Water flow from soil to plants depends on the properties of the soil next to roots, the rhizosphere. Although several studies showed that the rhizosphere has different properties than the bulk soil, effects of the rhizosphere on root water uptake are commonly neglected. To investigate the rhizosphere's properties we used neutron radiography to image water content distributions in soil samples planted with lupins during drying and subsequent rewetting. During drying, the water content in the rhizosphere was 0.05 larger than in the bulk soil. Immediately after rewetting, the picture reversed and the rhizosphere remained markedly dry. During the following days the water content of the rhizosphere increased and after 60 h it exceeded that of the bulk soil. The rhizosphere's thickness was approximately 1.5 mm. Based on the observed dynamics, we derived the distinct, hysteretic and time-dependent water retention curve of the rhizosphere. Our hypothesis is that the rhizosphere's water retention curve was determined by mucilage exuded by roots. The rhizosphere properties reduce water depletion around roots and weaken the drop of water potential towards roots, therefore favoring water uptake under dry conditions, as demonstrated by means of analytical calculation of water flow to a single root.

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Resistance against ␤-lactam antibiotics is a growing challenge for managing severe bacterial infections. The rapid and costefficient determination of ␤-lactam resistance is an important prerequisite for the choice of an adequate antibiotic therapy. ␤-Lactam resistance is based mainly on the expression/overexpression of ␤-lactamases, which destroy the central ␤-lactam ring of these drugs by hydrolysis. Hydrolysis corresponds to a mass shift of ؉18 Da, which can be easily detected by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Therefore, a MALDI-TOF MS-based assay was set up to investigate different enterobacteria for resistance against different ␤-lactam antibiotics: ampicillin, piperacillin, cefotaxime, ceftazidime, ertapenem, imipenem, and meropenem. ␤-Lactamases are enzymes that have a high turnover rate. Therefore, hydrolysis can be detected by MALDI-TOF MS already after a few hours of incubation of the bacteria to be tested with the given antibiotic. The comparison of the MS-derived data with the data from the routine procedure revealed identical classification of the bacteria according to sensitivity and resistance. The MALDI-TOF MS-based assay delivers the results on the same day. The approved routine procedures require at least an additional overnight incubation.

VAMP proteins are important components of the machinery controlling docking and/or fusion of secretory vesicles with their target membrane. We investigated the expression of VAMP proteins in pancreatic beta‐cells and their implication in the exocytosis of insulin. cDNA cloning revealed that VAMP‐2 and cellubrevin, but not VAMP‐1, are expressed in rat pancreatic islets and that their sequence is identical to that isolated from rat brain. Pancreatic beta‐cells contain secretory granules that store and secrete insulin as well as synaptic‐like microvesicles carrying gamma‐aminobutyric acid. After subcellular fractionation on continuous sucrose gradients, VAMP‐2 and cellubrevin were found to be associated with both types of secretory vesicle. The association of VAMP‐2 with insulin‐containing granules was confirmed by confocal microscopy of primary cultures of rat pancreatic beta‐cells. Pretreatment of streptolysin‐O permeabilized insulin‐secreting cells with tetanus and botulinum B neurotoxins selectively cleaved VAMP‐2 and cellubrevin and abolished Ca(2+)‐induced insulin release (IC50 approximately 15 nM). By contrast, the pretreatment with tetanus and botulinum B neurotoxins did not prevent GTP gamma S‐stimulated insulin secretion. Taken together, our results show that pancreatic beta‐cells express VAMP‐2 and cellubrevin and that one or both of these proteins selectively control Ca(2+)‐mediated insulin secretion.

Abstract. SNAP-25 is known as a neuron specific molecule involved in the fusion of small synaptic vesicles with the presynaptic plasma membrane. By immunolocalization and Western blot analysis, it is now shown that SNAP-25 is also expressed in pancreatic endocrine cells. Botulinum neurotoxins (BoNT) A and E were used to study the role of SNAP-25 in insulin secretion. These neurotoxins inhibit transmitter release by cleaving SNAP-25 in neurons.Cells from a pancreatic B cell line (HIT) and primary rat islet cells were permeabilized with streptolysin-O to allow toxin entry. SNAP-25 was cleaved by BoNT/A and BoNT/E, resulting in a molecular mass shift of ~ol and 3 kD, respectively. Cleavage was accompanied by an inhibition of Ca÷÷-stimulated insulin release in both cell types. In HIT cells, a concentration of 30-40 nM BoNT/E gave maximal inhibition of stimulated insulin secretion of '~60%, coinciding with essentially complete cleavage of SNAP-25. Half maximal effects in terms of cleavage and inhibition of insulin release were obtained at a concentration of 5-10 nM. The A type toxin showed maximal and halfmaximal effects at concentrations of 4 and 2 nM, respectively. In conclusion, the results suggest a role for SNAP-25 in fusion of dense core secretory granules with the plasma membrane in an endocrine cell typethe pancreatic B cell. IN pancreatic B cells, proinsulin is sorted in the transGolgi network for delivery to secretory granules where it is processed to insulin (23,40). Insulin is packed and stored in large dense core granules (LDCG) ~ and is released when exocytosis is stimulated by secretagogues such as glucose (23). Current understanding of B cell stimulussecretion coupling suggests that nutrient stimuli cause depolarization of the cell membrane, which leads to an influx of Ca ++ triggering the fusion of granules with the plasma membrane (48). Although sensitivity to glucose is a peculiarity of the pancreatic B cell, the other steps leading from the trans-Golgi network to LDCGs and the fusion of granules with the plasma membrane are, most probably, Please address all correspondence to Dr. Karin Sadoul, Laboratoires de Recherche Louis Jeantet, Centre Medical Universitaire, 1, rue MichelServet, CH-1211 Geneva 4, Switzerland. Tel.: 41 22 7025537. Fax.: 41 22 3473334. Abbreviations used in this paper:BoNT, botulinum neurotoxin; LDCG, large dense core granule; LDCV, large dense core vesicle; NSE N-ethylmaleimid-sensitive factor; SLMV, synaptic-like microvesicle; SNAP, soluble NSF attachment protein; SNAP-25, synaptosomal-associated protein of 25 kD; SSV, small synaptic vesicle.common to all cells possessing the regulated secretory pathway.The molecular mechanism for docking and fusion of LDCGs in endocrine cells has not so far been studied. Conversely, recent data from S611ner et al. have established a detailed model for docking and fusion of small synaptic vesicles (SSV) in neuronal cells (57,58). Using cosedimentation and immunoprecipitation techniques they could identify a 20-S fusion complex. The complex is...