| 2003 |
TRPV6 forms homotetramers and can form heterotetramers with TRPV5; the tetrameric stoichiometry was confirmed by sucrose gradient sedimentation (~400 kDa), co-immunoprecipitation from oocytes, and electrophysiological analysis of concatemeric channels. Heterotetrameric TRPV5/TRPV6 complexes show distinct Ca2+-dependent inactivation, Ba2+ selectivity, and pharmacological block compared to homotetramers. |
Sucrose gradient sedimentation, co-immunoprecipitation, concatemeric channel electrophysiology in HEK293 cells |
The EMBO journal |
High |
12574114
|
| 2003 |
S100A10 binds specifically to the conserved C-terminal VATTV motif of TRPV6 (with the first threonine, T600, being critical); S100A10 forms a heterotetrameric complex with annexin 2, and the S100A10–annexin 2 complex is required for routing TRPV6 to the plasma membrane. Mutation T600A abolishes S100A10 binding, redistributes TRPV6 to a sub-plasma-membrane compartment, and eliminates channel activity. Knockdown of annexin 2 by siRNA inhibits TRPV6-mediated currents. |
Yeast two-hybrid, GST pull-down, co-immunoprecipitation, site-directed mutagenesis, siRNA knockdown, electrophysiology in HEK293 cells, immunofluorescence |
The EMBO journal |
High |
12660155
|
| 2004 |
Calmodulin (CaM) binds TRPV6 in a Ca2+-dependent manner and positively regulates TRPV6 channel activity. CaM binding sites were localized to the transmembrane domain, an N-terminal 1-5-10 motif (residues 88–97), and a C-terminal 1-8-14 motif (residues 643–656). Overexpression of Ca2+-insensitive CaM mutants (CaM1234, CaM34) significantly reduced Ca2+ and Na+ currents of TRPV6-expressing HEK293 cells but not TRPV5 cells, implicating EF-hands 3 and 4 of CaM's C-lobe. |
GST pull-down, co-immunoprecipitation, whole-cell patch clamp, chimeric channel analysis in HEK293 cells |
The Journal of biological chemistry |
High |
15123711
|
| 2016 |
Crystal structure of rat TRPV6 at 3.25 Å resolution revealed: tetrameric assembly with an intracellular 'skirt' involved in allosteric modulation; Ca2+ selectivity determined by a ring of aspartate side chains in the selectivity filter; multiple cation-binding sites along the pore axis and extracellular vestibule defining a Ca2+ permeation mechanism. |
X-ray crystallography at 3.25 Å resolution |
Nature |
High |
27296226
|
| 2017 |
Cryo-EM structures of human TRPV6 in open and closed states revealed that channel opening involves an α-to-π helical transition in the pore-lining S6 helix at an alanine hinge below the selectivity filter, causing S6 helices to bend and rotate (iris-like gating). The selectivity filter adopts similar conformations in both states. Open probability is increased by PIP2. |
Cryo-electron microscopy, electrophysiology |
Nature |
High |
29258289
|
| 2018 |
2-APB binds to TRPV6 in a pocket formed by the cytoplasmic half of the S1–S4 transmembrane helix bundle. 2-APB induces channel closure by modulating protein–lipid interactions, as shown by comparing wild-type and high-affinity Y467A mutant structures. Mutagenesis and functional analysis confirmed the binding site. |
Crystal and cryo-EM structures of human and rat TRPV6 bound to 2-APB, mutagenesis, functional assays |
Nature communications |
High |
29941865
|
| 2008 |
Crystal structure of the TRPV6 ankyrin repeat domain (ARD) at 1.7 Å revealed a conserved large twist between repeats 4 and 5 and a variable third finger loop with putative regulatory phosphorylation sites. Pull-down assays showed the TRPV6 ARD does not bind ATP or calmodulin, contrasting with TRPV1 ARD, indicating a distinct regulatory role for the TRPV6 ARD. |
X-ray crystallography (1.7 Å), ATP-agarose and calmodulin-agarose pull-down assays, size exclusion chromatography |
Biochemistry |
High |
18232717
|
| 2006 |
GDP-bound Rab11a directly and specifically interacts with TRPV5 and TRPV6 via a conserved stretch in their C-termini. Rab11a colocalizes with TRPV6 in vesicular structures beneath the apical membrane. Co-expression of GDP-locked Rab11a significantly decreased TRPV6-mediated Ca2+ uptake by reducing channel surface expression. Rab11a thus mediates TRPV6 trafficking to the plasma membrane in a novel GDP-bound, direct cargo-interaction mode. |
Co-immunoprecipitation, confocal colocalization, Ca2+ uptake assays, surface expression analysis in epithelial cells and oocytes |
Molecular and cellular biology |
High |
16354700
|
| 2008 |
Klotho (acting as a β-glucuronidase) and β-glucuronidase specifically increase TRPV6 (and TRPV5) activity but not TRPV4 or TRPM6, suggesting activation via N-oligosaccharide hydrolysis on the channel. Deglycosylation by endoglycosidase-F also stimulated TRPV6 activity. |
Ca2+-influx measurements in transfected HEK293 cells, enzymatic deglycosylation |
Nephrology, dialysis, transplantation |
Medium |
18495742
|
| 2006 |
RGS2 interacts with the N-terminal domain of TRPV6 in a Ca2+-independent manner (identified by yeast two-hybrid and GST pull-down). RGS2 overexpression reduces Na+ and Ca2+ currents of TRPV6 but not TRPV5. The inhibitory effect is not due to altered channel trafficking (cell-surface biotinylation was unchanged), suggesting direct effects on gating. DeltaN-RGS2 lacking the N-terminal domain does not inhibit TRPV6, and the effect is GPCR-independent. |
Yeast two-hybrid, GST pull-down, whole-cell electrophysiology, cell-surface biotinylation in HEK293 cells |
The Journal of biological chemistry |
High |
16895908
|
| 2008 |
Ca2+ influx through TRPV6 activates phospholipase C, causing depletion of PIP2, which contributes to Ca2+-induced inactivation of TRPV6. Ba2+ (which does not activate PLC) does not cause inactivation. Dialysis of DiC8-PIP2 inhibited Ca2+-dependent inactivation; PIP2 activated TRPV6 in excised patches; rapamycin-inducible PI(4,5)P2 5-phosphatase inhibited TRPV6 currents. PI(4)P had no significant effect. |
Whole-cell and excised patch-clamp, rapamycin-inducible phosphatase system, Ca2+ imaging |
The Journal of biological chemistry |
High |
18390907
|
| 2011 |
PIP2 is a direct activator of TRPV6: reconstituted purified TRPV6 in planar lipid bilayers showed high activity in presence of PIP2 but not PI(4)P. Intracellular ATP supports TRPV6 activity indirectly by serving as substrate for type III PI4-kinases to resynthesize PIP2; MgATP reactivated TRPV6 after rundown in excised patches, an effect blocked by PI4K inhibitors. |
Excised inside-out patch clamp, planar lipid bilayer reconstitution with purified channel, pharmacological inhibition of PI4K |
FASEB journal |
High |
21810903
|
| 2008 |
Cyclophilin B (CypB) co-purifies with TRPV6 from human placenta (also with annexin A2) and co-expression of CypB with TRPV6 in Xenopus oocytes significantly increases TRPV6-mediated Ca2+ uptake. This stimulatory effect is reversed by cyclosporin A (which inhibits CypB's peptidyl-prolyl cis/trans isomerase activity), suggesting CypB activates TRPV6 through its isomerase activity. |
Protein enrichment/co-purification from human placenta, Xenopus oocyte Ca2+ uptake assay, pharmacological inhibition with cyclosporin A |
The Journal of biological chemistry |
Medium |
18445599
|
| 2009 |
ATP binds directly to TRPV6 (at sites within the ankyrin repeat domain and C-terminus), reduces whole-cell current increments, and prevents channel rundown (EC50 ~380 µM). PKCβII-mediated phosphorylation at defined ARD and C-terminal sites counteracts ATP-dependent stabilization, constituting a metabolic switch for Ca2+ influx. |
Biochemical ATP-binding studies, patch-clamp electrophysiology, site-directed mutagenesis of ARD and C-terminal residues |
FASEB journal |
Medium |
19805577
|
| 2010 |
Nedd4-2 (HECT ubiquitin E3 ligase) down-regulates TRPV6 protein abundance and Ca2+ influx by increasing TRPV6 ubiquitination and decreasing its stability (partially via proteasomal but not lysosomal pathway). The HECT domain is essential for both inhibition and association. WW1 and WW2 domains interact with TRPV6 terminal regions and their integrity limits the degree of TRPV6 ubiquitination. |
Xenopus oocyte co-expression, Ca2+ influx assays, ubiquitination assays, proteasome/lysosome inhibitors, mutagenesis |
The Journal of biological chemistry |
Medium |
20843805
|
| 2008 |
The annexin 2–S100A10 complex association with TRPV6 is regulated by cAMP/PKA/calcineurin A (CnA) signaling: forskolin-stimulated cAMP increases annexin 2–S100A10 complex formation (blocked by PKA or CnA inhibitors), and CnA-dependent dephosphorylation of annexin 2 mediates the interaction with TRPV6. PKA and CnA inhibitors attenuated 45Ca uptake in Caco-2 cells. |
Co-immunoprecipitation, 45Ca uptake assays, pharmacological inhibitors (PKA, CnA), cell biology in 16HBE14o- and Caco-2 cells |
Cell calcium |
Medium |
18187190
|
| 2006 |
NHERF4 (PDZ protein) interacts with TRPV6 via the fourth PDZ domain binding a conserved region in the TRPV6 C-terminus, distinct from the NHERF2 binding site. Interaction confirmed by yeast two-hybrid, GST pull-down, and co-immunoprecipitation. |
Yeast two-hybrid, GST pull-down, co-immunoprecipitation in HEK293 cells and oocytes |
Pflugers Archiv |
Medium |
16565876
|
| 2008 |
Nipsnap1 was identified as a novel TRPV6-interacting protein (by GST pull-down/bioinformatics) that abolishes TRPV6 currents without reducing plasma membrane expression of TRPV6, suggesting direct inhibition of channel gating rather than trafficking. |
GST pull-down, electrophysiology, cell-surface biotinylation, RT-PCR, immunohistochemistry |
Pflugers Archiv |
Medium |
18392847
|
| 2011 |
Male mice carrying the TRPV6-D541A pore mutation (rendering the channel nonfunctional) show severely impaired fertility due to failure of Ca2+ absorption by epididymal epithelium, resulting in ~10-fold higher Ca2+ in cauda epididymis luminal fluid. TRPV6 was localized to the apical membrane of epididymal epithelium (not in spermatozoa), establishing that TRPV6-mediated Ca2+ absorption in the epididymis is essential for sperm motility and viability. |
Knock-in mouse model (D541A), Ca2+ concentration measurements, Ca2+ absorption assays, immunolocalization, sperm functional assays |
Science signaling |
High |
21540454
|
| 2012 |
Trpv6 knockout mice (deletion of transmembrane/pore-forming and C-terminal regions) phenocopy the TRPV6-D541A pore mutant regarding epididymal Ca2+ handling and male fertility defects, arguing against residual channel-independent functions of the D541A mutant. |
Trpv6 knockout mouse (exon deletion), comparison with D541A knock-in, Ca2+ uptake assays, sperm function tests |
The Journal of biological chemistry |
High |
22427671
|
| 2008 |
TRPV6 is required for maternal-fetal Ca2+ transport: Trpv6 knockout fetuses had significantly lower fetal blood Ca2+, lower amniotic fluid Ca2+, ~40% lower 45Ca transport from mother to fetus, and lower ash weight (bone mineralization). TRPV6 mRNA and protein were localized in intraplacental and extraplacental yolk sac, co-localizing with calbindin-D9K. |
Trpv6 knockout mice, 45Ca transport assays, immunohistochemistry, qRT-PCR |
Journal of bone and mineral research |
High |
18348695
|
| 2012 |
TRPV6-D541A knockin mice (non-functional pore) show significantly impaired intestinal Ca2+ uptake under Ca2+-deficient diet conditions, establishing that functional TRPV6 channels mediate transepithelial Ca2+ absorption in the intestine, particularly under low-Ca2+ dietary challenge. |
TRPV6-D541A knockin mouse model, 45Ca intestinal absorption assay, metabolic cage measurements, protein expression analysis |
American journal of physiology. Gastrointestinal and liver physiology |
High |
22878123
|
| 2004 |
Under physiological intracellular Ca2+ buffering conditions, TRPV6 does not form constitutively open channels in HEK293 cells; channel activity requires chelation of intracellular Ca2+ (with EGTA). Monovalent cations permeate TRPV6 when Ca2+ permeation is absent, indicating extracellular divalent cation block. The C-terminal domain required for calmodulin binding is not necessary for this regulation; the pore residue D542 is essential for channel function. |
Whole-cell and perforated patch clamp, Fura-2 microfluorimetry, site-directed mutagenesis (D542A, truncation mutant) in HEK293 cells |
The Journal of biological chemistry |
High |
15184369
|
| 2013 |
The in vivo TRPV6 protein has an extended N-terminus: translation initiates at a non-AUG codon (ACG, decoded as methionine) upstream of the annotated AUG (which is not used). The extended full-length protein has similar channel properties to the shorter form in vitro, but increased trafficking to the plasma membrane and provides an additional scaffold for channel assembly. |
Mass spectrometry of endogenous placenta protein, cell biology/trafficking assays, electrophysiology |
The Journal of biological chemistry |
Medium |
23612980
|
| 2011 |
G protein-coupled receptor PSGR activates endogenous TRPV6 channels in prostate cells via Src kinase. PSGR stimulation enhances cytosolic Ca2+ through TRPV6; the Ca2+ signal has biophysical characteristics of TRPV6 currents; Src kinase activation occurs independently of G-protein activation, presumably through direct PSGR–Src interaction. siRNA knockdown of TRPV6 confirmed channel identity. |
Electrophysiology, live-cell Ca2+ imaging, siRNA knockdown, biochemical co-immunoprecipitation, Src kinase inhibition |
The Journal of biological chemistry |
Medium |
21349844
|
| 2009 |
WNK3 positively regulates TRPV6 activity in a kinase-dependent manner; the kinase-inactive WNK3-D294A mutation abolishes the effect. WNK3 increases Ca2+ influx and Na+ current mediated by TRPV6, and enhances delivery of mature (glycosylated) TRPV6 to the plasma membrane via the secretory pathway; the effect is abolished by the microtubule inhibitor colchicine. |
Xenopus oocyte co-expression, Ca2+ uptake assays, voltage clamp, surface biotinylation, exocytosis assays, kinase-dead mutagenesis |
American journal of physiology. Renal physiology |
Medium |
18768590
|
| 2009 |
SGK1 and PKB/Akt increase TRPV6 activity and plasma membrane abundance. PIKfyve, phosphorylated by SGK1 at S318, further augments TRPV6 activity when co-expressed with active SGK1; the S318A-PIKfyve mutant lacking the SGK1 phosphorylation site fails to enhance TRPV6 activity. |
Xenopus oocyte co-expression, Ca2+-activated Cl- current recording as indirect readout of TRPV6-mediated Ca2+ influx, immunohistochemistry, Western blotting |
The Journal of membrane biology |
Low |
20041238
|
| 2014 |
TRPV6 translocates to the plasma membrane in prostate cancer cells via an Orai1/TRPC1-mediated Ca2+/Annexin I/S100A11 pathway. TRPV6-mediated Ca2+ entry increases cell survival (proliferation and apoptosis resistance). Xenograft models confirmed increased aggressiveness of TRPV6-overexpressing tumors. |
Co-immunoprecipitation, Ca2+ imaging, siRNA knockdown, xenograft mouse models, immunohistochemistry |
Proceedings of the National Academy of Sciences |
Medium |
25172921
|
| 2007 |
TRPV6 mediates Ca2+ entry in human keratinocytes and is required for Ca2+-induced differentiation: siRNA-mediated TRPV6 silencing impairs differentiated phenotype, reduces differentiation markers (involucrin, transglutaminase-1, cytokeratin-10), disrupts cell morphology and stratification. 1,25-Dihydroxyvitamin D3 dose-dependently increases TRPV6 expression and Ca2+ uptake, promoting differentiation. |
siRNA knockdown, Ca2+ uptake assays (Fura-2), RT-PCR, immunoblotting in primary human keratinocytes |
The Journal of biological chemistry |
Medium |
17550901
|
| 2007 |
TRPV6 controls prostate cancer cell (LNCaP) proliferation: siRNA knockdown of TRPV6 decreases proliferation rate, reduces S-phase accumulation, and decreases PCNA expression. TRPV6-mediated Ca2+ entry activates NFAT transcription factor downstream. TRPV6 also contributes to apoptosis resistance. |
siRNA knockdown, cell cycle analysis, Ca2+ imaging, NFAT reporter assays in LNCaP cells |
Oncogene |
Medium |
17533368
|
| 2019 |
Activation of the Ca2+-sensing receptor (CaSR) on the basolateral membrane of intestinal epithelium directly attenuates Ca2+ absorption via TRPV6: cinacalcet (calcimimetic) decreased net Ca2+ flux in Ussing chambers in a TRPV6-dependent manner (absent in TRPV6-D541A mice), and inhibited Ca2+ flux through TRPV6 when co-expressed with CaSR in oocytes. PLC inhibitor U73122 prevented cinacalcet-mediated TRPV6 inhibition. |
Ussing chamber Ca2+ flux assays, TRPV6-D541A knockin mice, Xenopus oocyte co-expression, pharmacological inhibition |
JCI insight |
High |
31013259
|
| 2010 |
TRPV6 permeates heavy metal cations including Zn2+, Cd2+, Ba2+, Sr2+, Mn2+, La3+, and Gd3+ in addition to Ca2+, as shown by live-cell ion imaging (Fura-2, Newport Green DCF) and patch clamp in hTRPV6-expressing HEK293 cells. At higher concentrations, Cd2+, La3+, and Gd3+ are efficient inhibitors of TRPV6-mediated Ca2+ influx. |
Live-cell ion imaging (Fura-2, Newport Green DCF), whole-cell patch clamp, 45Ca uptake assays in HEK293 cells |
Cell calcium |
Medium |
21146870
|
| 2012 |
Numb1 isoform interacts with TRPV6 via an aspartate at residue 716 (TRPV6) and arginine at residue 434 (Numb1), as identified by co-immunoprecipitation, FRET, and C-terminal truncation mutagenesis. Numb1 negatively regulates TRPV6 activity: its expression decreases cytosolic Ca2+ in TRPV6-transfected cells, and a Numb1 mutant lacking the TRPV6-binding site fails to inhibit TRPV6. |
Co-immunoprecipitation, FRET, C-terminal truncation mutagenesis, Ca2+ measurements in HEK293 and cancer cells |
Cell calcium |
Medium |
23140583
|
| 2020 |
TRPV6 autoinhibition is maintained by two intramolecular interactions: S4–S5 linker to C-terminal TRP helix (L/C; mediated by Arg470:Trp593) and N-terminal pre-S1 to TRP helix (N/C; mediated by Trp321:Ile597). Disruption of either interaction by mutations or blocking peptides activates TRPV6. The N/C interaction depends on L/C but not vice versa. PIP2 binds three cationic residues in S5/C-terminus, suppresses both interactions, and activates TRPV6. |
Site-directed mutagenesis, blocking peptides, patch-clamp electrophysiology, Ca2+ imaging |
iScience |
Medium |
32829285
|
| 2016 |
In rat cauda epididymal principal cells, a constitutively active TRPV6-like Ca2+ current is coupled to a calcium-activated chloride conductance (TMEM16A). TRPV6 and TMEM16A proteins co-localize at the apical membrane. Removal of extracellular Ca2+ attenuates both currents; lanthanide block inhibits the TRPV6-like component first, then CaCC. In vivo perfusion showed substantial Ca2+ reabsorption from epididymal lumen suppressed by ruthenium red. |
Patch clamp in single epididymal cells, pharmacological inhibition, in vivo luminal perfusion, immunofluorescence colocalization, mRNA detection |
The Journal of general physiology |
Medium |
27481714
|
| 2019 |
In zebrafish, Trpv6 functions as a cell-autonomous regulator of epithelial quiescence: Trpv6-mediated constitutive Ca2+ influx activates PP2A, which suppresses IGF-mediated Akt-Tor and Erk signaling to maintain cellular quiescence. Genetic deletion or pharmacological blockade of Trpv6 caused epithelial cells to exit quiescence and re-enter the cell cycle; re-introduction of Trpv6 but not a channel-dead mutant restored quiescence. |
Zebrafish genetic deletion, pharmacological inhibition, cell cycle analysis, Ca2+ imaging, PP2A activity assays, IGF signaling readouts, human colon cancer cell experiments |
eLife |
Medium |
31526479
|
| 2022 |
TRPV6 is required for alcohol-induced intestinal barrier dysfunction: ethanol and acetaldehyde activate TRPV6 ionic currents in Caco-2 cells; TRPV6 deficiency (Trpv6-/- mice and organoids) attenuates ethanol/acetaldehyde-induced Ca2+ influx, tight junction disruption, and barrier dysfunction. Photoaffinity labeling of 3-azibutanol identified a histidine as a putative alcohol-binding site; substitution of this histidine (and a nearby arginine) reduces ethanol-activated TRPV6 currents. |
Patch clamp electrophysiology, Ca2+ imaging, Trpv6-/- mouse model, intestinal organoids, Caco-2 monolayers, photoaffinity labeling, site-directed mutagenesis |
Cell reports |
High |
35705057
|
| 2021 |
TRPV6 promotes breast cancer metastasis via NFATC2IP phosphorylation at Ser204 (with CDK5 as candidate kinase), which activates NFATC2 transcription factor, leading to upregulation of ADAMTS6 and increased cell migration. TRPV6 overexpression accelerates migration; TRPV6 suppression decreases it. |
Overexpression and siRNA knockdown, phosphorylation analysis, reporter assays, migration assays in breast cancer cells |
Cancer letters |
Low |
34265397
|
| 2020 |
TRPV6 suppresses osteoclastogenesis via the IGF-PI3K-AKT signaling pathway: TRPV6 knockout mice develop osteoporosis with enhanced osteoclast differentiation and bone resorption. TRPV6 located on the osteoclast cell membrane decreases p-IGF/total-IGF, p-PI3K/total-PI3K, and p-AKT/total-AKT ratios. Blocking IGF-PI3K-AKT with inhibitors relieved the inhibitory effect of TRPV6 on osteoclasts. |
Trpv6 knockout mice, lentiviral overexpression/silencing in osteoclasts, micro-CT, TRAP staining, pit formation assay, Western blot for pathway components |
Cell proliferation |
Medium |
33159483
|