Homer 1 Mediates Store- and Inositol 1,4,5-Trisphosphate Receptor-dependent Translocation and Retrieval of TRPC3 to the Plasma Membrane

Abstract

Ca2ϩ influx is a critical component of the receptor-evoked Ca 2ϩ signal and plays a role in many physiological functions (1). The best described form of Ca 2ϩ influx is mediated by the store-operated Ca 2ϩ channels (SOCs), 3 which are activated by agonist-dependent or agonist-independent depletion of Ca 2ϩ stored in the ER (1). The molecular identity of the SOCs and I crac is still not known with certainty, although recent work points to ORAI1/CRACM1/olf186-F as a potential I crac (2-5). However, accumulating evidence indicates that members of the transient receptor potential (TRP) family of ion channels are associated with SOCs in mammalian cells. Thus, deletion of TRPC4 in mice (6, 7) or of TRPC1, TRPC3, TRPC6, and TRPC7 by antisense or siRNA (8 -10) and dominant negative TRPC1, TRPC3, partially inhibit SOCs and/or receptor-stimulated Ca 2ϩ influx. The mechanism by which agonist stimulation activates Ca 2ϩ influx by TRPC channels is not well understood. TRPC1, -4, and -5 can be activated by store depletion, whereas TRPC3, -6, and -7 can be activated by the lipid diacylglycerol (11,15,16). However, depending on cell type and expression levels, TRPC3 can also be activated by store depletion (17)(18)(19). Several mechanisms have been proposed to explain how store depletion leads to activation of SOCs and TRPC channels; conformational coupling between TRPC channels and IP 3 receptors (IP 3 Rs) (18, 20 -22), exocytotic insertion of the channels in the plasma membrane (PM) (23)(24)(25), and activation by a diffusible messenger (26,27). Biochemical and functional evidence showed regulatory interaction between IP 3 Rs and several TRPC channels, including TRPC1 and TRPC3 (18,[28][29][30][31][32] Rs (30,32).A newly discovered and apparently a general regulatory mechanism of TRPC channel activity is agonist-stimulated translocation of the channels to the PM. In HEK293 cells, receptor stimulation but not passive store depletion was reported to stimulate the translocation of TRPC3 to the PM in a mechanism that was inhibited by cleavage of VAMP2 (vesicleassociated membrane protein 2) with tetanus toxin (38). Another form of regulation of TRPC3 is by interaction with phospholipase C␥ (22, 39). However, unlike the role of VAMP2, phospholipase C␥ does not affect the acute expression or translocation of TRPC3 but rather the steady-state level of TRPC3 in the PM (22). Stimulation of the epidermal growth factor receptor resulted in translocation of TRPC5 to the PM in a mechanism that was dependent on phosphoinositide 3-kinase, the Rho GTPase Rac1, and phosphatidylinositol-4-phosphate 5-kinase (PIP(5)K␣) (24). Finally, stimulation of the muscarinic M3 receptor resulted in translocation of TRPC6 to the PM in a time course that coincides with activation of Ca 2ϩ influx (25). For the most part, TRPC channel translocation has been studied in cell lines. Whether such a mechanism also operates in native cells is not known. Furthermore, TRPC3, -5, and -6 bind Homers and IP 3 Rs (33) (present work). The potential role of Homer and IP 3 R...

Homer proteins in Ca2+ signaling by excitable and non-excitable cells

“…In addition to the mGluRs, many Ca 2+ signaling proteins express Homer ligands and bind Homer. Among them are the scaffolding protein Shank [8], PLCβ [16,17], IP 3 receptors (IP 3 Rs) [15,18], TRPC channels [15,18], ryanodine receptors (RyRs) [19][20][21][22] and selective L-type Ca 2+ channel isoforms [23][24][25].…”

“…It should be possible to resolve this issue with the availability of mice from which single and combination of the Homers are deleted. The knockout mice were instrumental in discovering the acceleration of RGS proteins GAP activity by Homer2 [30] and the role of Homer1 in conformational coupling and excitation-contraction coupling (see below and [15,18]). These studies showed that H1a facilitates, while H1b/c impedes Ca 2+ signaling by regulating communication between Ca 2+ release channels at the ER/SR and Ca 2+ influx channels at the plasma membrane.…”

“…Receptor-stimulated translocation of TRPC1 was reported to be dependent on the small GTP-binding protein RhoA, [48], which is a regulator of vesicle trafficking and exocytosis. Similarly, stimulation of the M3 receptor results in translocation of TRPC3 [18,49] and requires the function of the SNARE protein VAMP2 [49]. Another study reported translocation of TRPC3 in response to stimulation of the EGF receptor but failed to observe translocation of TRPC3 in response to M3 receptor stimulation [50].…”

“…Binding of IP 3 to the IP 3 Rs dissociates the interaction between IP 3 Rs and Homer1 but not between Homer1 and TRPC3 to form IP 3 Rs-TRPC3-H1b/c complexes. Dissociation of the complexes results in robust translocation of the TRPC3 to the plasma membrane and their retrieval upon termination of cell stimulation [18]. Analysis of TRPC3 translocation in WT and Homer1 −/− pancreatic acinar cells showed that Homer1 regulates the rate of translocation and retrieval of TRPC3 channels from the plasma membrane.…”

“…Fig. 3B illustrates a possible mechanism by which the assembly of the TRPC3-H1b/c-IP3Rs complexes by H1b/c mediates both the translocation of TRPC3-containing vesicles to the plasma membrane and gating of TRPC3 by IP 3 Rs [18].…”