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1.
Protein and Molecular Characterization of Hippocampal Protein Kinase C in C57BL/6 and DBA/2 Mice 总被引:3,自引:1,他引:2
Barbara J. Bowers Steven C. Christensen James R. Pauley Richard Paylor Lisa Yuva Scott E. Dunbar † Jeanne M. Wehner 《Journal of neurochemistry》1995,64(6):2737-2746
Abstract: Measures of protein kinase C (PKC) in C57BL/6 and DBA/2 mice using [3 H]phorbol 12,13-dibutyrate binding to tissue homogenates and brain slices demonstrated that levels of activated, membrane-bound PKC were greater in C57BL hippocampus than in DBA hippocampus. Western analysis of α-, βI -, βII -, γ-, δ-, and ɛ-PKC using isozyme-specific antibodies indicated that the increase observed in C57BL hippocampus was due primarily to the γ-PKC protein, whose immunoreactivity was greater in the membrane-bound fraction in C57BL mice. Characterization of α-, βI,II -, and γ-PKC hippocampal mRNA using northern analysis and isozyme-specific nucleic acid probes did not reveal differences between the strains in levels of gene expression. Restriction fragment length polymorphisms (RFLP) were found in the α- and γ-, but not β-PKC genomic DNA. The RFLPs appeared to be located in noncoding, nonregulatory regions of the gene. These findings suggest that the γ-PKC isozyme is largely responsible for the PKC activity difference in C57BL and DBA hippocampus that has been reported previously and may be closely associated with differences in learning ability observed in these strains. 相似文献
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Austin L. Hughes 《Journal of molecular evolution》1996,43(1):4-10
The immunoglobulin-related chains of cell-surface receptors for the Fc region of immunoglobulins (FCERIα, FcγRI, FcγRII, and
FcγRIIIα) are encoded by members of a gene family. Phylogenetic analysis of representative members of this family from mammals
revealed that FcγRIIIα genes of human, mouse, and rat are not orthologous to one another in the region of the gene encoding
the Immunoglobulin C2-set domains. In phylogenetic trees of this region, FcγRIIIα and FcγRII clustered together. However,
in trees based on both coding and noncoding regions 5′ and 3′ to the C2 domains, FcγRIIIα genes of human, mouse, and rat clustered
together. This pattern of relationship is most easily explained as a result of two independent recombinational events occurring
in the mouse and rat after these two species diverged, in each of which the exons encoding the C2 domains were donated to
an FcγRIIIα gene by an FcγRII gene. 相似文献
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Abstract: Polyclonal antibodies were raised to the C-terminal part of the γ-aminobutyric acidA (GABAA ) receptor α4-subunit. These anti-peptide α4 (517–523) antibodies specifically identified a protein with apparent molecular mass 67 kDa in rat brain membranes. This protein was enriched by immunoaffinity chromatography of brain membrane extracts on Affigel 10 coupled to the anti-peptide α4 (517–523) antibodies and could then be identified by the anti-α4-antibodies as well as by the GABAA receptor subunit-specific monoclonal antibody bd-28. This appears to indicate that the 67-kDa protein is the α4-subunit of GABAA receptors. Intact GABAA receptors appeared to be retained by the immunoaffinity column because other GABAA receptor subunit proteins like the β2/β3-subunits and the γ2-subunit were detected in the immunoaffinity column eluate. Furthermore, in addition to the 67-kDa protein, a 51-kDa protein could be detected by the antibody bd-28 and the anti-peptide α4 (517–523) antibody in the immunoaffinity column eluate. A protein with similar apparent molecular mass was identified by the α1-subunit-specific anti-peptide α1 (1–9) antibody. In contrast to the α1-subunit, the 51-kDa protein identified by the anti-α4 antibody could not be deglycosylated by N -Glycanase. The identity of the 51-kDa protein identified by the anti-α4-antibodies thus must be further investigated. 相似文献
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† E. R. Korpi T. Kuner †P. Kristo M. Köhler A. Herb H. Lüddens P. H. Seeburg 《Journal of neurochemistry》1994,63(3):1167-1170
Abstract: Sequence variation was found in cDNA coding for the extracellular domain of the rat γ-aminobutyric acid type A (GABAA ) receptor α6 subunit. About 20% of polymerase chain reaction (PCR)-amplified α6 cDNA prepared from rat cerebellar mRNA lacked nucleotides 226–255 as estimated by counting single-stranded phage plaques hybridized specifically to the short (α6S) and long (wild-type) forms of the α6 mRNA. Genomic PCR revealed an intron located upstream of the 30-nucleotide sequence. Both splice forms were detected in the cerebellum by in situ hybridization. Recombinant receptors, resulting from coexpression of the α6S subunit with the GABAA receptor β2 and γ2 subunits in human embryonic kidney 293 cells, were inactive at binding [3 H]muscimol and [3 H]Ro 15-4513. In agreement, injection of complementary RNAs encoding the same subunits into Xenopus oocytes produced only weak GABA-induced currents, indistinguishable from those produced by β2γ2 receptors. Therefore, the 10 amino acids encoded by the 30-nucleotide fragment may be essential for the correct assembly or folding of the α6 subunit-containing receptors. 相似文献
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Latitudinal variation for two enzyme loci and an inversion polymorphism in Drosophila melanogaster from Central and South America 总被引:2,自引:0,他引:2
Van 't Land J Van Putten WF Villarroel H Kamping A Van Delden W 《Evolution; international journal of organic evolution》2000,54(1):201-209
Abstract.— Many organisms show latitudinal variation for various genetically determined traits. Such clines may involve neutral variation and originate from historical events or their maintenance may be explained by selection. For Drosophila melanogaster , latitudinal variation for allozymes, inversions, and quantitative traits has been found on several continents. We sampled D. melanogaster populations in Panama and along a transect of 40 latitudinal degrees on the west coast of South America. Negative correlations with latitude were found for Adhs and α GpdhF allele frequencies and for the frequency of the cosmopolitan inversion In(2L)t in Adhs αpdhF chromosomes. A positive correlation existed between wing length and latitude. Significant correlations were found between these traits and climatic variables like temperature and rainfall. The observed clines show considerable resemblance to those found on other continents. Gametic disequilibrium between Adhs and αGpdhF occurred predominantly at higher latitudes and was caused by the presence of In(2L)t . The reasons for the clinal distributions are discussed and it is argued that selection is the most likely explanation. However, the exact nature of the selective force and the interactions of allozymes with each other and with In(2L)t are complex and not fully understood. In tropical regions In(2L)t -containing genotypes have higher fitness than ST/ST and Adh and αGpdh hitchhike with the inversion, but there is also evidence for balancing selection at the Adh locus. 相似文献
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SUSUMU OHNO 《Differentiation; research in biological diversity》1981,18(1-3):65-74
The means employed for immunoglobulin heavy chain class switch appears to be no different from that by which meiotic intergenic crossing-overs are accomplished. As with other intergenic spacers, the 5' noncoding sequence of each Ig CH (immunoglobulin heavy chain constant region) gene apparently undergoes unconstrained sequence changes due to randomly sustained base substitutions, deletions, and duplications. Yet, there remains sufficient regional sequence homology between the Ig Cμ 5' noncoding sequence and those of its somatic recombination partners, e.g., Ig Cγ1 , Ig Cγ2b , Ig Cα , because each of these 5' noncoding sequences is made of multiple copies in various stages of degeneracy of one primordial 20 base pair-long sequence: (AGCTG) (AGCTG) (AGCTG) (GGGTG). 相似文献
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Abstract: The aim of this study was to evaluate the effect of citalopram, a second generation antidepressant agent producing no β-down-regulation, on the receptors and second messenger systems related to noradrenergic transmission in the cerebral cortex of the rat. We confirmed that citalopram does not bind to α1 -, α2 -, and β2 -adrenoceptors, but we found that it attenuates the inhibitory action of the protein kinase C activator, 12- O -tetradecanoylphorbol 13-acetate, on the noradrenergic response from α1 -adrenoceptor. In contrast to most antidepressants, chronic treatment with citalopram does not produce β-down-regulation, but increases the responses to noradrenaline from β-adrenoceptors without increasing the β1 ,-adrenoceptor density. Chronic treatment with citalopram also increases the maximal response from α1 -adrenoceptor. The results indicate that β-down-regulation is not a necessary characteristic of an efficient antidepressant drug. 相似文献
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Determination of Brain Enolase Isozymes with an Enzyme Immunoassay at the Level of Single Neurons 总被引:19,自引:16,他引:3
Abstract Ultrasensitive enzyme immunoassay systems for the assay of rat brain enolase isozymes ( αα , αγ , and γγ forms) were prepared by use of β- d -galactosidase from Escherichia coli as label and the purified rabbit antibodies to αα and γγ enolases. The antibodies were purified from the immunoglobulin G (IgG) fractions of antisera by immunoaffinity chromatography with a column of the corresponding antigen-coupled Sepharose. Sandwich-type immunoassay systems with the galactosidase-labeled antibody Fab'fragments and the antibody F(abapos;)2 -immobilized polystyrene beads could determine amounts as small as 1 amol (10−18 mol) of each isozyme. Purkinje cell bodies picked up from the bulk-separated fraction by means of a nylon loop were subjected to the assay at the level of single cells. In contrast to previous report, this neuron contained not only the γγ but also the αγ and αα enolases at a level of amol per cell body, although the concentration of γγ was the highest. Immunohistochemical experiments on the cerebellum with the peroxidase-labeled antirabbit IgG antibody and the unlabeled antibody method confirmed the above results, and indicated that both α and γ subunits of the enolase were stained intensely in axons. 相似文献
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Role of the Conserved Lysine Residue in the Middle of the Predicted
Extracellular Loop Between M2 and M3 in the GABAA Receptor 总被引:2,自引:1,他引:1
Abstract : In α1, β2, and γ2 subunits of the γ-aminobutyric acid A (GABAA ) receptor, a conserved lysine residue occupies the position in the middle of the predicted extracellular loop between the transmembrane M2 and M3 regions. In all three subunits, this residue was mutated to alanine. Whereas the mutation in α1 and β2 subunits results each in about a sixfold shift of the concentration-response curve for GABA to higher concentrations, no significant effect by mutation in the γ subunit was detected. The affinity for the competitive inhibitor bicuculline methiodide was not affected by the mutations in either the α1 subunit or the β2 subunit. Concentration-response curves for channel activation by pentobarbital were also shifted to higher concentrations by the mutation in the α and β subunits. Binding of [3 H]Ro 15-1788 was unaffected by the mutation in the α subunit, whereas the binding of [3 H]muscimol was shifted to lower affinity. Mutation of the residue in the α1 subunit to E, Q, or R resulted in an about eight-, 10-, or fivefold shift, respectively, to higher concentrations of the concentration-response curve for GABA. From these observations, it is concluded that the corresponding residues on the α1 and β2 subunits are involved more likely in the gating of the channel by GABA than in the binding of GABA or benzodiazepines. 相似文献
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Virginie Redeker Filippo Rusconi Jean Mary Danielle Promé Jean Rossier 《Journal of neurochemistry》1996,67(5):2104-2114
Abstract: A combination of posttranslational modifications contributes to the high heterogeneity of brain tubulin in mammals. In this report, the structures of the detyrosinated carboxy-terminal peptides of α-tubulin from newborn and adult mouse brain were compared. The heterogeneity of these carboxy-terminal peptides was observed to increase from newborn to adult brain tubulin. The major part of this increased heterogeneity is due to the posttranslational excision of Glu450 , which makes α-tubulin nontyrosinatable (Δ-2 tubulin). The structures of the polyglutamyl side chain of the bi- and triglutamylated peptides were analyzed in this work. In polyglutamylation of α-tubulin, the first glutamyl residue can only be amide-linked to the γ-carboxyl group of Glu445 , but the additional residues may be linked either to the γ- or to the α-carboxyl groups of the preceding one. By optimized reverse-phase separations and comparison with synthetic peptides corresponding to all possible linkages for the biglutamylated (γ1α2, γ1γ2) and triglutamylated (γ1α2α3, γ1γ2γ3, γ1α2γ3, γ1γ2α3, γ1γ2α2) tubulin peptides, it was possible to conclude that the mode of linkage connecting the second and third additional glutamyl residues corresponds mostly to α-bond structures, for both newborn and adult mice. 相似文献
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Abstract: The multisubunit γ-aminobutyric acid type A (GABAA ) receptor is heterogeneous in molecular and pharmacological aspects. We used quantitative autoradiographic techniques to generate detailed pharmacological profiles for the binding of the GABAA -receptor ionophore ligand tert -[35 S]butylbicyclophosphorothionate ([35 S]TBPS) and its modulation by GABA and the GABAA antagonists bicuculline and 2'-(3'-carboxy-2',3'-propyl)-3-amino-6- p -methoxyphenylpyrazinium bromide (SR 95531). Regional differences in the actions of bicuculline and SR 95531 were correlated with the expression of 13 GABAA subunits in brain as reported previously. In some brain regions SR 95531 reduced [35 S]TBPS binding much more than bicuculline, as illustrated by high ratios of bicuculline- to SR 95531-modulated [35 S]TBPS binding. This ratio correlated positively with α2-subunit mRNA levels. Binding that was equally affected by SR 95531 and bicuculline occurred prominently in regions with abundant α1 mRNA expression. The present findings thus reveal a novel pharmacological heterogeneity based on differences between α1 and α2 subunit-containing GABAA receptors. The data aid in developing GABAA -receptor subtype-specific antagonists and in establishing receptor domains critical for the actions of GABAA antagonists. 相似文献
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Ela Balic Uwe Rudolph† Jean-Marc Fritschy Hanns Mohler Dietmar Benke 《Journal of neurochemistry》2009,110(1):244-254
GABAA receptors are pentameric ligand-gated ion channels that are major mediators of fast inhibitory neurotransmission. Clinically relevant GABAA receptor subtypes are assembled from α5(1-3, 5), β1-3 and the γ2 subunit. They exhibit a stoichiometry of two α, two β and one γ subunit, with two GABA binding sites located at the α/β and one benzodiazepine binding site located at the α/γ subunit interface. Introduction of the H105R point mutation into the α5 subunit, to render α5 subunit-containing receptors insensitive to the clinically important benzodiazepine site agonist diazepam, unexpectedly resulted in a reduced level of α5 subunit protein in α5(H105R) mice. In this study, we show that the α5(H105R) mutation did not affect cell surface expression and targeting of the receptors or their assembly into macromolecular receptor complexes but resulted in a severe reduction of α5-selective ligand binding. Immunoprecipitation studies suggest that the diminished α5-selective binding is presumably due to a repositioning of the α5(H105R) subunit in GABAA receptor complexes containing two different α subunits. These findings imply an important role of histidine 105 in determining the position of the α5 subunit within the receptor complex by determining the affinity for assembly with the γ2 subunit. 相似文献
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Clostridium putrefaciens grew well in most media used routinely for culturing anaerobes, but produced spores only on lactose-egg yolk agar. The D80° was 8–14 min, z value was 4°–6° and D γ , 0.16 Mrad. The inhibitory interactions of pH, NaCl, NaNO2 and incubation temperature are described. 相似文献
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GABAA Receptors Containing the α4-Subunit: Prevalence, Distribution, Pharmacology, and Subunit Architecture In Situ 总被引:1,自引:0,他引:1
Abstract: Recombinant GABAA receptors, expressed from α-, β-, and γ2-subunits, are diazepam-insensitive when the α-subunit is either α4 or α6. In situ, diazepam-insensitive receptors containing the α6-subunit are almost exclusively expressed in the granule cell layer of the cerebellum. However, diazepam-insensitive receptors are also expressed in forebrain areas. Here, we report on the presence of diazepam-insensitive GABAA receptors in various brain areas containing the α4-subunit. GABAA receptors immunoprecipitated with a newly developed α4-subunit-specific antiserum displayed a drug binding profile that was indistinguishable from those of α4β2γ2-recombinant receptors and diazepam-insensitive [3 H]Ro 15-4513 binding sites in rat brain membranes. In addition, α4-subunit containing receptors and forebrain diazepam-insensitive receptors are present at comparably low abundance in rat brain and exhibit virtually identical patterns of distribution. Analysis of the subunit architecture of α4-subunit containing receptors revealed that the α4-subunit contributes to several receptor subtypes. Depending on the brain region, the α4-subunit can be coassembled with a second type of α4-subunit variant being α1, α2, or α3. The data demonstrate that native receptors containing the α4-subunit are structurally heterogeneous, expressed at very low abundance in the brain, and display the drug binding profile of diazepam-insensitive [3 H]Ro 15-4513 binding sites. Pharmacologically, these receptors may contribute to the actions of nonclassical ligands such as Ro 15-4513 and bretazenil. 相似文献
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Abstract: The pentameric subunit composition of a large population (36%) of the cerebellar granule cell GABAA receptors that show diazepam (or clonazepam)-insensitive [3 H]Ro 15-4513 binding has been determined by immunoprecipitation with subunit-specific antibodies. These receptors have α6 , α1 , γ2S , γ2L , and β2 or β3 subunits colocalizing in the same receptor complex. 相似文献