The versatile nature of the calcium-permeable cation channel TRPP2

TRPP2 is a member of the transient receptor potential (TRP) superfamily of cation channels, which is mutated in autosomal dominant polycystic kidney disease (ADPKD). TRPP2 is thought to function with polycystin 1-a large integral protein-as part of a multiprotein complex involved in transducing Ca(2+)-dependent information. TRPP2 has been implicated in various biological functions including cell proliferation, sperm fertilization, mating behaviour, mechanosensation and asymmetric gene expression. Although its function as a Ca(2+)-permeable cation channel is well established, its precise role in the plasma membrane, the endoplasmic reticulum and the cilium is controversial. Recent studies suggest that TRPP2 function is highly dependent on the subcellular compartment of expression, and is regulated by many interactions with adaptor proteins. This review summarizes the most pertinent evidence about the properties of TRPP2 channels, focusing on the compartment-specific functions of mammalian TRPP2.     

Activation of TRPP2 through mDia1-dependent voltage gating

The TRPP2 cation channel is directly responsible for approximately 15% of all cases of autosomal dominant polycystic kidney disease. However, the mechanisms underlying fundamental properties of TRPP2 regulation, such as channel gating and activation, are unknown. We have shown that TRPP2 was activated by EGF and physically interacted with the mammalian diaphanous-related formin 1 (mDia1), a downstream effector of RhoA. Now, we show that mDia1 regulates TRPP2 by specifically blocking its activity at negative but not positive potentials. The voltage-dependent unblock of TRPP2 by mDia1 at positive potentials is mediated through RhoA-induced molecular switching of mDia1 from its autoinhibited state at negative potentials to its activated state at positive potentials. Under physiological resting potentials, EGF activates TRPP2 by releasing the mDia1-dependent block through the activation of RhoA. Our data reveal a new role of mDia1 in the regulation of ion channels and suggest a molecular basis for the voltage-dependent gating of TRP channels.     

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

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 Ca²⁺ 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 Ca²⁺ 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 Ca²⁺ influx that may lead to dysregulated cell growth in ADPKD.     

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

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+) signalling.     

Two members of the TRPP family of ion channels, Pkd1l3 and Pkd2l1, are co-expressed in a subset of taste receptor cells

Taste receptors cells are responsible for detecting a wide variety of chemical stimuli. Several molecules including both G protein coupled receptors and ion channels have been shown to be involved in the detection and transduction of tastants. We report on the expression of two members of the transient receptor potential (TRP) family of ion channels, PKD1L3 and PKD2L1, in taste receptor cells. Both of these channels belong to the larger polycystic kidney disease (PKD or TRPP) subfamily of TRP channels, members of which have been demonstrated to be non-selective cation channels and permeable to both Na(+) and Ca(2+). Pkd1l3 and Pkd2l1 are co-expressed in a select subset of taste receptor cells and therefore may, like other PKD channels, function as a heteromer. We found the taste receptor cells expressing Pkd1l3 and Pkd2l1 to be distinct from those that express components of sweet, bitter and umami signal transduction pathways. These results provide the first evidence for a role of TRPP channels in taste receptor cell function.     

The S4–­­S5 linker – gearbox of TRP channel gating

Transient receptor potential (TRP) channels are cation channels which participate in a wide variety of physiological processes in organisms ranging from fungi to humans. They fulfill roles in body homeostasis, are sensors for noxious chemicals and temperature in the mammalian somatosensory system and are activated by light stimulated phospholipase C activity in Drosophila or by hypertonicity in yeast. The transmembrane topology of TRP channels is similar to that of voltage-gated cation channels. TRP proteins assemble as tetramers with each subunit containing six transmembrane helices (S1–S6) and intracellular N- and C-termini. Here we focus on the emerging functions of the cytosolic S4–S5 linker on TRP channel gating. Most of this knowledge comes from pathogenic mutations within the S4–S5 linker that alter TRP channel activities. This knowledge has stimulated forward genetic approaches to identify additional residues around this region which are essential for channel gating and is supported, in part, by recent structures obtained for TRPV1, TRPV2, TRPV6, TRPA1, and TRPP2.     

TRP channels: an overview

The TRP ("transient receptor potential") family of ion channels now comprises more than 30 cation channels, most of which are permeable for Ca2+, and some also for Mg2+. On the basis of sequence homology, the TRP family can be divided in seven main subfamilies: the TRPC ('Canonical') family, the TRPV ('Vanilloid') family, the TRPM ('Melastatin') family, the TRPP ('Polycystin') family, the TRPML ('Mucolipin') family, the TRPA ('Ankyrin') family, and the TRPN ('NOMPC') family. The cloning and characterization of members of this cation channel family has exploded during recent years, leading to a plethora of data on the roles of TRPs in a variety of tissues and species, including mammals, insects, and yeast. The present review summarizes the most pertinent recent evidence regarding the structural and functional properties of TRP channels, focusing on the regulation and physiology of mammalian TRPs.     

TRPP2 channels regulate apoptosis through the Ca2+ concentration in the endoplasmic reticulum

Ca²⁺ is an important signalling molecule that regulates multiple cellular processes, including apoptosis. Although Ca²⁺ influx through transient receptor potential (TRP) channels in the plasma membrane is known to trigger cell death, the function of intracellular TRP proteins in the regulation of Ca²⁺‐dependent signalling pathways and apoptosis has remained elusive. Here, we show that TRPP2, the ion channel mutated in autosomal dominant polycystic kidney disease (ADPKD), protects cells from apoptosis by lowering the Ca²⁺ concentration in the endoplasmic reticulum (ER). ER‐resident TRPP2 counteracts the activity of the sarcoendoplasmic Ca²⁺ ATPase by increasing the ER Ca²⁺ permeability. This results in diminished cytosolic and mitochondrial Ca²⁺ signals upon stimulation of inositol 1,4,5‐trisphosphate receptors and reduces Ca²⁺ release from the ER in response to apoptotic stimuli. Conversely, knockdown of TRPP2 in renal epithelial cells increases ER Ca²⁺ release and augments sensitivity to apoptosis. Our findings indicate an important function of ER‐resident TRPP2 in the modulation of intracellular Ca²⁺ signalling, and provide a molecular mechanism for the increased apoptosis rates in ADPKD upon loss of TRPP2 channel function.     

瞬时感受器电位通道与疾病

哺乳动物瞬时感受器电位(transient receptor potential,TRP)通道超家族由TRPC、TRPM、TRPV、TRPA、TRPP和TRPML六个亚家族组成。这些亚家族的29个离子通道几乎表达于所有的组织和细胞。大多对单价和二价阳离子都有通透性。TRP通道与多种生物学功能有关,包括高血压、温度觉、血管炎症、刺激感、肿瘤增生、细胞内离子稳态及神经细胞信号转导。对这些通道的生理功能及其与人类疾病的关系的研究有助于开发具有潜在治疗价值的TRP通道调节剂。     

Mechanotransduction by TRP Channels: General Concepts and Specific Role in the Vasculature

Transient receptor potential (TRP) ion channel superfamily is involved in sensing and transmission of a broad variety of external or internal stimuli, including but not limited to mechanical stress. Based on homology analysis, genetic and molecular studies have recently identified TRP channels in different tissues, comprising blood vessels. In invertebrates, many TRP channels including five TRPV channels identified in Caenorhabditis elegans and two in Drosophila have been implicated in mechanosensory behaviors as molecular basis of volume regulation, hearing and touch sensitivity. Consistently, in mammals many TRP family members such as TRPC1, TRPC3, TRPC6, TRPM4, TRPM7, TRPN1, TRPA1, TRPY1, TRPP1, TRPP2, and notably, TRPV1, TPRV2 as well as TRPV4 have been reported to be involved in mechanotransduction. This review summarizes recent and at times controversial findings on the role and regulation of TRP channels in mechanotransduction. Specifically, we highlight the relevance of TRPV channels in vascular regulation and focus on TRPV4 in the vascular system of the lung, which is constantly exposed to a unique combination of circumferential and longitudinal strains. In light of our observation in intact pulmonary microvessels that mechanical stress induced Ca²⁺ signaling in endothelial cells is closely related to TRPV4 activity, we postulate that TRPV4 plays a critical role in lung vascular mechanotransduction. The progress in this rapidly expanding field may allow for the identification of new molecular targets and the development of new therapeutic approaches in a number of intractable diseases related to mechanical stress.     

TRP channels in kidney disease

Mammalian TRP channel proteins form six-transmembrane cation-permeable channels that may be grouped into six subfamilies on the basis of amino acid sequence homology (TRPC, TRPV, TRPM, TRPA, TRPP, and TRPML). Recent studies of TRP channels indicate that they are involved in numerous fundamental cell functions and are considered to play an important role in the pathophysiology of many diseases. Many TRPs are expressed in kidney along different parts of the nephron and growing evidence suggest that these channels are involved in hereditary, as well as acquired kidney disorders. TRPC6, TRPM6, and TRPP2 have been implicated in hereditary focal segmental glomerulosclerosis (FSGS), hypomagnesemia with secondary hypocalcemia (HSH), and polycystic kidney disease (PKD), respectively. In addition, the highly Ca(2+)-selective channel, TRPV5, contributes to several acquired mineral (dys)regulation, such as diabetes mellitus (DM), acid-base disorders, diuretics, immunosuppressant agents, and vitamin D analogues-associated Ca(2+) imbalance whereas TRPV4 may function as an osmoreceptor in kidney and participate in the regulation of sodium and water balance. This review presents an overview of the current knowledge concerning the distribution of TRP channels in kidney and their possible roles in renal physiology and kidney diseases.     

Distinct protein domains regulate ciliary targeting and function of C. elegans PKD-2

TRPP2 (transient receptor potential polycystin-2) channels function in a range of cells where they are localized to specific subcellular regions including the endoplasmic reticulum (ER) and primary cilium. In humans, TRPP2/PC-2 mutations severely compromise kidney function and cause autosomal dominant polycystic kidney disease (ADPKD). The Caenorhabditis elegans TRPP2 homolog, PKD-2, is restricted to the somatodendritic (cell body and dendrite) and ciliary compartments of male specific sensory neurons. Within these neurons PKD-2 function is required for sensation. To understand the mechanisms regulating TRPP2 subcellular distribution and activity, we performed in vivo structure-function-localization studies using C. elegans as a model system. Our data demonstrate that somatodendritic and ciliary targeting requires the transmembrane (TM) region of PKD-2 and that the PKD-2 cytosolic termini regulate subcellular distribution and function. Within neuronal cell bodies, PKD-2 colocalizes with the OSM-9 TRP vanilloid (TRPV) channel, suggesting that these TRPP and TRPV channels may function in a common process. When human TRPP2/PC-2 is heterologously expressed in transgenic C. elegans animals, PC-2 does not visibly localize to cilia but does partially rescue pkd-2 null mutant defects, suggesting that human PC-2 and PKD-2 are functional homologs.     

The transient receptor potential family of ion channels

The transient receptor potential (TRP) multigene superfamily encodes integral membrane proteins that function as ion channels. Members of this family are conserved in yeast, invertebrates and vertebrates. The TRP family is subdivided into seven subfamilies: TRPC (canonical), TRPV (vanilloid), TRPM (melastatin), TRPP (polycystin), TRPML (mucolipin), TRPA (ankyrin) and TRPN (NOMPC-like); the latter is found only in invertebrates and fish. TRP ion channels are widely expressed in many different tissues and cell types, where they are involved in diverse physiological processes, such as sensation of different stimuli or ion homeostasis. Most TRPs are non-selective cation channels, only few are highly Ca²⁺ selective, some are even permeable for highly hydrated Mg²⁺ ions. This channel family shows a variety of gating mechanisms, with modes of activation ranging from ligand binding, voltage and changes in temperature to covalent modifications of nucleophilic residues. Activated TRP channels cause depolarization of the cellular membrane, which in turn activates voltage-dependent ion channels, resulting in a change of intracellular Ca²⁺ concentration; they serve as gatekeeper for transcellular transport of several cations (such as Ca²⁺ and Mg²⁺), and are required for the function of intracellular organelles (such as endosomes and lysosomes). Because of their function as intracellular Ca²⁺ release channels, they have an important regulatory role in cellular organelles. Mutations in several TRP genes have been implicated in diverse pathological states, including neurodegenerative disorders, skeletal dysplasia, kidney disorders and pain, and ongoing research may help find new therapies for treatments of related diseases.     

Molecular and functional characterization of a novel mouse transient receptor potential protein homologue TRP7. Ca(2+)-permeable cation channel that is constitutively activated and enhanced by stimulation of G protein-coupled receptor.

Characterization of mammalian homologues of Drosophila transient receptor potential protein (TRP) is an important clue to understand molecular mechanisms underlying Ca(2+) influx activated in response to stimulation of G(q) protein-coupled receptors in vertebrate cells. Here we have isolated cDNA encoding a novel seventh mammalian TRP homologue, TRP7, from mouse brain. TRP7 showed abundant RNA expression in the heart, lung, and eye and moderate expression in the brain, spleen, and testis. TRP7 recombinantly expressed in human embryonic kidney cells exhibited distinctive functional features, compared with other TRP homologues. Basal influx activity accompanied by reduction in Ca(2+) release from internal stores was characteristic of TRP7-expressing cells but was by far less significant in cells expressing TRP3, which is structurally the closest to TRP7 in the TRP family. TRP7 induced Ca(2+) influx in response to ATP receptor stimulation at ATP concentrations lower than those necessary for activation of TRP3 and for Ca(2+) release from the intracellular store, which suggests that the TRP7 channel is activated independently of Ca(2+) release. In fact, TRP7 expression did not affect capacitative Ca(2+) entry induced by thapsigargin, whereas TRP7 greatly potentiated Mn(2+) influx induced by diacylglycerols without involvement of protein kinase C. Nystatin-perforated and conventional whole-cell patch clamp recordings from TRP7-expressing cells demonstrated the constitutively activated and ATP-enhanced inward cation currents, both of which were initially blocked and then subsequently facilitated by extracellular Ca(2+) at a physiological concentration. Impairment of TRP7 currents by internal perfusion of the Ca(2+) chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid revealed an essential role of intracellular Ca(2+) in activation of TRP7, and their potent activation by the diacylglycerol analogue suggests that the TRP7 channel is a new member of diacylglycerol-activated cation channels. Relative permeabilities indicate that TRP7 is slightly selective to divalent cations. Thus, our findings reveal an interesting correspondence of TRP7 to the background and receptor stimulation-induced cation currents in various native systems.     

TRPP2 ion channels: Critical regulators of organ morphogenesis in health and disease

Ion channels control the membrane potential and mediate transport of ions across membranes. Archetypical physiological functions of ion channels include processes such as regulation of neuronal excitability, muscle contraction, or transepithelial ion transport. In that regard, transient receptor potential ion channel polycystin 2 (TRPP2) is remarkable, because it controls complex morphogenetic processes such as the establishment of properly shaped epithelial tubules and left-right-asymmetry of organs. The fascinating question of how an ion channel regulates morphogenesis has since captivated the attention of scientists in different disciplines. Four loosely connected key insights on different levels of biological complexity ranging from protein to whole organism have framed our understanding of TRPP2 physiology: 1) TRPP2 is a non-selective cation channel; 2) TRPP2 is part of a receptor-ion channel complex; 3) TRPP2 localizes to primary cilia; and 4) TRPP2 is required for organ morphogenesis. In this review, we will discuss the current knowledge in these key areas and highlight some of the challenges ahead.     

Structural insights into group II TRP channels

The seven members of the TRP channel superfamily are divided into two main groups with five members comprising group I (TRPC/V/M/N/A) and TRPML (TRP MucoLipin) and TRPP (TRP Polycystin) making up group II. Group II channels share a high sequence homology on their transmembrane domains and are distinct from group I members as they contain a large luminal/extracellular domain between transmembrane helix 1 (S1) and S2. Since 2016, there are more than ten research papers reporting various structures of group II channels by either cryo-EM or X-ray crystallography. These studies along with recent functional analysis by the other groups have considerably strengthened our knowledge on TRPML and TRPP channels. In this review, we summarize and discuss these reports providing molecular insights into the group II TRP channel family.     

Cloning and functional expression of human short TRP7, a candidate protein for store-operated Ca2+ influx.

The regulation and control of plasma membrane Ca(2+) fluxes is critical for the initiation and maintenance of a variety of signal transduction cascades. Recently, the study of transient receptor potential channels (TRPs) has suggested that these proteins have an important role to play in mediating capacitative calcium entry. In this study, we have isolated a cDNA from human brain that encodes a novel transient receptor potential channel termed human TRP7 (hTRP7). hTRP7 is a member of the short TRP channel family and is 98% homologous to mouse TRP7 (mTRP7). At the mRNA level hTRP7 was widely expressed in tissues of the central nervous system, as well as some peripheral tissues such as pituitary gland and kidney. However, in contrast to mTRP7, which is highly expressed in heart and lung, hTRP7 was undetectable in these tissues. For functional analysis, we heterologously expressed hTRP7 cDNA in an human embryonic kidney cell line. In comparison with untransfected cells depletion of intracellular calcium stores in hTRP7-expressing cells, using either carbachol or thapsigargin, produced a marked increase in the subsequent level of Ca(2+) influx. This increased Ca(2+) entry was blocked by inhibitors of capacitative calcium entry such as La(3+) and Gd(3+). Furthermore, transient transfection of an hTRP7 antisense expression construct into cells expressing hTRP7 eliminated the augmented store-operated Ca(2+) entry. Our findings suggest that hTRP7 is a store-operated calcium channel, a finding in stark contrast to the mouse orthologue, mTRP7, which is reported to enhance Ca(2+) influx independently of store depletion, and suggests that human and mouse TRP7 channels may fulfil different physiological roles.     

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

Autosomal dominant polycystic kidney disease is characterized by the loss-of-function of a signaling complex involving polycystin-1 and polycystin-2 (TRPP2, an ion channel of the TRP superfamily), resulting in a disturbance in intracellular Ca²⁺ signaling. Here, we identified the molecular determinants of the interaction between TRPP2 and the inositol 1,4,5-trisphosphate receptor (IP₃R), an intracellular Ca²⁺ channel in the endoplasmic reticulum. Glutathione S-transferase pulldown experiments combined with mutational analysis led to the identification of an acidic cluster in the C-terminal cytoplasmic tail of TRPP2 and a cluster of positively charged residues in the N-terminal ligand-binding domain of the IP₃R as directly responsible for the interaction. To investigate the functional relevance of TRPP2 in the endoplasmic reticulum, we re-introduced the protein in TRPP2^−/− mouse renal epithelial cells using an adenoviral expression system. The presence of TRPP2 resulted in an increased agonist-induced intracellular Ca²⁺ release in intact cells and IP₃-induced Ca²⁺ release in permeabilized cells. Using pathological mutants of TRPP2, R740X and D509V, and competing peptides, we demonstrated that TRPP2 amplified the Ca²⁺ signal by a local Ca²⁺-induced Ca²⁺-release mechanism, which only occurred in the presence of the TRPP2-IP₃R interaction, and not via altered IP₃R channel activity. Moreover, our results indicate that this interaction was instrumental in the formation of Ca²⁺ microdomains necessary for initiating Ca²⁺-induced Ca²⁺ release. The data strongly suggest that defects in this mechanism may account for the altered Ca²⁺ signaling associated with pathological TRPP2 mutations and therefore contribute to the development of autosomal dominant polycystic kidney disease.     

Structure and Functional Characterization of a Novel Human Low-Voltage Activated Calcium Channel

Abstract : We have isolated and characterized overlapping cDNAs encoding a novel, voltage-gated Ca²⁺ channel α₁ subunit, α_1H, from a human medullary thyroid carcinoma cell line. The α_1H subunit is structurally similar to previously described α₁ subunits. Northern blot analysis indicates that α_1H mRNA is expressed throughout the brain, primarily in the amygdala, caudate nucleus, and putamen, as well as in several nonneuronal tissues, with relatively high levels in the liver, kidney, and heart. Ba²⁺ currents recorded from human embryonic kidney 293 cells transiently expressing α_1H activated at relatively hyperpolarized potentials (-50 mV), rapidly inactivated (τ = 17 ms), and slowly deactivated. Similar results were observed in Xenopus oocytes expressing α_1H. Singlechannel measurements in human embryonic kidney 293 cells revealed a single-channel conductance of ~9 pS. These channels are blocked by Ni²⁺ (IC₅₀ = 6.6 μ M ) and the T-type channel antagonists mibefradil (~50% block at 1 μ M ) and amiloride (IC₅₀ = 167 μ M ). Thus, α_1H-containing channels exhibit biophysical and pharmacological properties characteristic of low voltage-activated, or T-type, Ca²⁺ channels.     

TRPC1 and TRPC5 form a novel cation channel in mammalian brain

TRP proteins are cation channels responding to receptor-dependent activation of phospholipase C. Mammalian (TRPC) channels can form hetero-oligomeric channels in vitro, but native TRPC channel complexes have not been identified to date. We demonstrate here that TRPC1 and TRPC5 are subunits of a heteromeric neuronal channel. Both TRPC proteins have overlapping distributions in the hippocampus. Coexpression of TRPC1 and TRPC5 in HEK293 cells resulted in a novel nonselective cation channel with a voltage dependence similar to NMDA receptor channels, but unlike that of any reported TRPC channel. TRPC1/TRPC5 heteromers were activated by G(q)-coupled receptors but not by depletion of intracellular Ca(2+) stores. In contrast to the more common view of the TRP family as comprising store-operated channels, we propose that many TRPC heteromers form diverse receptor-regulated nonselective cation channels in the mammalian brain.