16 research outputs found

    Acyl-Protein Thioesterase 2 Catalizes the Deacylation of Peripheral Membrane-Associated GAP-43

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    An acylation/deacylation cycle is necessary to maintain the steady-state subcellular distribution and biological activity of S-acylated peripheral proteins. Despite the progress that has been made in identifying and characterizing palmitoyltransferases (PATs), much less is known about the thioesterases involved in protein deacylation. In this work, we investigated the deacylation of growth-associated protein-43 (GAP-43), a dually acylated protein at cysteine residues 3 and 4. Using fluorescent fusion constructs, we measured in vivo the rate of deacylation of GAP-43 and its single acylated mutants in Chinese hamster ovary (CHO)-K1 and human HeLa cells. Biochemical and live cell imaging experiments demonstrated that single acylated mutants were completely deacylated with similar kinetic in both cell types. By RT-PCR we observed that acyl-protein thioesterase 1 (APT-1), the only bona fide thioesterase shown to mediate deacylation in vivo, is expressed in HeLa cells, but not in CHO-K1 cells. However, APT-1 overexpression neither increased the deacylation rate of single acylated GAP-43 nor affected the steady-state subcellular distribution of dually acylated GAP-43 both in CHO-K1 and HeLa cells, indicating that GAP-43 deacylation is not mediated by APT-1. Accordingly, we performed a bioinformatic search to identify putative candidates with acyl-protein thioesterase activity. Among several candidates, we found that APT-2 is expressed both in CHO-K1 and HeLa cells and its overexpression increased the deacylation rate of single acylated GAP-43 and affected the steady-state localization of diacylated GAP-43 and H-Ras. Thus, the results demonstrate that APT-2 is the protein thioesterase involved in the acylation/deacylation cycle operating in GAP-43 subcellular distribution

    Diagnostic tools in Rhinology EAACI position paper

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    This EAACI Task Force document aims at providing the readers with a comprehensive and complete overview of the currently available tools for diagnosis of nasal and sino-nasal disease. We have tried to logically order the different important issues related to history taking, clinical examination and additional investigative tools for evaluation of the severity of sinonasal disease into a consensus document. A panel of European experts in the field of Rhinology has contributed to this consensus document on Diagnostic Tools in Rhinology

    Hippocampal protein kinase C signaling mediates the short-term memory impairment induced by delta9-tetrahydrocannabinol

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    Cannabis affects cognitive performance through the activation of the endocannabinoid system, and the molecular mechanisms involved in this process are poorly understood. Using the novel object-recognition memory test in mice, we found that the main psychoactive component of cannabis, delta9-tetrahydrocannabinol (THC), alters short-term object-recognition memory specifically involving protein kinase C (PKC)-dependent signaling. Indeed, the systemic or intra-hippocampal pre-treatment with the PKC inhibitors, prevented the short-term, but not the long-term, memory impairment induced by THC. In contrast, systemic pre-treatment with mammalian target of rapamycin complex 1 (mTORC1) inhibitors, known to block the amnesic-like effects of THC on long-term memory, did not modify such a short-term cognitive deficit. Immunoblot analysis revealed a transient increase in PKC signaling activity in the hippocampus after THC treatment. Thus, THC administration induced the phosphorylation of a specific Ser residue in the hydrophobic-motif at the C-terminal tail of several PKC isoforms. This significant immunoreactive band that paralleled cognitive performance did not match in size with the major PKC isoforms expressed in the hippocampus except for PKCθ. Moreover, THC transiently enhanced the phosphorylation of the postsynaptic calmodulin-binding protein neurogranin in a PKC dependent manner. These data demonstrate that THC alters short-term object-recognition memory through hippocampal PKC/neurogranin signaling.Predoctoral fellowship from the Spanish Ministry of Education (A. B-G. and E.P.). Partial support by FP7-PEOPLE-2013-IEF-623638 (A.B-G). Partial support by FRAXA Research Foundation (M.G-G.). Predoctoral fellowship by FPI (MINEICO/FEDER, EU) (V.S-M. and L.G-L). Research supported by BFU2015-68568P (A.O.), (MINECO/FEDER, UE); Redes temáticas de investigación cooperativa en salud (RETICS) del Instituto de Salud Carlos III (#RD16/0017/0020)-Red de trastornos adictivos (RTA) (R.M.); Generalitat de Catalunya, AGAUR (#2014-SGR-1547) (R.M.); ICREA (Institució Catalana de Recerca i Estudis Avançats) Acadèmia (R.M. and A.O.
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