Homer Binds TRPC Family Channels and Is Required for Gating of TRPC1 by IP 3 Receptors (original) (raw)
Related papers
Journal of Biological Chemistry, 2006
The mechanism of TRP channel translocation in response to store depletion and agonist stimulation is not known. Here we use TRPC3 as a model to show that IP 3 and the scaffold Homer 1 (H1) regulate the rate of translocation and retrieval of TRPC3 from the PM. In resting cells, TRPC3 exists in TRPC3-H1b/c-IP 3 Rs complexes that are located in part at the PM and in part in intracellular vesicles. Binding of IP 3 to the IP 3 Rs dissociates the interaction between IP 3 Rs and H1 but not between H1 and TRPC3 to form IP 3 Rs-TRPC3-H1b/c. TIRFM and biotinylation assays show robust receptor-and store-dependent translocation of the TRPC3 to the PM and their retrieval upon termination of cell stimulation. The translocation requires depletion of stored Ca 2؉ and is prevented by inhibition of the IP 3 Rs. In HEK293, dissociating the H1b/c-IP 3 R complex with H1a results in TRPC3 translocation to the PM, where it is spontaneously active. The TRPC3-H1b/c-IP 3 Rs complex is reconstituted by infusing H1c into these cells. Reconstitution is inhibited by IP 3 . Deletion of H1 in mice markedly reduces the rates of translocation and retrieval of TRPC3. Conversely, infusion of H1c into H1 ؊/؊ cells eliminates spontaneous channel activity and increases the rate of channel activation by agonist stimulation. The effects of H1c are inhibited by IP 3 . These findings together with our earlier studies demonstrating gating of TRPC3 by IP 3 Rs were used to develop a model in which assembly of the TRPC3-H1b/c-IP 3 Rs complexes by H1b/c mediates both the translocation of TRPC3containing vesicles to the PM and gating of TRPC3 by IP 3 Rs.
Cellular localization of TRPC3 channel in rat brain: preferential distribution to oligodendrocytes
Neuroscience Letters, 2004
In the present work we describe the cellular localization of TRPC3 in non-excitable cells as compared to the neurons in normal rat brain. We performed a double labeling study for TRPC3 and one of the following cell-specific markers: mouse anti-glial fibrillary acidic protein (GFAP) for astrocytes; mouse anti-RIP for oligodendrocytes, or mouse anti-OX42 for microglia, or mouse anti-NeUN for neuronal nuclei or mouse anti-tyrosine hydroxylase (TH) for detection of dopaminergic neurons of the substantia nigra. Our double label immunofluorescence study showed that that TRPC3 is mainly localized in oligodendrocytes. These result were confirmed by the electron microscopy study, which showed TRPC3 immunoreactivity in oligodendrocytes. Consistent with the evidence that calcium homeostasis is important to oligodendrocytes for development, myelination, and demyelination [Microsc. Res. Tech. 52 (2001) 672], we can speculate that the distribution of TRPC3 in oligodendrocytes plays a role in myelination and or demyelination processes.
Physiological Function and Characterization of TRPCs in Neurons
Cells, 2014
Ca 2+ entry is essential for regulating vital physiological functions in all neuronal cells. Although neurons are engaged in multiple modes of Ca 2+ entry that regulates variety of neuronal functions, we will only discuss a subset of specialized Ca 2+ -permeable non-selective Transient Receptor Potential Canonical (TRPC) channels and summarize their physiological and pathological role in these excitable cells. Depletion of endoplasmic reticulum (ER) Ca 2+ stores, due to G-protein coupled receptor activation, has been shown to activate TRPC channels in both excitable and non-excitable cells. While all seven members of TRPC channels are predominately expressed in neuronal cells, the ion channel properties, mode of activation, and their physiological responses are quite distinct. Moreover, many of these TRPC channels have also been suggested to be associated with neuronal development, proliferation and differentiation. In addition, TRPCs also regulate neurosecretion, long-term potentiation and synaptic plasticity. Similarly, perturbations in Ca 2+ entry via the TRPC channels have been also suggested in a spectrum of neuropathological conditions. Hence, understanding the precise involvement of TRPCs in neuronal function and in neurodegenerative conditions would presumably unveil avenues for plausible therapeutic interventions for these devastating neuronal diseases.
The TRPC2 channel forms protein-protein interactions with Homer and RTP in the rat vomeronasal organ
BMC neuroscience, 2010
Background: The signal transduction cascade operational in the vomeronasal organ (VNO) of the olfactory system detects odorants important for prey localization, mating, and social recognition. While the protein machinery transducing these external cues has been individually well characterized, little attention has been paid to the role of protein-protein interactions among these molecules. Development of an in vitro expression system for the transient receptor potential 2 channel (TRPC2), which establishes the first electrical signal in the pheromone transduction pathway, led to the discovery of two protein partners that couple with the channel in the native VNO.
N-linked protein glycosylation is a major determinant for basal TRPC3 and TRPC6 channel activity
Journal of Biological …, 2003
The TRPC family of receptor-activated cation channels (TRPC channels) can be subdivided into four subfamilies based on sequence homology as well as functional similarities. Members of the TRPC3/6/7 subfamily share common biophysical characteristics and are activated by diacylglycerol in a membrane-delimited manner. At present, it is only poorly understood whether members of the TRPC3/6/7 subfamily are functionally redundant or whether they serve distinct cellular roles. By electrophysiological and fluorescence imaging strategies we show that TRPC3 displays considerable constitutive activity, while TRPC6 is a tightly regulated channel. To identify potential molecular correlates accounting for the functional difference, we analyzed the glycosylation pattern of TRPC6 compared with TRPC3. Two NX(S/T) motifs in TRPC6 were mutated (Asn to Gln) by in vitro mutagenesis to delete one or both extracellular N-linked glycosylation sites. Immunoblotting analysis of HEK 293 cell lysates expressing TRPC6 wild type and mutants favors a model of TRPC6 that is dually glycosylated within the first (e1) and second extracelluar loop (e2) as opposed to the monoglycosylated TRPC3 channel (Vannier, B., Zhu, X., Brown, D., and Birnbaumer, L. (1998) J. Biol. Chem. 273, 8675-8679). Elimination of the e2 glycosylation site, missing in the monoglycosylated TRPC3, was sufficient to convert the tightly receptor-regulated TRPC6 into a constitutively active channel, displaying functional characteristics of TRPC3. Reciprocally, engineering of an additional second glycosylated site in TRPC3 to mimic the glycosylation status in TRPC6 markedly reduced TRPC3 basal activity. We conclude that the glycosylation pattern plays a pivotal role for the tight regulation of TRPC6 through phospholipase C-activating receptors.
Corticolimbic Expression of TRPC4 and TRPC5 Channels in the Rodent Brain
PLoS One, 2007
The canonical transient receptor potential (TRPC) channels are a family of non-selective cation channels that are activated by increases in intracellular Ca 2+ and G q /phospholipase C-coupled receptors. We used quantitative real-time PCR, in situ hybridization, immunoblots and patch-clamp recording from several brain regions to examine the expression of the predominant TRPC channels in the rodent brain. Quantitative real-time PCR of the seven TRPC channels in the rodent brain revealed that TRPC4 and TRPC5 channels were the predominant TRPC subtypes in the adult rat brain. In situ hybridization histochemistry and immunoblotting further resolved a dense corticolimbic expression of the TRPC4 and TRPC5 channels. Total protein expression of HIP TRPC4 and 5 proteins increased throughout development and peaked late in adulthood (6-9 weeks). In adults, TRPC4 expression was high throughout the frontal cortex, lateral septum (LS), pyramidal cell layer of the hippocampus (HIP), dentate gyrus (DG), and ventral subiculum (vSUB). TRPC5 was highly expressed in the frontal cortex, pyramidal cell layer of the HIP, DG, and hypothalamus. Detailed examination of frontal cortical layer mRNA expression indicated TRPC4 mRNA is distributed throughout layers 2-6 of the prefrontal cortex (PFC), motor cortex (MCx), and somatosensory cortex (SCx). TRPC5 mRNA expression was concentrated specifically in the deep layers 5/6 and superficial layers 2/3 of the PFC and anterior cingulate. Patch-clamp recording indicated a strong metabotropic glutamate-activated cation current-mediated depolarization that was dependent on intracellular Ca 2+ and inhibited by protein kinase C in brain regions associated with dense TRPC4 or 5 expression and absent in regions lacking TRPC4 and 5 expression. Overall, the dense corticolimbic expression pattern suggests that these Gq/PLC coupled nonselective cation channels may be involved in learning, memory, and goal-directed behaviors.
Immunohistochemical study on the distribution of TRPC channels in the rat hippocampus
Brain Research, 2006
Objective: To identify glycogen synthase kinase (GSK) 3a expression in a mouse model of familial amyotrophic lateral sclerosis (ALS), we investigated the changes of GSK3a in the central nervous system of SOD1 G93A transgenic mice by immunohistochemistry. Methods: We used 12 SOD1 G93A transgenic and ten wild-type (wt) SOD1 transgenic mice bred by 'The Jackson Laboratory' under the strain designations B6SJL-TgN (SOD1 G93A ) 1 Gur/J and B6SJL-TgN (SOD1) 2 Gur/J, respectively. Immunohistochemistry was performed in accordance with the free-floating method described earlier.
Distribution of TRPC1 and TRPC5 in medial temporal lobe structures of mice
Cell and Tissue Research, 2005
The transient receptor potential (TRP) superfamily comprises a group of non-selective cation channels that have been implicated in both receptor and store-operated channel functions. The family of the classical TRPs (TRPCs) consists of seven members (TRPC1-7). The presence of TRPC1 and TRPC5 mRNA in the brain has previously been demonstrated by real-time polymerase chain reaction. However, the distribution of these receptors within different brain areas of mice has not been investigated in detail. We have used antibodies directed against TRPC1 and TRPC5 to study the distribution and localization of these channels in murine medial temporal lobe structures. Both TRPC1 and TRPC5 channels are present in the various nuclei of the amygdala, in the hippocampus, and in the subiculum and the entorhinal cortex. We have found that TRPC1 channels are primarily expressed on cell somata and on dendrites, whereas TRPC5 channels are exclusively located on cell bodies. Moreover, TRPC1 channels are selectively expressed by neurons, whereas TRPC5 channels are mainly expressed by neurons, but also by non-neuronal cells. The expression of TRPC1 and TRPC5 channels in mammalian temporal lobe structures suggests their involvement in neuronal plasticity, learning and memory.
Group I metabotropic glutamate receptor-dependent TRPC channel trafficking in hippocampal neurons
Journal of Neurochemistry, 2007
The group I metabotropic glutamate receptor agonist (S)-3,5dihydroxyphenylglycine (DHPG) elicited two phases of synchronized neuronal (epileptiform) discharges in hippocampal slices: an initial phase of short duration discharges followed by a phase of prolonged discharges. We assessed the involvement of transient receptor potential canonical (TRPC) channels in these responses. Pre-treatment of hippocampal slices with TRPC channel blockers, 1-[beta-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenethyl]-1H-imidazole hydrochloride (SKF96365) or 2-aminoethoxydiphenyl borate, did not affect the short epileptiform discharges but blocked the prolonged epileptiform discharges. SKF96365 suppressed ongoing DHPG-induced prolonged epileptiform discharges. Western blot analysis showed that the total TRPC4 or TRPC5 proteins in hippocampal slices were unchanged following DHPG. DHPG increased TRPC4 and TRPC5 in the cyto-plasmic compartment and decreased these proteins in the plasma membrane. Translocation of TRPC4 and TRPC5 was suppressed when the epileptiform discharges were blocked by ionotropic glutamate receptor blockers. Translocation of TRPC4 and TRPC5 was also prevented in slices from phospholipase C (PLC) b1 knockout mice, even when synchronized discharges were elicited by the convulsant 4-aminopyridine. The results suggest that TRPC channels are involved in generating DHPG-induced prolonged epileptiform discharges. This function of TRPC channels is associated with a neuronal activity-and PLCb1-dependent translocation of TRPC4 and TRPC5 proteins from the plasmalemma to the cytoplasmic compartment. acid; DHPG, (S)-3,5-dihydroxyphenylglycine; ERK1/2, extracellular signal-regulated kinases 1 and 2; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HRP, horseradish peroxidase; iGluR, ionotropic glutamate receptor; IP 3 , inositol 1,4,5-trisphosphate; mGluR, metabotropic glutamate receptor; PLC, phospholipase C; SDS, sodium dodecyl sulfate; SKF96365, 1-[beta-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenethyl]-1H-imidazole hydrochloride; TBST, Trisbuffered saline-Tween 20; TRP, transient receptor potential; TRPC, transient receptor potential canonical.