Specific distribution of gabarap, gec1/gabarap Like 1, gate16/gabarap Like 2, lc3 messenger RNAs in rat brain areas by quantitative real-time PCR
ABSTRACT GABARAP and GEC1/GABARAPL1 interact with tubulin and GABA(A) receptor and belong to a new protein family. This family includes GATE 16 and LC3, potentially involved in intracellular transport processes. In this study, we combined brain dissection and quantitative real-time reverse transcription polymerase chain reaction to study discriminatively gabarap, gec1/gabarapL1, gate16/gabarapL2, lc3 mRNA distribution in multiple rat brain areas.
- SourceAvailable from: Gen Kaneko
[Show abstract] [Hide abstract]
- "Since its discovery, GABARAP and its orthologs have been found in almost all organisms as diverse from yeast to human. Human GABARAPs, which are linker proteins between microtubules and g subunit of GABA A receptors are expressed ubiquitously in the central nervous system, and a few in endocrine organs and other tissues  . Amphioxus GABARAPs are expressed ubiquitously, with the highest expression in the enteric nervous system, notochord, and ovary, although its precise function is unknown  . "
ABSTRACT: γ-Aminobutyric acid receptor type A-associated protein (GABARAP) and its homologs constitute a protein family found in many eukaryotes from yeast to human, and are known to be involved in intracellular membrane trafficking of GABAA receptors and autophagy. In this study, we cloned cDNA-encoding GABARAP from the monogonont rotifer Brachionus plicatilis and examined for its tissue distribution at the protein level in neonates, males and females. Using reverse transcription (RT)-PCR and rapid amplification of cDNA ends (RACE) techniques, we showed that like other GABARAPs, rotifer GABARAP was also composed of 117 amino acids and highly homologous to vertebrate GABARAP2 ortholog (74–76% identity). GABARAP was demonstrated with its specific antibody to be ubiquitously distributed, irrespective of neonates, males, and females, in the coronal area that covers brain and contains most mechano- and chemoreceptors. Rotifer GABARAP was also expressed in the mature eggs but not in immature eggs. Double immunostaining with mammalian anti-GABA γ receptor antibody showed that rotifer GABARAP co-localized with GABA receptor, suggesting the association of the two proteins. The presence of GABARAP in rotifer implies that it is highly conserved during evolution, and plays important roles in various biological processes.International Review of Hydrobiology 03/2014; 99(1-2):188-197. DOI:10.1002/iroh.201301720 · 1.01 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: Fast inhibitory synaptic transmission is predominantly mediated by GABA(A) receptor (GABA(A)R) in the CNS. Although several types of neuronal activity-dependent plasticity at GABAergic synapses have been reported, the detailed mechanism is elusive. Here we show that binding of structurally altered GABA(A)R-associated protein (GABARAP) to GABA(A)R gamma2 subunit and to tubulin is critical for long-term potentiation [called rebound potentiation (RP)] at inhibitory synapses on a cerebellar Purkinje neuron (PN). Either inhibition of GABARAP association with GABA(A)Rgamma2 or deletion of tubulin binding region of GABARAP impaired RP. Inhibition of tubulin polymerization also suppressed RP. Thus, precise regulation of GABA(A)Rgamma2-GABARAP-microtubule interaction is critical for RP. Furthermore, competitive inhibition of GABARAP binding to GABA(A)Rgamma2 after the RP establishment attenuated the potentiated response, suggesting that GABARAP is critical not only for the induction but also for the maintenance of RP. Fluorescence resonance energy transfer analysis revealed that GABARAP underwent sustained structural alteration after brief depolarization of a PN depending on the activity of Ca2+/calmodulin-dependent protein kinase II (CaMKII), which is required for the RP induction. The susceptibility of GABARAP to undergo structural alteration was abolished by an amino acid replacement in GABARAP. Furthermore, RP was impaired by expression of the mutant GABARAP with the replacement. Together, we conclude that GABA(A)R association with structurally altered GABARAP downstream of CaMKII activation is essential for RP.The Journal of Neuroscience : The Official Journal of the Society for Neuroscience 07/2007; 27(25):6788-99. DOI:10.1523/JNEUROSCI.1981-07.2007 · 6.75 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: GEC1 protein shares high identity with GABARAP (GABA(A) Receptor-Associated Protein), interacts with tubulin and GABA(A) receptors and is potentially involved in intracellular transport processes. Recently, using quantitative real time PCR, we have reported the gec1 mRNA expression in different rat brain areas. In the present study, we investigated the cell types expressing gec1 in rat brain. Sense and anti-sense gec1 RNA probes, corresponding to the 3'-untranslated region, were generated. In northern blotting experiments, the anti-sense probe revealed only the 1.75 kb gec1 mRNAs. On the other hand, in immunohistochemistry experiments, GEC1 polyclonal antibodies did not discriminate between GEC1 and GABARAP proteins. Therefore, we used digoxigenin-labeled RNA probes for in situ hybridization (ISH) experiments to map the gec1 expression. Using the anti-sense probe, we detected the gec1 mRNAs specifically in neurons throughout the rostrocaudal extent of the brain as well as in the spinal cord. Although a majority of neurons expressed the gec1 mRNAs, different intensities of labeling were observed depending on the areas: the strongest labeling was observed in the isocortex, hippocampus, basal telencephalon, some thalamic and most of hypothalamic nuclei, cerebellum, and numerous brainstem nuclei. Furthermore, the gec1 mRNAs were intensely expressed in neurons involved in somatomotor and neuroendocrine functions and weakly expressed in sensory and reticular structures. These results corroborate the putative role of the GEC1 protein in the trafficking of receptor GABA(A).Brain Research 06/2008; 1210:103-15. DOI:10.1016/j.brainres.2008.02.077 · 2.83 Impact Factor