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481.
Munc13-1 is required for the sustained release of insulin from pancreatic beta cells 总被引:1,自引:0,他引:1
Munc13-1 is a presynaptic protein that is essential for synaptic vesicle priming. Deletion of Munc13-1/unc13 causes total arrest of synaptic transmission due to a complete loss of fusion-competent synaptic vesicles. The requirement of Munc13-1 for large dense-core vesicles (LDCVs), however, has not been established. In the present study, we use Munc13-1 knockout (KO) and diacylglycerol (DAG) binding-deficient Munc13-1H567K mutant knockin (KI) mice to determine the role of Munc13-1 in the secretion of insulin-containing LDCVs from primary cultured pancreatic β cells. We show that Munc13-1 is required for the sustained insulin release upon prolonged stimulation. The sustained release involves signaling of DAG second messenger, since it is also reduced in KI mice. Insulin secretion in response to glucose stimulation is characterized by a biphasic time course. Our data show that Munc13-1 plays an essential role in the development of the second phase of insulin secretion by priming insulin-containing LDCVs. 相似文献
482.
Spiders are omnipresent, occurring in almost all terrestrial habitats. They are generalist predators and important for conservation biocontrol in agricultural fields, helping to reduce pesticide applications. In this study, we examined how spider families and spider web types in rice fields are related to local and landscape management. Samples were taken in differently managed rice fields adjoining either homegarden polycultures or banana monocultures. Furthermore, landscape structure, prey abundance, herb richness and cover were taken into account. The results showed that prey availability explained most of the variation in spiders and their web’s abundance, indicating that spiders in the rice fields can potentially contribute to pest control. Adjacent habitat had no effect on the spider populations, but maintaining fallow fields in the surrounding landscape seems to be a useful measure to promote Erigoninae in rice fields. There was no evidence that local management practices such as fertiliser and pesticide use had an impact on spider families, which appeared to be due to the low level of these inputs. Spider web sampling can complement spider sampling as it detects spiders hidden at the base of the rice tillers, which are likely to be missed by sweep netting. Additionally, tetragnathid webs are easy to observe and thus can be used as an indication for farmers not to spray pesticides as spiders are potentially controlling the pest species. Interviews with farmers made clear that many farmers in the study area showed their interest in management methods that promote biological pest control. 相似文献
483.
Genetic analysis of amidase mutants of Pseudomonas aeruginosa 总被引:8,自引:0,他引:8
484.
Structure of Saccharomyces cerevisiae mating hormone a-factor. Identification of S-farnesyl cysteine as a structural component 总被引:45,自引:0,他引:45
R J Anderegg R Betz S A Carr J W Crabb W Duntze 《The Journal of biological chemistry》1988,263(34):18236-18240
Mating type a cells of the yeast Saccharomyces cerevisiae produce a mating hormone, the a-factor, that we have previously characterized as a very hydrophobic, modified dodecapeptide (Betz, R., Crabb, J. W., Meyer, H. E., Wittig, R., and Duntze, W. (1987) J. Biol. Chem. 262, 546-548). We have investigated the molecular structure in detail using mass spectrometry and proton NMR spectrometry of the intact hormone and authentic component molecules. Tandem mass spectrometry confirms the previously determined peptide sequence of the hormone and shows that it contains additional structural components with masses of 205 and 15 daltons. These were identified by proton NMR and mass spectrometry as a farnesyl (C15H25) residue and a terminal methyl ester group. The farnesyl moiety is attached to the sulfur atom of the carboxyl-terminal cysteine residue, as revealed by NMR of synthetic S-farnesyl cysteine methyl ester. The stereochemical configuration of the farnesyl moiety was determined to be trans,trans by comparison of gas chromatography retention times, mass spectra, and NMR spectra with those of standards. These results define the structure of a-factor as: (Sequence: see text). Replacement of the farnesyl by a methyl group leads to a partial reduction in specific biological activity of the a-factor, whereas hydrolysis of the carboxyl-terminal methyl ester causes a complete loss of activity. 相似文献
485.
Clostridial neurotoxins compromise the stability of a low energy SNARE complex mediating NSF activation of synaptic vesicle fusion. 总被引:7,自引:0,他引:7
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A 20S complex composed of the cytosolic fusion proteins NSF and SNAP and the synaptosomal SNAP receptors (SNAREs) synaptobrevin, syntaxin and SNAP-25 is essential for synaptic vesicle exocytosis. Formation of this complex is thought to be regulated by synaptotagmin, the putative calcium sensor of neurotransmitter release. Here we have examined how different inhibitors of neurotransmitter release, e.g. clostridial neurotoxins and a synaptotagmin peptide, affect the properties of the 20S complex. Cleavage of synaptobrevin and SNAP-25 by the neurotoxic clostridial proteases tetanus toxin and botulinum toxin A had no effect on assembly and disassembly of the 20S complex; however, the stability of its SDS-resistant SNARE core was compromised. This SDS-resistant low energy conformation of the SNAREs constitutes the physiological target of NSF, as indicated by its ATP-dependent disassembly in the presence of SNAP and NSF. Synaptotagmin peptides caused inhibition of in vitro binding of this protein to the SNAREs, a result that is inconsistent with synaptotagmin's proposed role as a regulator of SNAP binding. Our data can be reconciled by the idea that NSF and SNAP generate synaptotagmin-containing intermediates in synaptic vesicle fusion, which catalyse neurotransmitter release. 相似文献
486.
Betz J Bauwens A Kunsmann L Bielaszewska M Mormann M Humpf HU Karch H Friedrich AW Müthing J 《Biological chemistry》2012,393(3):133-147
Membrane microdomain association of the glycosphingolipids (GSLs) globotriaosylceramide (Gb3Cer) and globotetraosylceramide (Gb4Cer), the highly and less effective receptors, respectively, for Shiga toxins (Stxs), is assumed as a functional requirement for Stx-mediated cytotoxicity. In a previous study, we demonstrated predominant localization of Stx receptors in cholesterol-enriched membrane microdomains of moderately Stx-sensitive human brain microvascular endothelial cells (HBMECs) by means of detergent-resistant membranes (DRMs). Here we report a different preferential distribution of Stx receptors in non-DRM fractions of human glomerular microvascular endothelial cells (GMVECs), the major targets of Stxs in the human kidney. Full structural characterization of Stx receptors using electrospray ionization (ESI) mass spectrometry revealed Gb3Cer and Gb4Cer lipoforms with ceramide moieties mainly composed of C24:0/C24:1 or C16:0 fatty acid and sphingosine (d18:1) in GMVECs comparable to those previously found in HBMECs. Thin-layer chromatography immunostaining demonstrated an approximately 2-fold higher content of Gb3Cer and a 1.4-fold higher content of Gb4Cer in GMVECs than in HBMECs. However, this does not explain the remarkable higher cytotoxic action of Stx1 and Stx2 toward GMVECs as compared with HBMECs. Our finding opens new questions on the microdomain association of Stx receptors and the functional role of GSLs in the membrane assembly of GMVECs. 相似文献
487.
Use of isolated brain capillaries and cultured endothelial cells to study the blood-brain barrier 总被引:5,自引:0,他引:5
Brain capillary endothelial cells are responsible for forming the blood-brain barrier (BBB). Methods are now available to isolate microvessels from brain and study their biochemical and transport characteristics. From these investigations, new ideas have been proposed concerning the role of endothelial cells in the function of the BBB. More recently, success in culturing endothelial cells from brain microvessels has opened the way for novel approaches to the study of the regulation of endothelial cell permeability. We anticipate continued rapid progress in this area and expect that this will lead to a better understanding of the mechanisms involved in the regulation of BBB permeability and brain capillary function. 相似文献
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