Transport of amino acids and ammonium in mycelium of Agaricus bisporus.

Mycelium of Agaricus bisporus took up methylamine (MA), glutamate, glutamine and arginine by high-affinity transport systems following Michaelis-Menten kinetics. The activities of these systems were influenced by the nitrogen source used for mycelial growth. Moreover, MA, glutamate and glutamine uptakes were derepressed by nitrogen starvation, whereas arginine uptake was repressed. The two ammonium-specific transport systems with different affinities and capacities were inhibited by NH(+)(4), with a K(i) of 3.7 microM for the high-velocity system. The K(m) values for glutamate, glutamine and arginine transport were 124, 151 and 32 microM, respectively. Inhibition of arginine uptake by lysine and histidine showed that they are competitive inhibitors. MA, glutamate and glutamine uptake was inversely proportional to the intracellular NH(+)(4) concentration. Moreover, increase of the intracellular NH(+)(4) level caused by PPT (DL-phosphinotricin) resulted in an immediate cessation of MA, glutamine and glutamate uptake. It seems that the intracellular NH(+)(4) concentration regulates its own influx by feedback-inhibition of the uptake system and probably also its efflux which becomes apparent when mycelium is grown on protein. Addition of extracellular NH(+)(4) did not inhibit glutamine uptake, suggesting that NH(+)(4) and glutamine are equally preferred nitrogen sources. The physiological importance of these uptake systems for the utilization of nitrogen compounds by A. bisporus is discussed.     

Massive deformation of the scrotal wall by idiopathic calcinosis of the scrotum

Idiopathic calcinosis of the scrotum is a rare disease that may cause massive deformation of the scrotal wall. The first patient we present was also known to have neurofibromatosis. In the second patient we describe, nodules of idiopathic calcinosis of the scrotum were seen with walls that evidenced no epithelial lining, as well as calcification in epithelial cysts. At present, the only possible treatment is excision, and we excised the afflicted skin without problems in primary wound closure.     

Assignment of the human homologue of Pim-1, a mouse gene implicated in leukemogenesis,to the pter-q12 region of chromosome 6

Summary Viral leukemogenesis in mice is frequently initiated by proviral activation of a highly conserved cellular gene called Pim-1. Here we report the chromosomal localization of the human homologue by Southern blot analyses of DNAs obtained from human-rodent somatic cell hybrids. The single copy human homologue was assigned to the 6pter-q12 segment.     

NP7 protects from cell death induced by oxidative stress in neuronal and glial midbrain cultures from parkin null mice

Parkin mutations produce Parkinson’s disease (PD) in humans and nigrostriatal dopamine lesions related to increased free radicals in mice. We examined the effects of NP7, a synthetic, marine derived, free radical scavenger which enters the brain, on H₂O₂ toxicity in cultured neurons and glia from wild-type (WT) and parkin null mice (PK-KO).NP7, 5-10 μM, prevented the H₂O₂ induced apoptosis and necrosis of midbrain neuronal and glial cultures from WT and PK-KO mice. NP7 suppressed microglial activation and the H₂O₂ induced drop-out of dopamine neurons_. Furthermore, NP7 prevented the increased phosphorylation of ERK and AKT induced by H₂O₂. NP7 may be a promising neuroprotector against oxidative stress in PD.     

DIATOM SILICA BIOMINERALIZATION: AT NANOSCALE LEVEL A CHEMICALLY UNIFORM PROCESS

Using a high-brilliance synchrotron X-ray source, combined small- and wide-angle X-ray scattering (SAXS and WAXS) was applied to study nanoscale characteristics, in particular pore size in the range of 3 to 65 nm, of a variety of unialgal cultures of centric and pennate diatoms, and of mixed diatom populations sampled in the field. Results of scattering analysis were compared with details of pore size, structure and orientation visible at the electron microscopic level. WAXS patterns did not reveal any crystalline phase or features of microcrystallinity (resolution 0.07 to 0.51 nm), which implies a totally amorphous character of the SiO₂ matrix of the frustule material. SAXS data (resolution 3 to 65 nm) provided information on geometry, size, and distribution of pores in the silica. Overall, two pore regions were recognized that were common to the silica of all samples: the smallest (d less than 10 nm) regularly spaced and shaped spherically, the larger (up to 65 nm) being cylinders or slits. Apparently, at a nanoscale level diatomaceous silica is quite homologous among species, in agreement with the chemical principles of silica polymerization under the conditions of pH and precursor concentrations inside the silicon deposition vesicle. The final frustule "macro"-morphology is of course species-specific, being determined genetically. Synthetically-derived MCM-type silicas have a similarly organized pore distribution in an amorphous silica matrix as we found in all diatom species studied. We therefore suggest that organic molecules of a kind used as structure-directing agents to produce these artificial silicas play a role in the nucleation of the silica polymerization reaction and the shaping of pore morphology inside the silicon deposition vesicle of diatoms. Structure-directing molecules now await isolation from the SDV, followed by identification and characterisation by molecular techniques.     

Determination of deuterium-labeled tryptophan in proteins by means of high-performance liquid chromatography and thermospray mass spectrometry

In order to develop direct methods for determining the extent of metabolic incorporation of isotopically labeled amino acids into a protein, the determination of deuterated tryptophan in [2H5]tryptophan-bacteriorhodopsin was investigated. The isotopically modified protein was subjected to alkaline hydrolysis. After phenyl isothiocyanate derivatization of the hydrolysate, the mixture was separated by reversed-phase liquid chromatography. Field desorption mass spectrometry and thermospray mass spectrometry were investigated for their ability to determine the ratio between [2H5]tryptophan and total tryptophan in the collected fractions. In order to check the procedure a set of known tryptophan/[2H5]tryptophan mixtures were passed through the same derivatization, HPLC separation, and lyophilization procedure as used for the biological samples.     

Hydrophobia, lectin binding, and molecular order in the stratum corneum of adult and neonatal rats and in human breast cells in culture.

A search for differences due to ANS staining (hydrophobia), Con A and PNA binding capacity, and birefringence was carried out on stratified epithelia of rat skin and human breast cells (HBC) in culture. Microfluorimetric measurements confirm that the ANS fluorescence of the stratum corneum from adults is higher than that of newborns. HBC exhibited an unexpected deep ANS-fluorescence. Differences in the binding capacity of the epithelial layers to Con A and PNA were detected with advancing age. Retardation measurements revealed that the form birefringence of the stratum corneum is higher in adult animals specially as revealed by the fact that its form birefringence curve branch from n = 1.414 to n = 1.479 is steeper, i.e. depict higher values. The strong birefringence of the cytoplasmic tonofilaments presented by cultured human breast cells was considered an unexpected finding and attributed to changes that the cells underwent following the in vitro conditions.     

Evolution of sprouting versus seeding inAspalathus linearis

We have tested the hypothesis that reseeding is a plesiomorphic character state and that sprouting is a derived state inAspalathus linearis, and that the latter is an adaptation to ensure fire-survival in a fireprone environment. Samples of five seeder and four sprouter populations of A.linearis were examined by horizontal starch gel electrophoresis to assess the amount of genetic differentiation within and between sprouting and seeding populations, and to determine the extent of gene flow between the populations. Leaf extracts were surveyed for ten enzymes and gene products revealed genetic variation at 13 (76%) of 17 protein coding loci. Allele frequency differences were found between sprouting and seeding populations and genetic distance values show that the sprouters are grouped separate from the seeders, thus providing support for the morphological data on which the above mentioned hypothesis is based. It is evident that evolution operates at the population level inA. linearis.