1,721,025 research outputs found
Phosphorylated cAMP response element binding protein in the mouse brain after fear conditioning: relationship to Fos production
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The use of multiple ion chromatograms in on-line HPLC-MS for the characterization of post-translational and chemical modifications of proteins
No full textCharacterization of peptide-protein interactions using photoaffinity labeling and LC/MS
No full textThe combination of photoaffinity labeling (PAL) with modern mass spectrometric techniques is a powerful approach for the characterization of peptide-protein interactions. Depending on the analytical strategy applied, a PAL experiment can provide different levels of information ranging from the identification of interaction partners to the structural characterization of ligand-binding sites. On the basis of LC/MS data generated in the framework of the identification of the binding site of the neuropeptide corticotropin-releasing factor (CRF) on its binding protein (CRFBP), the key role of LC/MS in the characterization of photoadducts on different structural levels was demonstrated. Covalent photoadducts of rat CRFBP (rCRFBP) were obtained by PAL with different mono- and bifunctional benzophenone photoprobes designed on the basis of the sequence of the synthetic CRF fragment human/rat CRF(6-33) which binds to CRFBP with high affinity. In view of the stoichiometry, LC/MS analysis revealed that the photoadducts consisted of one molecule of photoprobe and one molecule of rCRFBP. For a further characterization of the photoadducts on the oligopeptide level, enzymatic digests of unlabeled rCRFBP and of the respective photoadduct were compared by peptide mapping monitored with LC/MS. Thereby, it was found that the photoprobe that contained the photophore at its N-terminus labeled the amino acid sequence rCRFBP(34-38), whereas the photoprobe that contained the photophore at its C-terminus labeled rCRFBP(12-26). On the basis of the characterization of the photoadduct formed by rCRFBP and the bifunctional photoprobe that contained photophores on both termini, semiquantitative comparison of different enzymatic digests was accomplished by application of the mass-selective multiple ion chromatogram strategy
Phosphorylation of hippocampal Erk-1/2, Elk-1, and p90-Rsk-1 during contextual fear conditioning: Interactions between Erk-1/2 and Elk-1
No full textThe phosphorylation of proteins involved in the MAP kinase signal transduction pathway was investigated during associative learning of C57BL/6J mice. Context-dependent fear conditioning, consisting of a single exposure of mice to a context followed by foot shock, was employed as a learning paradigm. Control groups consisted of mice exposed to context only or an immediate shock in the context. Coincident up-regulation of phosphorylated Erk-1/2 and Elk-1 was observed in the CA3 hippocampal subfield and dentate gyrus 30 min after fear conditioning but not after the control paradigms. Phosphorylated Erk-1/2 and Elk-1 were associated and predominantly colocalized in the mossy fibers. In vitro kinase assays showed that hippocampal Erk-1/2 phosphorylates Elk-1. Notably, Elk-1 in turn enhances the phosphorylation of Erk-1/2 and its downstream target p90Rsk-1. Increased phosphorylation and nuclear translocation of p90Rsk-1 was also demonstrated in the CA3 hippocampal area in vivo during contextual fear conditioning. The observed interactions between hippocampal Elk-1 and Erk-1/2 proteins may affect the consolidation of contextual memories through activation of the downstream nuclear targets of Erk-1/2, such as p90Rsk-1, without requiring nuclear translocation of Elk-1 and Erk-1/2
Modulation of Hypothalamic NMDA Receptor Function by Cyclic AMP-Dependent Protein Kinase and Phosphatases
No full textDifferential responsiveness of CRF receptor subtypes to N-terminal truncation of peptidic ligands
No full textIdentification of Two Corticotropin-Releasing Factor Receptors from Xenopus laevis with High Ligand Selectivity: Unusual Pharmacology of the Type 1 Receptor
No full textCdk5 in the Adult Non-Demented Brain
No full textCyclin-dependent kinase 5 (Cdk5), a Ser / Thr kinase, regulates the phosphorylation of neuronal proteins and thereby influences neuronal morphology, migration and axon growth. Tightly coordinated interactions between Cdk5 and its activator proteins p35 and p39 are critical for the developmental processes of postmitotic neurons as well as functioning of the adult CNS. Excessive up-regulation of Cdk5 activity leading to hyperphosphorylation of cytoskeletal proteins has been linked to neurodegenerative disorders, such as Alzheimers disease (AD). On this basis it was proposed that Cdk5 might be a promising drug target. The physiologic role of Cdk5 in the adult CNS has been addressed recently. It was demonstrated that Cdk5 is involved in striatal and hippocampal neuronal plasticity and long-term behavioral changes associated with these processes. On the basis of the newly identified role of Cdk5 in synaptic plasticity, learning and memory the view that Cdk5 represents a good drug target in AD accompanied by cognitive dysfunctions may have to be revisited. Alternatively, targeting the mechanisms up-stream of Cdk5 leading to deregulation of Cdk5 activity, such as proteolytic cleavage of its activating subunits may prove to be more beneficial as a therapeutical approach
Distinct roles of hippocampal de novo protein synthesis and actin rearrangement in extinction of contextual fear
No full textIt is believed that de novo protein synthesis is fundamentally linked to synaptic changes in neuronal circuits involved in acquisition and extinction of conditioned responses. Recent studies show that neuronal plasticity may be also altered by cytoskeletal rearrangement independently of protein synthesis. We investigated the role of these processes in the hippocampus during acquisition and extinction of context-dependent conditioned fear in mice. Intrahippocampal injections of the protein synthesis inhibitors anisomycin and puromycin, or of the actin rearrangement inhibitors cytochalasin D and latrunculin A, prevented the acquisition of context-dependent fear. Unexpectedly, anisomycin and puromycin enhanced extinction without erasing the fear memory. In contrast, cytochalasin D and latrunculin A prevented extinction of context-dependent freezing. On the basis of these findings, it is suggested that certain hippocampal mechanisms mediating extinction of conditioned contextual fear are inhibited by protein synthesis and involve actin rearrangement. Such mechanisms might predominantly elicit modifications of hippocampal circuits that store the conditioning memory
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