Modulation of Ca ²⁺ entry by polypeptides of the inositol 1,4,5-trisphosphate receptor (IP3R) that bind transient receptor potential (TRP): Evidence for roles of TRP and IP3R in store depletion-activated Ca ²⁺ entry

Abstract

Homologues ofS timulation of cells that elevates inositol 1,4,5-trisphosphate (IP3) causes the release of Ca 2ϩ from internal stores and its entry from the external milieu (for reviews see refs. 1-3). The release from internal stores occurs through channels formed by IP3 receptors (IP3Rs), and entry is mediated by a set of functionally heterogeneous but ubiquitous channels that are activated by the store depletion event per se. Transient receptor potential (TRP) proteins have been hypothesized to be structural components of Ca 2ϩ entry channels (4, 5) and to be activated by IP3R in response to IP3 or store depletion. However, neither has the presence of TRP in Ca 2ϩ entry channels been proven nor has the mechanism(s) by which the channels are activated been clearly elucidated. Indeed, the mechanism by which Ca 2ϩ entry channels are activated has received considerable attention, and arguments have been set forth (i) for activation by second messengers or mediators that include cGMP, IP3, diacylglycerol, a G protein, arachidonic acid derivatives, and a complex termed CIF (6-14), (ii) for translocation from internal pools with involvement of an exocytotic event (15, 16), and (iii) for shortrange physical coupling between the membrane delimiting the store and the plasma membrane (17, 18). The short-range physical-coupling model proposed that membrane Ca 2ϩ entry channels may be activated by the same protein that is responsible for store depletion, i.e., the IP3R (for details see ref. 1).The first functional evidence for a direct role of IP3R in Ca 2ϩ entry was obtained by Kiselyov et al. (19), who showed that Ca 2ϩ entry channels found in HEK cells expressing transfected TRP3 in stable form can be activated in inside-out membrane patches by addition of either IP3R-rich cerebellar microsomes or liposomes carrying recombinant IP3R protein truncated at its C terminus to inactivate its channel-forming capacity. However, this study did not determine whether the protein with which IP3R interacted was TRP3. Indeed, TRP was shown to cause changes in protein expression other than TRP, e.g., upregulation of IP3Rs, and attempts to coimmunoprecipitate IP3R and TRP3 failed (19). We now show that IP3R and TRP can be coimmunoprecipitated. We thus sought to identify interacting domains using in vitro protein:protein interaction tests and, if we found them, to test for their function. Such domains were identified and, upon expression in cells whose TRP and IP3R complement had not been manipulated, were found to modulate natural Ca 2ϩ entry stimulated by either a G protein-coupled pathway or store depletion. The data support a model in which store depletion-activated Ca 2ϩ entry is mediated by TRP-based channels that are activated by the IP3R. Materials and Methods

Heterologous expression of pneumococcal virulence factor PspC on the surface of Lactococcus lactis confers adhesive properties

“…4c). 2-APB is a pharmacological substance, commonly used as an InsP3 receptor antagonist (Birnbaumer et al, 2000;Boulay et al, 1999;Wu et al, 2000). These results validated the results of our previous studies conducted with pneumococci.…”

Novel role for the transient receptor potential channel TRPM2 in prostate cancer cell proliferation

“…A similar mechanism could also be present for TRPM2 considering evidence that inositol 1,4,5-trisphosphate receptors (IP 3 R) can migrate into the nucleus and IP 3 R directly binds to TRPC channels. 32,33 TRPM2 is expressed in many non-cancerous cells, such as brain and peripheral blood cells. 7,8,34 Considering the function of TRPM2 in these cells is to serve as a mediator of oxidative-stress-induced calcium influx (i.e., the channel is at plasma membrane) and thus apoptosis in those tissues, inhibition of TRPM2 may be beneficial in terms of continued survival.…”

RhoA Interaction with Inositol 1,4,5-Trisphosphate Receptor and Transient Receptor Potential Channel-1 Regulates Ca2+ Entry

“…Although evidence from several laboratories showed that IP 3 R coupling with TRPC can trigger Ca 2ϩ entry, the mechanisms responsible for this critical association event are not clear (23,25,34,(61)(62)(63)(64). Models involving chemical and conformation coupling have been suggested (1, 11, 14 -16, 65).…”