A Pathogenic C Terminus-truncated Polycystin-2 Mutant Enhances Receptor-activated Ca2+ Entry via Association with TRPC3 and TRPC7

Abstract

Mutations in PKD2 gene result in autosomal dominant polycystic kidney disease (ADPKD). PKD2 encodes polycystin-2 (TRPP2), which is a homologue of transient receptor potential (TRP) cation channel proteins. Here we identify a novel PKD2 mutation that generates a C-terminal tail-truncated TRPP2 mutant 697fsX with a frameshift resulting in an aberrant 17-amino acid addition after glutamic acid residue 697 from a family showing mild ADPKD symptoms. When recombinantly expressed in HEK293 cells, wild-type (WT) TRPP2 localized at the endoplasmic reticulum (ER) membrane significantly enhanced Ca 2؉ release from the ER upon muscarinic acetylcholine receptor (mAChR) stimulation. In contrast, 697fsX, which showed a predominant plasma membrane localization characteristic of TRPP2 mutants with C terminus deletion, prominently increased mAChR-activated Ca 2؉ influx in cells expressing TRPC3 or TRPC7. Coimmunoprecipitation, pulldown assay, and cross-linking experiments revealed a physical association between 697fsX and TRPC3 or TRPC7. 697fsX but not WT TRPP2 elicited a depolarizing shift of reversal potentials and an enhancement of single-channel conductance indicative of altered ion-permeating pore properties of mAChR-activated currents. Importantly, in kidney epithelial LLC-PK1 cells the recombinant 679fsX construct was codistributed with native TRPC3 proteins at the apical membrane area, but the WT construct was distributed in the basolateral membrane and adjacent intracellular areas. Our results suggest that heteromeric cation channels comprised of the TRPP2 mutant and the TRPC3 or TRPC7 protein induce enhanced receptor-activated Ca 2؉

Vasopressin and disruption of calcium signalling in polycystic kidney disease

“…Studies aimed at elucidating the role of the polycystins in this process and how it is altered in ADPKD are summarized in Table 2. 12,32–34,39,50–65 The results of these studies are not always consistent, likely due to different experimental conditions. For example, reduced IP3-induced calcium release has been observed in experiments where PC1 was either knocked down or overexpressed.…”

“…For example, PC1 interacts with the inositol 1,4,5-trisphosphate receptor (IP3R) and the stromal interaction molecule 1 (STIM1), while PC2 interacts with IP3R, the ryanodine receptor (RyR), TRPV4 and TRPC1, 3, 4 and 7. 32–39 …”

Transient receptor potential channelopathies

“…Recently, a new frameshift mutation, resulting in mild form of ADPKD, has been identified [91]. This mutation leads to the C-terminally truncated TRPP2 channel (E697X) that shows a predominant plasma membrane expression and prominently increased receptor-operated Ca 2+ (ROC) influx in cells expressing TRPC3 or TRPC7.…”

Polycystin-2 Activation by Inositol 1,4,5-Trisphosphate-induced Ca2+ Release Requires Its Direct Association with the Inositol 1,4,5-Trisphosphate Receptor in a Signaling Microdomain

“…Recently Li et al (24) demonstrated also that ER-localized PKD1 can interact with the IP 3 R, thereby inhibiting IP 3 -induced Ca 2ϩ release (IICR). Several studies observed an enhancement of intracellular Ca 2ϩ release that was attributed to an effect of TRPP2 (15,23,25,26), but the physiological mechanism of action was not elucidated. On the other hand, Wegierski and co-workers (27) observed that TRPP2 could also act as a passive Ca 2ϩ -leak channel in the ER, thereby lowering the Ca 2ϩ concentration in the ER ([Ca 2ϩ ] ER ), which resulted in an opposite effect and decreased the magnitude of IICR.…”