Effects of X-irradiation induced loss of cerebellar granule cells on the synaptosomal levels and the high affinity uptake of amino acids.

Abstract— Crude synaptosomal (P₂) preparations were obtained from the cerebella of rats in which the granule cell population had been selectively reduced by X-irradiation treatment and from the cerebella of control animals. In the P₂ fraction from control cerebella, the level of glutamate was greater than any other of the 5 amino acids measured and was 2-fold higher than taurine, which was present at the next highest level. The content of taurine was slightly higher than that found for aspartate and was 3-fold greater than that observed for GABA. Alanine and glycine were present in the lowest amounts. The levels of glutamate and aspartate were significantly (P < 0.05) lower by 25 and 15%, respectively, in the P₂ fraction isolated from the X-irradiated cerebella in comparison to control values. The content of taurine, GABA, glycine, and alanine were not changed by the X-irradiation treatment. The uptake of 1.0 μm -l -[³H]glutamate and l -[³H]aspartate was reduced approx 20% by X-irradiation treatment, whereas the uptake of 1.0 μm -[³H]GABA and [³H]taurine was unchanged. A more detailed kinetic analysis of l -[³H]glutamate uptake revealed there was a 20% decrease in the V_max value with X-irradiation treatment and no change in the apparent K_m value. In a second study, the uptake of l -[³H]glutamate, l -[³H]aspartate and [³H]GABA was measured using P₂ fractions obtained from the cerebella of rats in which the population of granule, stellate and basket cells had been reduced by X-irradiation treatment. The uptake of 1.0μm -l -[³H]glutamate, l -[³H]aspartate and [³H]GABA was significantly (P < 0.05) reduced to 57, 68, and 59%, respectively, of control values. A more detailed kinetic analysis of [³H]GABA uptake revealed no significant change in the apparent K_m and a 35% decrease in the V_max value. The data are discussed in terms of glutamate being the excitatory neurotransmitter released from granule cells and GABA being the inhibitory neurotransmitter released from basket cells.     

Effects of x-irradiation on the levels of glutamate,aspartate and GABA in different regions of the cerebellum of the rat

The effect of the x-irradiation-induced loss of cerebellar granule and stellate cells on the levels of glutamate, aspartate and GABA in regions of the rat cerebellum was determined. The level of glutamate was significantly lower in the neuron-depleted cerebellar cortex while GABA levels were higher than control values in the cerebellar cortex and white matter of the x-irradiated rats. Aspartate levels were not changed by x-irradiation in any cerebellar region. The data is discussed in terms of the proposed role of glutamate as the excitatory neurotransmitter released from granule cells.     

The identification of dimethylphenylarsine as a microbial metabolite using a simple method of chemofocusing

Candida humicola acts on benzenearsonic acid to produce dimethylphenylarsine, which was identified by mass spectroscopy following the chemofocusing of the volatile metabolite onto a mercuric chloride impregnated filter. The same technique established that trimethylarsine is the volatile metabolic product obtained from C. humicola treated with 4-NH₂-2-OHC₆H₃AsO(OH)₂ and (CH₃)₃AsO. Arsanilic acid, 4-NH₂C₆H₄AsO(OH)₂, is not metabolized to a volatile arsine.     

Methylation of FrzCD, a methyl-accepting taxis protein of Myxococcus xanthus, is correlated with factors affecting cell behavior.

Myxococcus xanthus, a nonflagellated gliding bacterium, exhibits multicellular behavior during vegetative growth and fruiting body formation. The frizzy (frz) genes are required to control directed motility for these interactions. The frz genes encode proteins that are homologous to all of the major enteric chemotaxis proteins, with the exception of CheZ. In this study, we characterized FrzCD, a protein which is homologous to the methyl-accepting chemotaxis proteins from the enteric bacteria. FrzCD, unlike the other methyl-accepting chemotaxis proteins, was found to be localized primarily in the cytoplasmic fraction of cells. FrzCD migrates as a ladder of bands on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, reflecting heterogeneity due to methylation or demethylation and to deamidation. FrzCD was shown to be methylated in vivo when cells were exposed to yeast extract or Casitone and demethylated when starved in buffer. We used the methylation state of FrzCD as revealed by Western blot (immunoblot) analyses to search for stimuli that are recognized by the frz signal transduction system. Common amino acids, nucleotides, vitamins, and sugars were not recognized, but certain lipids and alcohols were recognized. For example, the saturated fatty acids capric acid and lauric acid stimulated FrzCD methylation, whereas a variety of other saturated fatty acids did not. Lauryl alcohol and lipoic acid also stimulated methylation, as did phospholipids containing lauric acid. In contrast, several short-chain alcohols, such as isoamyl alcohol, and some other solvents caused demethylation. The relatively high concentrations of the chemicals required for a response may indicate that these chemicals are not the relevant signals recognized by M. xanthus in nature. Isoamyl alcohol and isopropanol also had profound effects on the behavior of wild-type cells, causing them to reverse continuously. Cells of frzB, frzF, and frzG mutants also reversed continuously in the presence of isoamyl alcohol, whereas cells of frzA, frzCD, or frzE mutants did not. On the basis of the data presented, we propose a model for the frz signal transduction pathway in M. xanthus.     

Organization, structure, and polymorphisms of the human profilaggrin gene

Profilaggrin is a major protein component of the keratohyalin granules of mammalian epidermis. It is initially expressed as a large polyprotein precursor and is subsequently proteolytically processed into individual functional filaggrin molecules. We have isolated genomic DNA and cDNA clones encoding the 5'- and 3'-ends of the human gene and mRNA. The data reveal the presence of likely "CAT" and "TATA" sequences, an intron in the 5'-untranslated region, and several potential regulatory sequences. While all repeats are of the same length (972 bp, 324 amino acids), sequences display considerable variation (10-15%) between repeats on the same clone and between different clones. Most variations are attributable to single-base changes, but many also involve changes in charge. Thus, human filaggrin consists of a heterogeneous population of molecules of different sizes, charges, and sequences. However, amino acid sequences encoding the amino and carboxyl termini are more conserved, as are the 5' and 3' DNA sequences flanking the coding portions of the gene. The presence of unique restriction enzyme sites in these conserved flanking sequences has enabled calculations on the size of the full-length gene and the numbers of repeats in it: depending on the source of genomic DNA, the gene contains 10, 11, or 12 filaggrin repeats that segregate in kindred families by normal Mendelian genetic mechanisms. This means that the human profilaggrin gene system is also polymorphic with respect to size due to simple allelic differences between different individuals. The amino- and carboxyl-terminal sequences of profilaggrin contain partial or truncated repeats with unusual un-filaggrin-like sequences on the termini.(ABSTRACT TRUNCATED AT 250 WORDS)     

A locus for familial hypertrophic cardiomyopathy is closely linked to the cardiac myosin heavy chain genes, CRI-L436, and CRI-L329 on chromosome 14 at q11-q12

We report that a gene responsible for familial hypertrophic cardiomyopathy (HC) is closely linked to the cardiac alpha and beta myosin heavy chain (MHC) genes on chromosome 14q11. We have recently shown that probe CRI-L436, derived from the anonymous DNA locus D14S26, detects a polymorphic restriction fragment that segregates with familial HC in affected members of a large Canadian family. Using chromosomal in situ hybridization, we have mapped CRI-L436 to chromosome 14 at q11-q12. Because the cardiac MHC genes also map to this chromosomal band, we have determined the genetic distances between the cardiac beta MHC gene, D14S26, and the familial HC locus. Data presented here show that these three loci are linked within 5 centimorgans on chromosome 14 at q11-q12. The possibility that defects in either the cardiac alpha or beta MHC genes are responsible for familial HC is discussed.     

Localization of human variable and constant region immunoglobulin heavy chain genes on subtelomeric band q32 of chromosome 14

Analysis of a group of human/rodent somatic cell hybrids with nucleic acid probes prepared from cloned human variable region (VH), junctional (JH), and constant region (C epsilon) heavy chain immunoglobulin genes indicates that all of these IgH genes are localized on the subtelomeric (q32) band of chromosome 14. Somatic cell hybrids were isolated in selective medium after fusing human fibroblasts with hprt- Chinese hamster cells. The human parental cells contained two translocation chromosomes representing a reciprocal translocation between chromosomes X and 14. Only those hybrid cell lines retaining a complete human autosome 14 or the X/14 translocation chromosome (i.e. containing band 14q32) retained the human IgH genes. Retention of these genes did not correlate with the presence of the other translocation chromosome, 14/X. These results indicate that all human IgH genes (VH, JH, and CH) map to the same chromosomal band (14q32) which is commonly involved in reciprocal translocations with human chromosome 8 (8q24) in B-cell neoplasms.