Formation of a new receptor‐operated channel by heteromeric assembly of TRPP2 and TRPC1 subunits

Abstract

Although several protein-protein interactions have been reported between transient receptor potential (TRP) channels, they are all known to occur exclusively between members of the same group. The only intergroup interaction described so far is that of TRPP2 and TRPC1; however, the significance of this interaction is unknown. Here, we show that TRPP2 and TRPC1 assemble to form a channel with a unique constellation of new and TRPP2/ TRPC1-specific properties. TRPP2/TRPC1 is activated in response to G-protein-coupled receptor activation and shows a pattern of single-channel conductance, amiloride sensitivity and ion permeability distinct from that of TRPP2 or TRPC1 alone. Native TRPP2/TRPC1 activity is shown in kidney cells by complementary gain-of-function and loss-of-function experiments, and its existence under physiological conditions is supported by colocalization at the primary cilium and by co-immunoprecipitation from kidney membranes. Identification of the heteromultimeric TRPP2/TRPC1 channel has implications in mechanosensation and cilium-based Ca 2 þ signalling.

Structural and molecular basis of the assembly of the TRPP2/PKD1 complex

“…This suggests that TRPP2 reaches the surface membrane either on its own or, perhaps, in complex with an unknown endogenous oocyte protein. One such candidate protein is TRPC1, which is endogenously expressed in oocytes (23) and directly associates with TRPP2 (17,21,24). Fluorescence spots from oocytes expressing TRPP2-EGFP alone showed the same EGFP bleaching pattern ( Fig.…”

“…These results suggest that the association of the fourth subunit is significantly weaker than the trimeric assembly and that this association is potentially dynamic and regulated. A tightly bound TRPP2 trimer may associate with other channel-forming subunits, such as TRPC1 (17,21,24) and TRPV4 (37), to form heterotetrameric channels, thereby greatly increasing its functional spectrum and versatility. It is of interest to note that the cyclic nucleotidegated channels of rod photoreceptors also have a 3:1 stoichiometry, with 3 A1 and 1 B1 subunits (38)(39)(40)(41)(42), and that this stoichiometry is determined by a trimer-forming leucine-zipper domain in the C terminus of the A1 subunit (40).…”

“…S3A). Finally, it has been reported that the TRPP2/PKD1 complex can be activated by an anti-PKD1 antibody raised against an epitope (E2939-N2956) in the extracellular N terminus in human PKD1 (20,21). We produced 2 different batches of anti-mouse PKD1 serum raised against the corresponding region (E2931-N2948) in mouse PKD1.…”

“…They also colocalize in the primary cilia of kidney epithelium (14). The 2 proteins associate physically (15)(16)(17)(18)(19) and form functional complexes (18,20,21). However, the subunit composition of this heteromeric complex and the molecular mechanism underlying its assembly are unknown.…”

Structural model of the TRPP2/PKD1 C-terminal coiled-coil complex produced by a combined computational and experimental approach

“…TRPP2 (also known as PKD2 or polycystin-2), a member of the TRPP subfamily (3), coassembles with PKD1 (also known as polycystin-1), an integral membrane protein that presumably functions as a cell surface receptor (4), to form an ion channel/receptor complex (5)(6)(7)(8)(9)(10)(11)(12). Both TRPP2 and PKD1 are widely distributed, with high expression levels in kidney epithelial cells (3,4,10,11), where they appear to colocalize in the primary cilia and may function as mechanosensitive, Ca 2þ -permeable cation channels (5,13).…”