Photosynthetic electron transport interaction of xanthorrhizol isolated from Iostephane heterophylla and its derivatives

A bioactivity-guided chemical study of Iostephane heterophylla (Asteraceae) led to the isolation of xanthorrhizol (1) as the compound that causes inhibition of ATP synthesis, H⁺-uptake and electron flow from water to methylviologen (basal, phosphorylating and uncoupled) in freshly lysed spinach chloroplasts, thus acting as an inhibitor of the Hill reaction. Acetyl (2), dihydro (3) and acetyl-dihydro (4) derivatives were synthesized. It was found that 4 was less active than 1 and 2 in ATP synthesis, whereas 3 was the most potent inhibitor of the Hill reaction and was also an inhibitor of H⁺-ATPase. Studies of the photosynthetic partial redox reactions from PQ to MV indicated that 1 partially inhibited the PQ pool, but that 3 did not. However, both inhibited the uncoupled electron transport in PSII from water to DCBQ. Uncoupled electron flow from water to silicomolybdate was completely inhibited by 3 and partially by 1. The reaction from DPC to DCPIP was inhibited by both 1 and 3. These results indicate that the inhibition site is located within PSII for 1 and 3 as was corroborated by fluorescence decay data.     

The γ subunits of the native GABAA/benzodiazepine receptors

Subunit-specific antibodies to all the γ subunit isoforms described in mammalian brain (γ₁, γ_2S, γ_L, and γ₃) have been made. The proportion of GABA_A receptors containing each γ subunit isoform in various brain regions has been determined by quantitative immunoprecipitation. In all tested regions of the rat brain, the γ₁, and γ₃ subunits are present in considerable smaller proportion of GABA_A receptor than the γ₂ subunit. Immunocytochemistry shows that γ₁ immunoreactivity concentrates in the stratum oriens and stratum radiatum of the CA1 region of the hippocampus. In the dentate gyrus, γ₁ immunoreactivity concentrates on the outer 2/3 of the molecular layer coinciding with the localization of the axospinous synapses of the perforant pathway. In contrast, γ₃ immunoreactivity concentrates on the basket cells and other GABAergic local circuit neurons of the hilus. These cells are also rich in γ_2S. In the cerebellu, γ₁ immunolabeling was localized on the Bergmann glia. The γ_2S and γ_2L subunits are differentially expressed in various brain regions. Thus the γ_2S is highly expressed in the olfactory bulb and hippocampus whereas the γ_2L is very abundant in inferior colliculus and cerebellum, particularly in Purkinje cells, as immunocytochemistry, in situ hybridization and immunoprecipitation techniques have revealed. The γ_2S and γ_2L coexist in some brain areas and cell types. Moreover, the γ_2S and γ_2L subunits can coexist in the same GABA_A receptor pentamer. We have shown that this is the case in some GABA_A receptors expressed in cerebellar granule cells. These GABA_A receptors also have α and β subunits forming the pentamer. Immunoblots have shown that the rat γ₁, γ_2S, γ_2L and γ₃ subunits are peptides of 47, 45, 47 and 44 kDa respectively. Results also indicate that there are aging-related changes in the expression of the γ_2S and γ_2L subunits in various brain regions which suggest the existence of aging-related changes in the subunit composition of the GABA_A receptors which in turn might lead to changes in receptor pharmacology. The results obtained with the various γ subunit isoforms are discussed in terms of the high molecular and binding heterogeneity of the native GABA_A receptors in brain. Special issue dedicated to Dr. Kinya Kuriyama     

The Subunit Composition of a GABAA/Benzodiazepine Receptor from Rat Cerebellum

Abstract: The pentameric subunit composition of a large population (36%) of the cerebellar granule cell GABA_A receptors that show diazepam (or clonazepam)-insensitive [³H]Ro 15-4513 binding has been determined by immunoprecipitation with subunit-specific antibodies. These receptors have α₆, α₁, γ_2S, γ_2L, and β₂ or β₃ subunits colocalizing in the same receptor complex.     

Big flies, small repeats: the "Thr-Gly" region of the period gene in Diptera

The region of the clock gene period (per) that encodes a repetitive tract of threonine-glycine (Thr-Gly) pairs has been compared between Dipteran species both within and outside the Drosophilidae. All the non- Drosophilidae sequences in this region are short and present a remarkably stable picture compared to the Drosophilidae, in which the region is much larger and extremely variable, both in size and composition. The accelerated evolution in the repetitive region of the Drosophilidae appears to be mainly due to an expansion of two ancestral repeats, one encoding a Thr-Gly dipeptide and the other a pentapeptide rich in serine, glycine, and asparagine or threonine. In some drosophilids the expansion involves a duplication of the pentapeptide sequence, but in Drosophila pseudoobscura both the dipeptide and the pentapeptide repeats are present in larger numbers. In the nondrosophilids, however, the pentapeptide sequence is represented by one copy and the dipeptide by two copies. These observations fulfill some of the predictions of recent theoretical models that have simulated the evolution of repetitive sequences.      

Do reproductive characteristics of the species explain differences in the investment in weapon size present in males?

Sexually selected traits, such as male weapons, are highly variable in shape and size across species. However, little is known about the mechanisms that may govern this variation. Because males with greater investment in weapon size tend to win more fights, but also pay higher costs related to weapon development and maintenance, larger weapons should be expected only in species in which victory in male–male fights generate reproductive benefits that outweigh investment costs. Here, we hypothesized that the reproductive characteristics that increase the chances of winners to access females or to fertilize eggs will favor the evolution of larger weapons. To evaluate this hypothesis, we conducted a meta-analysis using arthropods as model organisms. To measure investment in weapon size, we gathered both Pearson correlation coefficient and the standardized (but non centralized) slope values for the relationship between weapon size and body size. We found that none of the reproductive characteristics we investigated was related to male weapon size. Thus, it seems that greater certainty of accessing a female or fertilizing female eggs with a victory does not modulate the investment in male weapon size. Perhaps the cost–benefit ratio between weapon size investment and reproductive success is not the main factor driving the variation in weapon size.     

Genomic Organization and Evolution of Alternative Exons in a Drosophila Calcium Channel Gene

The genomic organization of a gene coding for an α1 subunit of a voltage-gated calcium channel of Drosophila melanogaster (Dmca1A) was determined. Thirty-four exons, distributed over 45 kb of genomic sequence, have been identified and mapped, including exons in three regions involved in alternative splicing and new sites potentially involved in RNA editing. The comparison of the intron/exon boundaries of this channel with a mammalian counterpart shows that the genomic structure of these two genes has remained fairly similar during evolution, with more than half of the Drosophila intron positions being perfectly conserved compared to the human channel. Phylogenetic analysis of the mutually exclusive alternative exons revealed that they have diverged considerably. It is suggested that this divergence, rather than reflecting evolutionary age, is the likely result of accelerated rates of evolution following duplication.     

Linkage disequilibrium, mutational analysis and natural selection in the repetitive region of the clock gene, period, in Drosophila melanogaster

We have used the method of disequilibrium pattern analysis to examine associations between the threonine-glycine (Thr-Gly) encoding repeat region of the clock gene period (per) of Drosophila melanogaster, and polymorphic sites both upstream and downstream of the repeat, in a number of European fly populations. The results are consistent with the view that selection may be operating on various haplotypes which share the Thr-Gly length alleles encoding 17, 20 and 23 dipeptide pairs, and that the repeat itself may be the focus for selection. These conclusions lend support to a number of other population and behavioural investigations which have provided evidence that selection is acting on the Thr-Gly region. The linkage analysis was also used to infer an approximate mutation rate (mu) for the repeat, of 10(-5) < mu < 4 x 10(-5) per gamete per generation. Direct measurements of the mutation rate using the polymerase chain reaction in a pedigree analysis of tens of thousands of individuals do not contradict this value. Consequently, the Thr-Gly repeat does not have a mutation rate that is as high as some of the non-coding minisatellites, but it is several orders of magnitude higher than the nucleotide substitution rate. The implications of this elevated mutation rate for linkage disequilibria and selection are discussed.     

N-Acetylaspartylglutamate Stimulates Metabotropic Glutamate Receptor 3 to Regulate Expression of the GABAAα6 Subunit in Cerebellar Granule Cells

Abstract: We have shown that the vertebrate neuropeptide N-acetylaspartylglutamate (NAAG) meets the criteria for a neurotransmitter, including function as a selective metabotropic glutamate receptor (mGluR) 3 agonist. Short-term treatment of cerebellar granule cells with NAAG (30 µM) results in the transient increase in content of GABA_Aα6 subunit mRNA. Using quantitative PCR, this increase was determined to be up to 170% of control values. Similar effects are seen following treatment with trans-1-aminocyclopentane-1,3-dicarboxylate and glutamate and are blocked by the mGluR antagonists (2S,3S,4S)-2-methyl-2-(carboxycyclopropyl)glycine and (2S)-α-ethylglutamic acid. The effect is pertussis toxin-sensitive. The increase in α6 subunit mRNA level can be simulated by activation of other receptors negatively linked to adenylate cyclase activity, such as adenosine A1, α2-adrenergic, muscarinic, and GABA_B receptors. Forskolin stimulation of cyclic AMP (cAMP) levels abolished the effect of NAAG. The change in α6 levels induced by 30 µM NAAG can be inhibited in a dose-dependent manner by simultaneous application of increasing doses of the β-adrenergic receptor agonist isoproterenol. The increase in α6 mRNA content is followed by a fourfold increase in α6 protein level 6 h posttreatment. Under voltage-clamped conditions, NAAG-treated granule cells demonstrate an increase in the furosemide-induced inhibition of GABA-gated currents in a concentration-dependent manner, indicating an increase in functional α6-containing GABA_A receptors. These data support the hypothesis that NAAG, acting through mGluR3, regulates expression of the GABA_Aα6 subunit via a cAMP-mediated pathway and that cAMP-coupled receptors for other neurotransmitters may similarly influence GABA_A receptor subunit composition.