ABSTRACT
Although addiction develops in a considerable number of regular cocaine users, molecular risk factors for cocaine dependence are still unknown. It was proposed that establishing drug use and memory formation might share molecular and anatomical pathways. Alpha-Ca(2+)/calmodulin-dependent protein kinase-II (αCaMKII) is a key mediator of learning and memory also involved in drug-related plasticity. The autophosphorylation of αCaMKII was shown to accelerate learning. Thus, we investigated the role of αCaMKII autophosphorylation in the time course of establishing cocaine use-related behavior in mice. We found that αCaMKII autophosphorylation-deficient αCaMKII(T286A) mice show delayed establishment of conditioned place preference, but no changes in acute behavioral activation, sensitization or conditioned hyperlocomotion to cocaine (20 mg kg(-1), intraperitoneal). In vivo microdialysis revealed that αCaMKII(T286A) mice have blunted dopamine (DA) and blocked serotonin (5-HT) responses in the nucleus accumbens (NAcc) and prefrontal cortex after acute cocaine administration (20 mg kg(-1), intraperitoneal), whereas noradrenaline responses were preserved. Under cocaine, the attenuated DA and 5-HT activation in αCaMKII(T286A) mice was followed by impaired c-Fos activation in the NAcc. To translate the rodent findings to human conditions, several CAMK2A gene polymorphisms were tested regarding their risk for a fast establishment of cocaine dependence in two independent samples of regular cocaine users from Brazil (n=688) and Switzerland (n=141). A meta-analysis across both samples confirmed that CAMK2A rs3776823 TT-allele carriers display a faster transition to severe cocaine use than C-allele carriers. Together, these data suggest that αCaMKII controls the speed for the establishment of cocaine's reinforcing effects.
Subject(s)
Behavior, Addictive/genetics , Calcium-Calmodulin-Dependent Protein Kinase Type 2/genetics , Cocaine-Related Disorders/genetics , Cocaine/genetics , Reinforcement, Psychology , Adult , Animals , Behavior, Animal/drug effects , Female , Humans , Male , MiceABSTRACT
A high-throughput screening assay for atom transfer catalysis has been developed. This assay is based on two probes, developed herein, which generate highly fluorescent products upon carbene or oxygen atom transfer. The emission wavelength of probes 1 and 5 shift significantly (up to 90 nm) upon epoxidation, allowing detection of product at 3% conversion. Probe 7 is not fluorescent, while fluorescence emission by carbene insertion/rearrangement product 8 allows detection at less than 1% conversion. Such sensitivity allows for examination of single-bead reactions in a high throughput array format (1536 wells per plate), and provides a broad detection window ranging from single to high turnover numbers. Thousands of metal complexes are evaluated in a single screening experiment. Preliminary screening of a diverse ligand library with probe 7 in the presence of Rh(II) uncovered new catalysts capable of cyclopropanation and C-H insertion.
ABSTRACT
RNA polymerase was isolated from Streptomyces granaticolor and protein kinase was partially purified from Streptomyces albus. When RNA polymerase was treated with protein kinase in vitro the activity of RNA polymerase was markedly enhanced. Furthermore, a protein of M = 65 kDa was isolated which, after being phosphorylated, stimulated RNA polymerase activity in vitro. Because neither the beta-subunits nor the alpha-subunits of RNA polymerase were phosphorylated it is assumed that phosphorylation of the 65 kDa protein may regulate the activity of RNA polymerase in streptomycetes.
Subject(s)
DNA-Directed RNA Polymerases/metabolism , Protein Kinases/physiology , Streptomyces/enzymology , Phosphorylation , Streptomyces/genetics , Transcription, Genetic/physiologyABSTRACT
Phosphorylation of synaptic plasma brain membranes (SPM) causes a decrease of the specific binding of demethylated tricyclic antidepressants (TCA) and changes the affinity of 3H-imipramine and 3H-desmethylimipramine binding. The decrease of TCA binding was found also in lymphocyte membranes. In platelet membranes a decreased binding was found only with demethylated dibenzazepine derivatives. Bmax and Kd values are also decreased in the presence of phosphatidic acid or alpha-glycerolphosphate.
Subject(s)
Antidepressive Agents, Tricyclic/metabolism , Synaptic Membranes/metabolism , Animals , Cattle , Dealkylation , Desipramine/metabolism , Dibenzazepines/metabolism , Humans , Imipramine/analogs & derivatives , Imipramine/metabolism , In Vitro Techniques , Kinetics , Ligands , PhosphorylationABSTRACT
In several reactions catalyzed by highly purified peptide elongation factor 1 from rabbit reticulocytes, GTP may be fully replaced by CTP but not by ATP or UTP. This holds true for the factor-dependent binding of aminoacyl-tRNA to ribosomes, GTPase activity, GTP-dependent autophosphorylation of the factor protein and binding of cholesteryl 14-methylhexadecanoate by the factor.
