An in situ study of beta-glucosidase activity in normal and gaucher fibroblasts with fluorogenic probes

Beta-glucosidase activity was evaluated in situ by means of fluorogenic probes in normal human fibroblasts and fibroblasts from homozygous carriers of the Gaucher trait. Probe internalization, targeting to lysosomes and post-cleavage probe retention were the primary concerns. Internalization and targeting were attempted by in situ photosensitized labilization of lysosomal membranes, lysosomotropic detergents and the use of low density lipid (LDL) or the receptor ligand apolipoprotein E (ApoE). Post-cleavage increase of fluorescence with fluoresceinyl (bis) betaglucopyranoside was appreciably above the rather large pre-cleavage emission. In cells incubated overnight with nonylumbelliferylbetaglucoside (UG9) in the presence of bovine serum albumin and in the absence of ApoE, the probe was dealt with as a cytotoxic agent, accumulating in a paranuclear cap, most likely comprising elements of the endoplasmic reticulum (ER) and Golgi apparatus. Targeting of UG9 to lysosomes occurred within 1 to 3 h of preincubation in the presence of ApoE. There was some evidence of specificity, as Gaucher fibroblasts exhibited weaker cleavage of UG9 (by 50 per cent or more) compared to normal fibroblasts, but in the Gaucher cells there was some residual beta-glucosidase activity. Cleavage of UG9 was nearly totally suppressed in Gaucher cells treated with the beta-glucosidase inhibitor, conduritol B epoxide, for 24 h to 7 days. Suppression in the control fibroblasts was evident but to a lesser degree. The in situ method of fluorogenic assay established for beta-glucosidase deficiency, is in principle applicable to enzyme deficiencies in other lysosomal storage diseases, or to evaluate enhanced enzyme activity following gene therapy.     

Molecular Exploration of the First-Century Tomb of the Shroud in Akeldama,Jerusalem

The Tomb of the Shroud is a first-century C.E. tomb discovered in Akeldama, Jerusalem, Israel that had been illegally entered and looted. The investigation of this tomb by an interdisciplinary team of researchers began in 2000. More than twenty stone ossuaries for collecting human bones were found, along with textiles from a burial shroud, hair and skeletal remains. The research presented here focuses on genetic analysis of the bioarchaeological remains from the tomb using mitochondrial DNA to examine familial relationships of the individuals within the tomb and molecular screening for the presence of disease. There are three mitochondrial haplotypes shared between a number of the remains analyzed suggesting a possible family tomb. There were two pathogens genetically detected within the collection of osteological samples, these were Mycobacterium tuberculosis and Mycobacterium leprae. The Tomb of the Shroud is one of very few examples of a preserved shrouded human burial and the only example of a plaster sealed loculus with remains genetically confirmed to have belonged to a shrouded male individual that suffered from tuberculosis and leprosy dating to the first-century C.E. This is the earliest case of leprosy with a confirmed date in which M. leprae DNA was detected.     

A compatible interaction of Alternaria brassicicola with Arabidopsis thaliana ecotype DiG: evidence for a specific transcriptional signature

Background  

The interaction of Arabidopsis with Alternaria brassicicola provides a model for disease caused by necrotrophs, but a drawback has been the lack of a compatible pathosystem. Infection of most ecotypes, including the widely-studied line Col-0, with this pathogen generally leads to a lesion that does not expand beyond the inoculated area. This study examines an ecotype, Dijon G (DiG), which is considered sensitive to A. brassicicola.     

Class similarity and viewpoint invariance in the recognition of 3D objects

 In human vision, the processes and the representations involved in identifying specific individuals are frequently assumed to be different from those used for basic level classification, because classification is largely viewpoint-invariant, but identification is not. This assumption was tested in psychophysical experiments, in which objective similarity between stimuli (and, consequently, the level of their distinction) varied in a controlled fashion. Subjects were trained to discriminate between two classes of computer-generated three-dimensional objects, one resembling monkeys and the other, dogs. Both classes were defined by the same set of 56 parameters, which encoded sizes, shapes, and placement of the limbs, ears, snout, etc. Interpolation between parameter vectors of the class prototypes yielded shapes that changed smoothly between monkey and dog. Within-class variation was induced in each trial by randomly perturbing all the parameters. After the subjects reached 90% correct performance on a fixed canonical view of each object, discrimination performance was tested for novel views that differed by up to 60° from the training view. In experiment 1 (in which the distribution of parameters in each class was unimodal) and in experiment 2 (bimodal classes), the stimuli differed only parametrically and consisted of the same geons (parts), yet were recognized virtually independently of viewpoint in the low-similarity condition. In experiment 3, the prototypes differed in their arrangement of geons, yet the subjects’ performance depended significantly on viewpoint in the high-similarity condition. In all three experiments, higher interstimulus similarity was associated with an increase in the mean error rate and, for misorientation of up to 45°, with an increase in the degree of viewpoint dependence. These results suggest that a geon-level difference between stimuli is neither strictly necessary nor always sufficient for viewpoint-invariant performance. Thus, basic and subordinate-level processes in visual recognition may be more closely related than previously thought. Received: 15 November 1993/Accepted in revised form: 14 July 1994     

Attempts to control sand flies by insecticide-sprayed strips along the periphery of a village.

The customary method for controlling leishmaniasis and sand fly bites in Israel is the spraying of large quantities of residual insecticides on walls of houses and neighboring surfaces. However, the high summer temperatures, strong radiation, and dust limit the efficacy of the method. The sand fly vectors of Leishmania appeared to come uphill to the village of Kfar Adumim. Therefore, to block their path and improve control, beta-cyfluthrine or DDT were repeatedly sprayed on a strip of cloth that was stretched on the ground along and near the peripheral houses. The efficacy of the treatments was examined by comparing the proportions of sand flies that had been caught before and after the treatments, below and above the belt. The capture near the houses of sand flies that had been marked by colored sugar baits below the protective belt indicated that they were moving uphill and crossing the strip. The catches showed a gradual uphill decrease, up to the houses, in the number of sand flies. However, the proportions of sand flies caught in the different sites were similar before and after insecticide treatments showing that the belt was ineffective. Laboratory tests of pieces of the sprayed strip and exposure of sand flies to similar pieces indicated that the lack of effect was not caused by deficiency of insecticides.     

香石竹花瓣中乙烯诱导合成的60kD蛋白功能的初步研究

越来越多的研究表明植物器官的衰老与蛋白质合成密切相关[1～3]。乙烯在许多花卉衰老和一些水果的成熟过程中起重要作用[1,3],实验结果显示花对乙烯的反应依赖于新的蛋白质合成[2～5]。深入研究那些与衰老相关的蛋白质,特别是乙烯诱导合成的蛋白质的生化功能,对于了解掌握...     

Structural and magnetic behavior of mono- and dinuclear nickel (II) complexes of N,N′-bis-(3,5-dipiperidin-1-yl-[2,4,6]triazin-1-yl)-pyridin-2-ylmethyl-ethane-1,2-diamine

Reaction of nickel (II) perchlorate with the ligand N,N′-bis-(3,5-dipiperidin-1-yl-[2,4,6]triazin-1-yl)-pyridin-2-ylmethyl-ethane-1,2-diamine yields an octahedral Ni(II) high-spin complex 1 ([C₄₀H₅₆N₁₄Ni(H₂O)(CH₃OH)](ClO₄)₂(CH₃OH)₂) with moderate zero-field splitting (ZFS) axial distortion parameter D/k_B = 5.37 K. The ligand contributes a N4 donor set; the remaining two coordinating positions are occupied by coordinating solvents molecules. Exchange of the coordinating solvents molecules in complex 1 to thiocyanate moieties leads to formation of complex 2 ([C₄₀H₅₆N₁₄Ni(NCS)₂](CHCl)₃) with an extended parameter D/k_B = 8.80 K. The analysis of the structural and magnetic properties of complexes 1 and 2 led to the design of dinuclear complex 3 ([C₄₀H₅₆N₁₄NiN₃]₂(ClO₄)₂(CH₃OH)₂), where two azido groups were utilized as bridging ligands. The double azido bridges in complex 3 cross each other to form a rarely observed non-coplanar (N₃)₂ structure. The magnetic behavior of complex 3 reveals ferromagnetic coupling interactions characterized by J/k_B = 23.25 K, D₁/k_B = 7.90 K, D₂/k_B = 0.54 K.