Direct binding of α‐actinin enhances TRPP3 channel activity

Abstract

Transient receptor potential (TRP) polycystin 2 and 3 (TRPP2 and 3) are homologous members of the TRP superfamily of cation channels but have different physiological functions. TRPP2 is part of a flow sensor, and is defective in autosomal dominant polycystic kidney disease and implicated in left-right asymmetry development. TRPP3 is reported to implicate in sour tasting in bipolar cells of taste buds of the tongue and in the regulation of pH-sensitive action potential in neurons surrounding the central canal of spinal cord. TRPP3 is present in both excitable and non-excitable cells in various tissues, such as retina, brain, heart, testis, and kidney, but its common and cell type-specific functional characteristics remain largely unknown. In this study, we investigated physical and functional interactions between TRPP3 and a-actinin, an actinbundling protein known to regulate several types of ion channels. We employed planer lipid bilayer electrophysiology system to study the function of TRPP3 channel that was affinity-purified from Madin-Darby canine kidney cells. Upon reconstitution in bilayer, TRPP3 exhibited cation channel activities that were substantially augmented by a-actinin. The TRPP3-a-actinin association was documented by coimmunoprecipitation using native cells and tissues, yeast two-hybrid, and in vitro binding assays. Further, TRPP3 was abundantly present in mouse brain where it associates with aactinin-2. Taken together, a-actinin not only attaches TRPP3 to the cytoskeleton but also up-regulates TRPP3 channel function. It remains to be determined whether the TRPP3-aactinin interaction is relevant to acid sensing and other functions in neuronal and non-neuronal cells. Keywords: acid sensing, lipid bilayer electrophysiology, protein-protein interaction, tandem affinity purification, transient receptor potential polycystin 3, a-actinin.

Regulation of the murine TRPP3 channel by voltage, pH, and changes in cell volume

“…It has been reported that human TRPP3 forms a nonselective cation channel when it is expressed in Xenopus oocytes [1][2][3]14], planar lipid bilayers [13], or mammalian cells [25]. The human TRPP3 channel is considered to be Ca 2+ -activated [1], although no functional consequence of this Ca 2+ regulation has been reported [25].…”

“…It has been shown that TRPP3 channels are voltage-dependent [13] and inhibited by monovalent cations larger than tetraethyl ammonium [2] as well as by multivalent cations such as Mg 2+ , Gd 3+ , and La 3+ [1,14]. In the case of murine TRPP3, however, neither the single-channel conductance nor the exact gating properties have been investigated so far.…”

Receptor for Activated C Kinase 1 (RACK1) Inhibits Function of Transient Receptor Potential (TRP)-type Channel Pkd2L1 through Physical Interaction

“…It is well known that RACK1 is abundantly expressed in brain and interacts with brain proteins such as NMDA and GABA receptors (38,39,49). On the other hand, Pkd2L1 was also recently found in several brain compartments such as hippocampus, thalamus, and spinal cord, although its physiological roles there remain unclear (10,15). We thus tested their interaction in mouse brain tissues by co-IP.…”

“…We also reported that Pkd2L1 physically interacts with ␣-actinin, an important component of the actin filament, in brain and other tissues and is functionally stimulated by ␣-actinin in vitro (10). Although overexpressed Pkd2L1 is targeted to the plasma membrane (PM) of Xenopus oocytes, it mostly localizes in intracellular membranes of mammalian cells when expressed alone (10 -12).…”

Regulation of TRPP3 Channel Function by N-terminal Domain Palmitoylation and Phosphorylation

“…In Xenopus laevis oocytes, TRPP3 targets to the plasma membrane when expressed alone and acts as a Ca 2ϩ -activated channel permeable to Na ϩ , K ϩ , and Ca 2ϩ (15,16). In human embryonic kidney (HEK) cell lines, TRPP3 mainly targets to the endoplasmic reticulum membrane when expressed alone but traffics to the plasma membrane when co-expressed with PKD1 or PKD1L3, a homologue of PKD1 (9,17,18). TRPP3 alone, or together with PKD1L3, mediates pH-dependent cation conductance in an off-response manner, i.e.…”