allows the inside and outside of the switch to be differentiated while the RBE sequence heterogeneity permits its ON and OFF forms to be distinguished by the recombinases. 相似文献
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Freeman Ernest J.; Ruehr Mary L.; Dorman Robert V. 《American journal of physiology. Cell physiology》1998,274(1):C282
The accumulation of radiolabeled arachidonicacid (AA), immunoblot analysis of subcellular fractions, andimmunofluorescence tagging of proteins in intact cells were used toexamine the coupling of ANG II receptors with the activity and locationof a cytosolic phospholipase A2(cPLA2) in vascular smoothmuscle cells (VSMC). ANG II induced the accumulation of AA, whichpeaked by 10 min and was downregulated by 20 min. A large proportion ofthe AA released in response to ANG II was due to the activation of a Ca2+-dependent lipase coupled toan AT1 receptor. However,regulation of Ca2+ availabilityfailed to completely block AA release, and a small but significantreduction in ANG II-mediated AA release was observed in the presence ofan AT2 antagonist. These findings,coupled with a 25% reduction in the ANG II-induced AA release by aninhibitor specific for aCa2+-independentPLA2, are consistent with thepresence and activation of aCa2+-independentPLA2. In contrast, immunoblotanalysis and immunofluorescence detection showed that the ANGII-mediated translocation of cPLA2 to a membrane fraction was exclusivelyAT1 dependent and regulated byCa2+ availability. Furthermore,the nucleus was the membrane target. We conclude that ANG II regulatesthe Ca2+-dependent activation andtranslocation of cPLA2 through anAT1 receptor and that this eventis targeted at the nucleus in VSMC. 相似文献
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An enhancement of glutamate release from hippocampal neurons has been implicated in long-term potentiation, which is thought to be a cellular correlate of learning and memory. This phenomenom appears to be involved the activation of protein kinase C and lipid second messengers have been implicated in this process. The purpose of this study was to examine how lipid-derived second messengers, which are known to potentiate glutamate release, influence the accumulation of intraterminal free Ca2+, since exocytosis requires Ca2+ and a potentiation of Ca2+ accumulation may provide a molecular mechanism for enhancing glutamate release. The activation of protein kinase C with phorbol esters potentiates the depolarization-evoked release of glutamate from mossy fiber and other hippocampal nerve terminals. Here we show that the activation of protein kinase C also enhances evoked presynaptic Ca2+ accumulation and this effect is attenuated by the protein kinase C inhibitor staurosporine. In addition, the protein kinase C-dependent increase in evoked Ca2+ accumulation was reduced by inhibitors of phospholipase A2 and voltage-sensitive Ca2+ channels, as well as by a lipoxygenase product of arachidonic acid metabolism. That some of the effects of protein kinase C activation were mediated through phospholipase A2 was also indicated by the ability of staurosporine to reduce the Ca2+ accumulation induced by arachidonic acid or the phospholipase A2 activator melittin. Similarly, the synergistic facilitation of evoked Ca2+ accumulation induced by a combination of arachidonic acid and diacylglycerol analogs was attenuated by staurosporine. We suggest, therefore, that the protein kinase C-dependent potentiation of evoked glutamate release is reflected by increases in presynaptic Ca2+ and that the lipid second messengers play a central role in this enhancement of chemical transmission processes. 相似文献
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Jeffrey G. Dorman Frederic S. Castruccio Enrique N. Curchitser Joan A. Kleypas Thomas M. Powell 《Coral reefs (Online)》2016,35(1):169-179
The Spratly Island archipelago is a remote network of coral reefs and islands in the South China Sea that is a likely source of coral larvae to the greater region, but about which little is known. Using a particle-tracking model driven by oceanographic data from the Coral Triangle region, we simulated both spring and fall spawning events of Acropora millepora, a common coral species, over a 46-yr period (1960–2005). Simulated population biology of A. millepora included the acquisition and loss of competency, settlement over appropriate benthic habitat, and mortality based on experimental data. The simulations aimed to provide insights into the connectivity of reefs within the Spratly Islands, the settlement of larvae on reefs of the greater South China Sea, and the potential dispersal range of reef organisms from the Spratly Islands. Results suggest that (1) the Spratly Islands may be a significant source of A. millepora larvae for the Palawan reefs (Philippines) and some of the most isolated reefs of the South China Sea; and (2) the relatively isolated western Spratly Islands have limited source reefs supplying them with larvae and fewer of their larvae successfully settling on other reefs. Examination of particle dispersal without biology (settlement and mortality) suggests that larval connectivity is possible throughout the South China Sea and into the Coral Triangle region. Strong differences in the spring versus fall larval connectivity and dispersal highlight the need for a greater understanding of spawning dynamics of the region. This study confirms that the Spratly Islands are likely an important source of larvae for the South China Sea and Coral Triangle region. 相似文献
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