The Experts below are selected from a list of 360 Experts worldwide ranked by ideXlab platform
Bernd Nilius - One of the best experts on this subject based on the ideXlab platform.
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Pharmacology of Vanilloid Transient Receptor Potential Cation Channels
Molecular Pharmacology, 2009Co-Authors: Joris Vriens, Giovanni Appendino, Bernd NiliusAbstract:Depending on their primary structure, the 28 mammalian transient receptor potential (TRP) cation channels identified so far can be sorted into 6 subfamilies: TRPC (“Canonical”), TRPV (“Vanilloid”), TRPM (“Melastatin”), TRPP (“Polycystin”), TRPML (“Mucolipin”), and TRPA (“Ankyrin”). The TRPV subfamily (vanilloid receptors) comprises channels critically involved in nociception and thermosensing (TRPV1, TRPV2, TRPV3, and TRPV4), whereas TRPV5 and TRPV6 are involved in renal Ca 2 absorption/reabsorption. Apart from TRPV1, the pharmacology of these channels is still insufficiently known. Furthermore, only few small-molecule ligands for non-TRPV1 vanilloid receptors have been identified, and little is known of their endogenous ligands, resulting in a substantial “orphan” state for these channels. In this review, we summarize the pharmacological properties of members of the TRPV subfamily, highlighting the critical issues and challenges facing their “deorphanization” and clinical exploitation.
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Vanilloid Transient Receptor Potential Cation Channels : An Overview
Current Pharmaceutical Design, 2008Co-Authors: Rudi Vennekens, Grzegorz Owsianik, Bernd NiliusAbstract:The mammalian branch of the Transient Receptor Potential (TRP) superfamily of cation channels consists of 28 members. They can be subdivided in six main subfamilies: the TRPC (‘Canonical’), TRPV (‘Vanilloid’), TRPM (‘Melastatin’), TRPP (‘Polycystin’), TRPML (‘Mucolipin’) and the TRPA (‘Ankyrin’) group. The TRPV subfamily comprises channels that are critically involved in nociception and thermo-sensing (TRPV1, TRPV2, TRPV3, TRPV4) as well as highly Ca2+ selective channels involved in Ca2+ absorption/ reabsorption in mammals (TRPV5, TRPV6). In this review we summarize fundamental physiological properties of all TRPV members in the light of various cellular functions of these channels and their significance in the systemic context of the mammalian organism.
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regulation of transient receptor potential trp channels by phosphoinositides
Pflügers Archiv: European Journal of Physiology, 2007Co-Authors: Tibor Rohacs, Bernd NiliusAbstract:This review summarizes the modulation of transient receptor potential (TRP) channels, by phosphoinositides. TRP channels are characterized by polymodal activation and a surprising complexity of regulation mechanisms. Possibly, most if not all TRP channels are modulated by phosphoinositides. Modulation by phosphatidylinositol 4,5-biphosphate (PIP2) has been shown in detail for TRP vanilloid (TRPV) 1, TRPV5, TRP melastatin (TRPM) 4, TRPM5, TRPM7, TRPM8, TRP polycystin 2, and the Drosophila TPR-like (TRPL) channels. This review describes mechanisms of modulation of TRP channels mainly by PIP2 and discusses some future challenges of this fascinating topic.
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the selectivity filter of the cation channel TRPM4
Journal of Biological Chemistry, 2005Co-Authors: Bernd Nilius, Grzegorz Owsianik, Annelies Janssens, Jean Prenen, Chunbo Wang, Michael X Zhu, Thomas VoetsAbstract:Transient receptor potential channel melastatin subfamily (TRPM) 4 and its close homologue, TRPM5, are the only two members of the large transient receptor potential superfamily of cation channels that are impermeable to Ca2+. In this study, we located the TRPM4 selectivity filter and investigated possible structural elements that render it Ca2+-impermeable. Based on homology with known cation channel pores, we identified an acidic stretch of six amino acids in the loop between transmembrane helices TM5 and TM6 (981EDMDVA986) as a potential selectivity filter. Substitution of this six-amino acid stretch with the selectivity filter of TRPV6 (TIIDGP) resulted in a functional channel that combined the gating hallmarks of TRPM4 (activation by Ca2+, voltage dependence) with TRPV6-like sensitivity to block by extracellular Ca2+ and Mg2+ as well as Ca2+ permeation. Neutralization of Glu981 resulted in a channel with normal permeability properties but a strongly reduced sensitivity to block by intracellular spermine. Neutralization of Asp982 yielded a functional channel that exhibited extremely fast desensitization (τ
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comparison of functional properties of the ca2 activated cation channels TRPM4 and trpm5 from mice
Cell Calcium, 2005Co-Authors: Nina Ullrich, Rudi Vennekens, Jean Prenen, Guy Droogmans, Thomas Voets, Karel Talavera, Bernd NiliusAbstract:Non-selective cation (NSC) channels activated by intracellular Ca2+ ([Ca2+]i) play an important role in Ca2+ signaling and membrane excitability in many cell types. TRPM4 and TRPM5, two Ca2+-activated cation channels of the TRP superfamily, are potential molecular correlates of NSC channels. We compared the functional properties of mouse TRPM4 and TRPM5 heterologously expressed in HEK 293 cells. Dialyzing cells with different Ca2+ concentrations revealed a difference in Ca2+ sensitivity between TRPM4 and TRPM5, with EC50 values of 20.2+/-4.0 microM and 0.70+/-0.1 microM, respectively. Similarly, TRPM5 activated at lower Ca2+ concentration than TRPM4 when [Ca2+]i was raised by UV uncaging of the Ca2+-cage DMNP-EDTA. Current amplitudes of TRPM4 and TRPM5 were not correlated to the rate of changes in [Ca2+]i. The Ca2+ sensitivity of both channels was strongly reduced in inside-out patches, resulting in approximately 10-30 times higher EC50 values than under whole-cell conditions. Currents through TRPM4 and TRPM5 deactivated at negative and activated at positive potentials with similar kinetics. Both channels were equally sensitive to block by intracellular spermine. TRPM4 displayed a 10-fold higher affinity for block by flufenamic acid. Importantly, ATP4- blocked TRPM4 with high affinity (IC50 of 0.8+/-0.1 microM), whereas TRPM5 is insensitive to ATP4- at concentrations up to 1 mM.
Thomas Gudermann - One of the best experts on this subject based on the ideXlab platform.
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role of kinase coupled trp channels in mineral homeostasis
Pharmacology & Therapeutics, 2017Co-Authors: Vladimir Chubanov, Lorenz Mittermeier, Thomas GudermannAbstract:Abstract Transient receptor potential (TRP) proteins TRPM6 and TRPM7 are α-kinase-coupled divalent cation-selective channels activated upon a reduction of cytosolic levels of Mg2+ and Mg·ATP. Emerging evidence indicate that one of the main physiological functions of TRPM6 and TRPM7 is maintaining of cellular metabolism of Mg2+ and likely other essential metals such as Ca2+ and Zn2+. Recent experiments with genetic animal models have shown that TRPM6 and TRPM7 are essential for epithelial Mg2+ transport in the placenta and intestine. In addition, mutations in TRPM6 or TRPM7 have been linked to Mg2+ deficiency in humans. However, many key functional aspects of these remarkable proteins as well as mechanisms of the associated channelopathies remain incompletely understood. The present review article highlights the recent significant progress in the field with the focus on the vital roles of TRPM7 and TRPM7 in mineral homeostasis.
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trpm6 and trpm7 differentially contribute to the relief of heteromeric trpm6 7 channels from inhibition by cytosolic mg 2 and mg atp
Scientific Reports, 2017Co-Authors: Silvia Ferioli, Susanna Zierler, Thomas Gudermann, Joanna Zaiserer, Johann Schredelseker, Vladimir ChubanovAbstract:TRPM6 and its homologue TRPM7 are α-kinase-coupled divalent cation-selective channels activated upon reduction of cytosolic levels of Mg2+ and Mg·ATP. TRPM6 is vital for organismal Mg2+ balance. However, mechanistically the cellular role and functional nonredundancy of TRPM6 remain incompletely understood. Comparative analysis of native currents in primary cells from TRPM6- versus TRPM7-deficient mice supported the concept that native TRPM6 primarily functions as a constituent of heteromeric TRPM6/7 channels. However, heterologous expression of the human TRPM6 protein engendered controversial results with respect to channel characteristics including its regulation by Mg2+ and Mg·ATP. To resolve this issue, we cloned the mouse TRPM6 (mTRPM6) cDNA and compared its functional characteristics to mouse TRPM7 (mTRPM7) after heterologous expression. Notably, we observed that mTRPM6 and mTRPM7 differentially regulate properties of heteromeric mTRPM6/7 channels: In the presence of mTRPM7, the extreme sensitivity of functionally expressed homomeric mTRPM6 to Mg2+ is tuned to higher concentrations, whereas mTRPM6 relieves mTRPM7 from the tight inhibition by Mg·ATP. Consequently, the association of mTRPM6 with mTRPM7 allows for high constitutive activity of mTRPM6/7 in the presence of physiological levels of Mg2+ and Mg·ATP, thus laying the mechanistic foundation for constant vectorial Mg2+ transport specifically into epithelial cells.
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Evolutionary determinants of divergent calcium selectivity of TRPM channels
The FASEB Journal, 2007Co-Authors: Michael Mederos Y Schnitzler, Thomas Gudermann, Janine Wäring, Vladimir ChubanovAbstract:The mammalian TRPM gene family can be subdivided into distinct categories of cation channels that are either highly permeable for Ca(2+) (TRPM3/6/7), nonselective (TRPM2/8), or even Ca(2+) impermeable (TRPM4/5). TRPM6/7 are fused to alpha-kinase domains, whereas TRPM2 is linked to an ADP-ribose phosphohydrolase (Nudix domain). At a molecular level, the evolutionary steps that gave rise to the structural and functional TRPM channel diversity remain elusive. Here, we provide phylogenetic evidence that Nudix-linked channels represent an ancestral type of TRPMs that is present in various phyla, ranging from protists to humans. Surprisingly, the pore-forming segments of invertebrate TRPM2-like proteins display high sequence similarity to those of Ca(2+)-selective TRPMs, while human TRPM2 is characterized by a loss of several conserved residues. Using the patch-clamp technique, Ca(2+) imaging, and site-directed mutagenesis, we demonstrate that restoration of only two "ancient" pore residues in human TRPM2 (Q981E/P983Y) significantly increased (approximately 4-fold) its permeability for Ca(2+). Conversely, introduction of a "modern" sequence motif into mouse TRPM7 (E1047Q/Y1049P) resulted in the loss of Ca(2+) permeation and a linear TRPM2-like current-voltage relationship. Overall, our findings provide an integrative view on the evolution of the domain architecture and the structural basis of the distinct ion permeation profiles of TRPM channels.
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hypomagnesemia with secondary hypocalcemia due to a missense mutation in the putative pore forming region of trpm6
Journal of Biological Chemistry, 2007Co-Authors: Vladimir Chubanov, Siegfried Waldegger, Michael Mederos Y Schnitzler, Karl P Schlingmann, Janine Wäring, Jolanta Heinzinger, Silke Kaske, Thomas GudermannAbstract:Abstract Hypomagnesemia with secondary hypocalcemia is an autosomal recessive disorder caused by mutations in the TRPM6 gene. Current experimental evidence suggests that TRPM6 may function in a specific association with TRPM7 by means of heterooligomeric channel complex formation. Here, we report the identification and functional characterization of a new hypomagnesemia with secondary hypocalcemia missense mutation in TRPM6. The affected subject presented with profound hypomagnesemia and hypocalcemia caused by compound heterozygous mutation in the TRPM6 gene: 1208(-1)G > A affecting the acceptor splice site preceding exon 11, and 3050C > G resulting in the amino acid change (P1017R) in the putative pore-forming region of TRPM6. To assess the functional consequences of the P1017R mutation, TRPM6P1017R and wild-type TRPM6 were co-expressed with TRPM7 in Xenopus oocytes and HEK 293 cells, and currents were assessed by two-electrode voltage clamp and whole cell patch clamp measurements, respectively. Co-expression of wild-type TRPM6 and TRPM7 resulted in a significant increase in the amplitude of TRPM7-like currents. In contrast, TRPM6P1017R suppressed TRPM7 channel activity. In line with these observations, TRPM7, containing the corresponding mutation P1040R, displayed a dominant-negative effect upon co-expression with wild-type TRPM7. Confocal microscopy and fluorescence resonance energy transfer recordings demonstrated that the P1017R mutation neither affects assembly of TRPM6 with TRPM7, nor co-trafficking of heteromultimeric channel complexes to the cell surface. We conclude that a functional defect in the putative pore of TRPM6/7 channel complexes is sufficient to impair body magnesium homeostasis.
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hypomagnesemia with secondary hypocalcemia due to a missense mutation in the putative pore forming region of trpm6
Journal of Biological Chemistry, 2007Co-Authors: Vladimir Chubanov, Siegfried Waldegger, Michael Mederos Y Schnitzler, Karl P Schlingmann, Janine Wäring, Jolanta Heinzinger, Silke Kaske, Thomas GudermannAbstract:Abstract Hypomagnesemia with secondary hypocalcemia is an autosomal recessive disorder caused by mutations in the TRPM6 gene. Current experimental evidence suggests that TRPM6 may function in a specific association with TRPM7 by means of heterooligomeric channel complex formation. Here, we report the identification and functional characterization of a new hypomagnesemia with secondary hypocalcemia missense mutation in TRPM6. The affected subject presented with profound hypomagnesemia and hypocalcemia caused by compound heterozygous mutation in the TRPM6 gene: 1208(-1)G > A affecting the acceptor splice site preceding exon 11, and 3050C > G resulting in the amino acid change (P1017R) in the putative pore-forming region of TRPM6. To assess the functional consequences of the P1017R mutation, TRPM6P1017R and wild-type TRPM6 were co-expressed with TRPM7 in Xenopus oocytes and HEK 293 cells, and currents were assessed by two-electrode voltage clamp and whole cell patch clamp measurements, respectively. Co-expression of wild-type TRPM6 and TRPM7 resulted in a significant increase in the amplitude of TRPM7-like currents. In contrast, TRPM6P1017R suppressed TRPM7 channel activity. In line with these observations, TRPM7, containing the corresponding mutation P1040R, displayed a dominant-negative effect upon co-expression with wild-type TRPM7. Confocal microscopy and fluorescence resonance energy transfer recordings demonstrated that the P1017R mutation neither affects assembly of TRPM6 with TRPM7, nor co-trafficking of heteromultimeric channel complexes to the cell surface. We conclude that a functional defect in the putative pore of TRPM6/7 channel complexes is sufficient to impair body magnesium homeostasis.
Vladimir Chubanov - One of the best experts on this subject based on the ideXlab platform.
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role of kinase coupled trp channels in mineral homeostasis
Pharmacology & Therapeutics, 2017Co-Authors: Vladimir Chubanov, Lorenz Mittermeier, Thomas GudermannAbstract:Abstract Transient receptor potential (TRP) proteins TRPM6 and TRPM7 are α-kinase-coupled divalent cation-selective channels activated upon a reduction of cytosolic levels of Mg2+ and Mg·ATP. Emerging evidence indicate that one of the main physiological functions of TRPM6 and TRPM7 is maintaining of cellular metabolism of Mg2+ and likely other essential metals such as Ca2+ and Zn2+. Recent experiments with genetic animal models have shown that TRPM6 and TRPM7 are essential for epithelial Mg2+ transport in the placenta and intestine. In addition, mutations in TRPM6 or TRPM7 have been linked to Mg2+ deficiency in humans. However, many key functional aspects of these remarkable proteins as well as mechanisms of the associated channelopathies remain incompletely understood. The present review article highlights the recent significant progress in the field with the focus on the vital roles of TRPM7 and TRPM7 in mineral homeostasis.
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trpm6 and trpm7 differentially contribute to the relief of heteromeric trpm6 7 channels from inhibition by cytosolic mg 2 and mg atp
Scientific Reports, 2017Co-Authors: Silvia Ferioli, Susanna Zierler, Thomas Gudermann, Joanna Zaiserer, Johann Schredelseker, Vladimir ChubanovAbstract:TRPM6 and its homologue TRPM7 are α-kinase-coupled divalent cation-selective channels activated upon reduction of cytosolic levels of Mg2+ and Mg·ATP. TRPM6 is vital for organismal Mg2+ balance. However, mechanistically the cellular role and functional nonredundancy of TRPM6 remain incompletely understood. Comparative analysis of native currents in primary cells from TRPM6- versus TRPM7-deficient mice supported the concept that native TRPM6 primarily functions as a constituent of heteromeric TRPM6/7 channels. However, heterologous expression of the human TRPM6 protein engendered controversial results with respect to channel characteristics including its regulation by Mg2+ and Mg·ATP. To resolve this issue, we cloned the mouse TRPM6 (mTRPM6) cDNA and compared its functional characteristics to mouse TRPM7 (mTRPM7) after heterologous expression. Notably, we observed that mTRPM6 and mTRPM7 differentially regulate properties of heteromeric mTRPM6/7 channels: In the presence of mTRPM7, the extreme sensitivity of functionally expressed homomeric mTRPM6 to Mg2+ is tuned to higher concentrations, whereas mTRPM6 relieves mTRPM7 from the tight inhibition by Mg·ATP. Consequently, the association of mTRPM6 with mTRPM7 allows for high constitutive activity of mTRPM6/7 in the presence of physiological levels of Mg2+ and Mg·ATP, thus laying the mechanistic foundation for constant vectorial Mg2+ transport specifically into epithelial cells.
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Evolutionary determinants of divergent calcium selectivity of TRPM channels
The FASEB Journal, 2007Co-Authors: Michael Mederos Y Schnitzler, Thomas Gudermann, Janine Wäring, Vladimir ChubanovAbstract:The mammalian TRPM gene family can be subdivided into distinct categories of cation channels that are either highly permeable for Ca(2+) (TRPM3/6/7), nonselective (TRPM2/8), or even Ca(2+) impermeable (TRPM4/5). TRPM6/7 are fused to alpha-kinase domains, whereas TRPM2 is linked to an ADP-ribose phosphohydrolase (Nudix domain). At a molecular level, the evolutionary steps that gave rise to the structural and functional TRPM channel diversity remain elusive. Here, we provide phylogenetic evidence that Nudix-linked channels represent an ancestral type of TRPMs that is present in various phyla, ranging from protists to humans. Surprisingly, the pore-forming segments of invertebrate TRPM2-like proteins display high sequence similarity to those of Ca(2+)-selective TRPMs, while human TRPM2 is characterized by a loss of several conserved residues. Using the patch-clamp technique, Ca(2+) imaging, and site-directed mutagenesis, we demonstrate that restoration of only two "ancient" pore residues in human TRPM2 (Q981E/P983Y) significantly increased (approximately 4-fold) its permeability for Ca(2+). Conversely, introduction of a "modern" sequence motif into mouse TRPM7 (E1047Q/Y1049P) resulted in the loss of Ca(2+) permeation and a linear TRPM2-like current-voltage relationship. Overall, our findings provide an integrative view on the evolution of the domain architecture and the structural basis of the distinct ion permeation profiles of TRPM channels.
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hypomagnesemia with secondary hypocalcemia due to a missense mutation in the putative pore forming region of trpm6
Journal of Biological Chemistry, 2007Co-Authors: Vladimir Chubanov, Siegfried Waldegger, Michael Mederos Y Schnitzler, Karl P Schlingmann, Janine Wäring, Jolanta Heinzinger, Silke Kaske, Thomas GudermannAbstract:Abstract Hypomagnesemia with secondary hypocalcemia is an autosomal recessive disorder caused by mutations in the TRPM6 gene. Current experimental evidence suggests that TRPM6 may function in a specific association with TRPM7 by means of heterooligomeric channel complex formation. Here, we report the identification and functional characterization of a new hypomagnesemia with secondary hypocalcemia missense mutation in TRPM6. The affected subject presented with profound hypomagnesemia and hypocalcemia caused by compound heterozygous mutation in the TRPM6 gene: 1208(-1)G > A affecting the acceptor splice site preceding exon 11, and 3050C > G resulting in the amino acid change (P1017R) in the putative pore-forming region of TRPM6. To assess the functional consequences of the P1017R mutation, TRPM6P1017R and wild-type TRPM6 were co-expressed with TRPM7 in Xenopus oocytes and HEK 293 cells, and currents were assessed by two-electrode voltage clamp and whole cell patch clamp measurements, respectively. Co-expression of wild-type TRPM6 and TRPM7 resulted in a significant increase in the amplitude of TRPM7-like currents. In contrast, TRPM6P1017R suppressed TRPM7 channel activity. In line with these observations, TRPM7, containing the corresponding mutation P1040R, displayed a dominant-negative effect upon co-expression with wild-type TRPM7. Confocal microscopy and fluorescence resonance energy transfer recordings demonstrated that the P1017R mutation neither affects assembly of TRPM6 with TRPM7, nor co-trafficking of heteromultimeric channel complexes to the cell surface. We conclude that a functional defect in the putative pore of TRPM6/7 channel complexes is sufficient to impair body magnesium homeostasis.
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hypomagnesemia with secondary hypocalcemia due to a missense mutation in the putative pore forming region of trpm6
Journal of Biological Chemistry, 2007Co-Authors: Vladimir Chubanov, Siegfried Waldegger, Michael Mederos Y Schnitzler, Karl P Schlingmann, Janine Wäring, Jolanta Heinzinger, Silke Kaske, Thomas GudermannAbstract:Abstract Hypomagnesemia with secondary hypocalcemia is an autosomal recessive disorder caused by mutations in the TRPM6 gene. Current experimental evidence suggests that TRPM6 may function in a specific association with TRPM7 by means of heterooligomeric channel complex formation. Here, we report the identification and functional characterization of a new hypomagnesemia with secondary hypocalcemia missense mutation in TRPM6. The affected subject presented with profound hypomagnesemia and hypocalcemia caused by compound heterozygous mutation in the TRPM6 gene: 1208(-1)G > A affecting the acceptor splice site preceding exon 11, and 3050C > G resulting in the amino acid change (P1017R) in the putative pore-forming region of TRPM6. To assess the functional consequences of the P1017R mutation, TRPM6P1017R and wild-type TRPM6 were co-expressed with TRPM7 in Xenopus oocytes and HEK 293 cells, and currents were assessed by two-electrode voltage clamp and whole cell patch clamp measurements, respectively. Co-expression of wild-type TRPM6 and TRPM7 resulted in a significant increase in the amplitude of TRPM7-like currents. In contrast, TRPM6P1017R suppressed TRPM7 channel activity. In line with these observations, TRPM7, containing the corresponding mutation P1040R, displayed a dominant-negative effect upon co-expression with wild-type TRPM7. Confocal microscopy and fluorescence resonance energy transfer recordings demonstrated that the P1017R mutation neither affects assembly of TRPM6 with TRPM7, nor co-trafficking of heteromultimeric channel complexes to the cell surface. We conclude that a functional defect in the putative pore of TRPM6/7 channel complexes is sufficient to impair body magnesium homeostasis.
Thomas Voets - One of the best experts on this subject based on the ideXlab platform.
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functional expression of transient receptor potential channels in human endometrial stromal cells during the luteal phase of the menstrual cycle
Human Reproduction, 2015Co-Authors: Katrien De Clercq, Thomas Voets, Rieta Van Bree, Karen Peeraer, Katharina Held, Christel Meuleman, Carla Tomassetti, Thomas Dhooghe, Joris VriensAbstract:STUDY QUESTION Are members of the transient receptor potential (TRP) channel superfamily functionally expressed in the human endometrial stroma? SUMMARY ANSWER The Ca(2+)-permeable ion channels TRPV2, TRPV4, TRPC6 and TRPM7 are functionally expressed in primary endometrial stromal cells. WHAT IS KNOWN ALREADY Intercellular communication between epithelial and stromal endometrial cells is required to initiate decidualization, a prerequisite for successful implantation. TRP channels are possible candidates as signal transducers involved in cell-cell communication, but no fingerprint is available of the functional distribution of TRP channels in the human endometrium during the luteal phase of the menstrual cycle. STUDY DESIGN, SIZE, DURATION Endometrial biopsy samples (previously frozen) from patients of reproductive age with regular menstrual cycles, who were undergoing diagnostic laparoscopic surgery for pain and/or infertility, were analysed. Samples were obtained from the menstrual (Days 1-5, n = 3), follicular (Days 6-14, n = 6), early luteal (Days 15-20, n = 5) and late luteal (Days 21-28, n = 5) phases. In addition, a total of 13 patient samples taken during the luteal phase were used to set up primary cell cultures for further experiments. PARTICIPANTS/MATERIALS, SETTING, METHODS Quantitative real-time PCR (qRT-PCR), immunocytochemistry, Fura2-based Ca(2+)-microfluorimetry and whole-cell patch clamp experiments were performed to study the functional expression pattern of TRP channels. Specific pharmacological agents, such as Δ(9)-tetrahydrocannabinol, GSK1016790A and 1-oleoyl-2-acetyl-glycerol, were used to functionally assess the expression of TRPV2, TRPV4 and TRPC6, respectively. MAIN RESULTS AND THE ROLE OF CHANCE Expression of TRPV2, TRPV4, TRPC1, TRPC4, TRPC6, TRPM4 and TRPM7 was detected at the mRNA level in endometrial biopsies (n = 19) and in primary endometrial stromal cell cultures obtained from patients during the luteal phase (n = 5) of the menstrual cycle. Messenger RNA levels of TRPV2, TRPC4 and TRPC6 were significantly increased (P < 0.01) in the late luteal phase compared with the early luteal phase. Immunocytochemistry experiments showed a positive staining for TRPV2, TRPV4, TRPC6 and TRPM7 in the plasma membrane and in the cytoplasm of primary endometrial stromal cells. Ca(2+)-microfluorimetry revealed significant increases (P < 0.001) in intracellular Ca(2+) levels when stromal cells were incubated with specific activators of TRPV2, TRPV4 and TRPC6. Further functional characterization was performed using whole-cell patch clamp experiments. Taken together, these data provide evidence for the functional activity of TRPV2, TRPV4, TRPC6 and TRPM7 channels in primary stromal cell cultures. LIMITATIONS, REASONS FOR CAUTION Although mRNA levels are detected for TRPV6, TRPC1, TRPC4 and TRPM4, the limited supply of specific antibodies and lack of selective pharmacological agents restricted any additional analysis of these ion channels. WIDER IMPLICATIONS OF THE FINDINGS Embryo implantation is a dynamic developmental process that integrates many signalling molecules into a precisely orchestrated programme. Our findings identified certain members of the TRP superfamily as candidate sensors in the epithelial-stromal crosstalk. These results are very helpful to unravel the signalling cascade required for successful embryo implantation. In addition, this knowledge could lead to new strategies to correct implantation failure and facilitate the development of novel non-hormonal contraceptives. STUDY FUNDING/ COMPETING INTERESTS This work was supported by grants from the Research Foundation-Flanders (G.0856.13N to J.V.), the Research Council of the KU Leuven (OT/13/113 to J.V. and T.D. and PF-TRPLe to T.V.) and by the Planckaert-De Waele fund (to J.V.). K.D.C. and K.H. are funded by the FWO Belgium. None of the authors have a conflict of interest.
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the ca2 activated cation channel TRPM4 is regulated by phosphatidylinositol 4 5 biphosphate
The EMBO Journal, 2006Co-Authors: Frank Mahieu, Rudi Vennekens, Annelies Janssens, Jean Prenen, Thomas VoetsAbstract:Transient receptor potential (TRP) channel, melastatin subfamily (TRPM)4 is a Ca2+-activated monovalent cation channel that depolarizes the plasma membrane and thereby modulates Ca2+ influx through Ca2+-permeable pathways. A typical feature of TRPM4 is its rapid desensitization to intracellular Ca2+ ([Ca2+]i). Here we show that phosphatidylinositol 4,5-biphosphate (PIP2) counteracts desensitization to [Ca2+]i in inside-out patches and rundown of TRPM4 currents in whole-cell patch-clamp experiments. PIP2 shifted the voltage dependence of TRPM4 activation towards negative potentials and increased the channel's Ca2+ sensitivity 100-fold. Conversely, activation of the phospholipase C (PLC)-coupled M1 muscarinic receptor or pharmacological depletion of cellular PIP2 potently inhibited currents through TRPM4. Neutralization of basic residues in a C-terminal pleckstrin homology (PH) domain accelerated TRPM4 current desensitization and strongly attenuated the effect of PIP2, whereas mutations to the C-terminal TRP box and TRP domain had no effect on the PIP2 sensitivity. Our data demonstrate that PIP2 is a strong positive modulator of TRPM4, and implicate the C-terminal PH domain in PIP2 action. PLC-mediated PIP2 breakdown may constitute a physiologically important brake on TRPM4 activity.
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the selectivity filter of the cation channel TRPM4
Journal of Biological Chemistry, 2005Co-Authors: Bernd Nilius, Grzegorz Owsianik, Annelies Janssens, Jean Prenen, Chunbo Wang, Michael X Zhu, Thomas VoetsAbstract:Transient receptor potential channel melastatin subfamily (TRPM) 4 and its close homologue, TRPM5, are the only two members of the large transient receptor potential superfamily of cation channels that are impermeable to Ca2+. In this study, we located the TRPM4 selectivity filter and investigated possible structural elements that render it Ca2+-impermeable. Based on homology with known cation channel pores, we identified an acidic stretch of six amino acids in the loop between transmembrane helices TM5 and TM6 (981EDMDVA986) as a potential selectivity filter. Substitution of this six-amino acid stretch with the selectivity filter of TRPV6 (TIIDGP) resulted in a functional channel that combined the gating hallmarks of TRPM4 (activation by Ca2+, voltage dependence) with TRPV6-like sensitivity to block by extracellular Ca2+ and Mg2+ as well as Ca2+ permeation. Neutralization of Glu981 resulted in a channel with normal permeability properties but a strongly reduced sensitivity to block by intracellular spermine. Neutralization of Asp982 yielded a functional channel that exhibited extremely fast desensitization (τ
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comparison of functional properties of the ca2 activated cation channels TRPM4 and trpm5 from mice
Cell Calcium, 2005Co-Authors: Nina Ullrich, Rudi Vennekens, Jean Prenen, Guy Droogmans, Thomas Voets, Karel Talavera, Bernd NiliusAbstract:Non-selective cation (NSC) channels activated by intracellular Ca2+ ([Ca2+]i) play an important role in Ca2+ signaling and membrane excitability in many cell types. TRPM4 and TRPM5, two Ca2+-activated cation channels of the TRP superfamily, are potential molecular correlates of NSC channels. We compared the functional properties of mouse TRPM4 and TRPM5 heterologously expressed in HEK 293 cells. Dialyzing cells with different Ca2+ concentrations revealed a difference in Ca2+ sensitivity between TRPM4 and TRPM5, with EC50 values of 20.2+/-4.0 microM and 0.70+/-0.1 microM, respectively. Similarly, TRPM5 activated at lower Ca2+ concentration than TRPM4 when [Ca2+]i was raised by UV uncaging of the Ca2+-cage DMNP-EDTA. Current amplitudes of TRPM4 and TRPM5 were not correlated to the rate of changes in [Ca2+]i. The Ca2+ sensitivity of both channels was strongly reduced in inside-out patches, resulting in approximately 10-30 times higher EC50 values than under whole-cell conditions. Currents through TRPM4 and TRPM5 deactivated at negative and activated at positive potentials with similar kinetics. Both channels were equally sensitive to block by intracellular spermine. TRPM4 displayed a 10-fold higher affinity for block by flufenamic acid. Importantly, ATP4- blocked TRPM4 with high affinity (IC50 of 0.8+/-0.1 microM), whereas TRPM5 is insensitive to ATP4- at concentrations up to 1 mM.
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decavanadate modulates gating of TRPM4 cation channels
The Journal of Physiology, 2004Co-Authors: Bernd Nilius, Annelies Janssens, Jean Prenen, Thomas Voets, Guy DroogmansAbstract:We have tested the effects of decavanadate (DV), a compound known to interfere with ATP binding in ATP-dependent transport proteins, on TRPM4, a Ca(2+)-activated, voltage-dependent monovalent cation channel, whose activity is potently blocked by intracellular ATP(4-). Application of micromolar Ca(2+) concentrations to the cytoplasmic side of inside-out patches led to immediate current activation followed by rapid current decay, which can be explained by an at least 30-fold decreased apparent affinity for Ca(2+). Subsequent application of DV (10 microm) strongly affected the voltage-dependent gating of the channel, resulting in large sustained currents over the voltage range between -180 and +140 mV. The effect of DV was half-maximal at a concentration of 1.9 microm. The Ca(2+)- and voltage-dependent gating of the channel was well described by a sequential kinetic scheme in which Ca(2+) binding precedes voltage-dependent gating. The effects of DV could be explained by an action on the voltage-dependent closing step. Surprisingly, DV did not antagonize the effect of ATP(4-) on TRPM4, but caused a nearly 10-fold increase in the sensitivity of the ATP(4-) block. TRPM5, which is the most homologous channel to TRPM4, was not modulated by DV. The effect of DV was lost in a TRPM4 chimera in which the C-terminus was substituted with that of TRPM5. Deletion of a cluster in the C-terminus of TRPM4 containing positively charged amino acid residues with a high homology to part of the decavanadate binding site in SERCA pumps, completely abolished the DV effect but also accelerated desensitization. Deletion of a similar site in the N-terminus had no effects on DV responses. These results indicate that the C-terminus of TRPM4 is critically involved in mediating the DV effects. In conclusion, decavanadate modulates TRPM4, but not TRPM5, by inhibiting voltage-dependent closure of the channel.
Joris Vriens - One of the best experts on this subject based on the ideXlab platform.
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functional expression of transient receptor potential channels in human endometrial stromal cells during the luteal phase of the menstrual cycle
Human Reproduction, 2015Co-Authors: Katrien De Clercq, Thomas Voets, Rieta Van Bree, Karen Peeraer, Katharina Held, Christel Meuleman, Carla Tomassetti, Thomas Dhooghe, Joris VriensAbstract:STUDY QUESTION Are members of the transient receptor potential (TRP) channel superfamily functionally expressed in the human endometrial stroma? SUMMARY ANSWER The Ca(2+)-permeable ion channels TRPV2, TRPV4, TRPC6 and TRPM7 are functionally expressed in primary endometrial stromal cells. WHAT IS KNOWN ALREADY Intercellular communication between epithelial and stromal endometrial cells is required to initiate decidualization, a prerequisite for successful implantation. TRP channels are possible candidates as signal transducers involved in cell-cell communication, but no fingerprint is available of the functional distribution of TRP channels in the human endometrium during the luteal phase of the menstrual cycle. STUDY DESIGN, SIZE, DURATION Endometrial biopsy samples (previously frozen) from patients of reproductive age with regular menstrual cycles, who were undergoing diagnostic laparoscopic surgery for pain and/or infertility, were analysed. Samples were obtained from the menstrual (Days 1-5, n = 3), follicular (Days 6-14, n = 6), early luteal (Days 15-20, n = 5) and late luteal (Days 21-28, n = 5) phases. In addition, a total of 13 patient samples taken during the luteal phase were used to set up primary cell cultures for further experiments. PARTICIPANTS/MATERIALS, SETTING, METHODS Quantitative real-time PCR (qRT-PCR), immunocytochemistry, Fura2-based Ca(2+)-microfluorimetry and whole-cell patch clamp experiments were performed to study the functional expression pattern of TRP channels. Specific pharmacological agents, such as Δ(9)-tetrahydrocannabinol, GSK1016790A and 1-oleoyl-2-acetyl-glycerol, were used to functionally assess the expression of TRPV2, TRPV4 and TRPC6, respectively. MAIN RESULTS AND THE ROLE OF CHANCE Expression of TRPV2, TRPV4, TRPC1, TRPC4, TRPC6, TRPM4 and TRPM7 was detected at the mRNA level in endometrial biopsies (n = 19) and in primary endometrial stromal cell cultures obtained from patients during the luteal phase (n = 5) of the menstrual cycle. Messenger RNA levels of TRPV2, TRPC4 and TRPC6 were significantly increased (P < 0.01) in the late luteal phase compared with the early luteal phase. Immunocytochemistry experiments showed a positive staining for TRPV2, TRPV4, TRPC6 and TRPM7 in the plasma membrane and in the cytoplasm of primary endometrial stromal cells. Ca(2+)-microfluorimetry revealed significant increases (P < 0.001) in intracellular Ca(2+) levels when stromal cells were incubated with specific activators of TRPV2, TRPV4 and TRPC6. Further functional characterization was performed using whole-cell patch clamp experiments. Taken together, these data provide evidence for the functional activity of TRPV2, TRPV4, TRPC6 and TRPM7 channels in primary stromal cell cultures. LIMITATIONS, REASONS FOR CAUTION Although mRNA levels are detected for TRPV6, TRPC1, TRPC4 and TRPM4, the limited supply of specific antibodies and lack of selective pharmacological agents restricted any additional analysis of these ion channels. WIDER IMPLICATIONS OF THE FINDINGS Embryo implantation is a dynamic developmental process that integrates many signalling molecules into a precisely orchestrated programme. Our findings identified certain members of the TRP superfamily as candidate sensors in the epithelial-stromal crosstalk. These results are very helpful to unravel the signalling cascade required for successful embryo implantation. In addition, this knowledge could lead to new strategies to correct implantation failure and facilitate the development of novel non-hormonal contraceptives. STUDY FUNDING/ COMPETING INTERESTS This work was supported by grants from the Research Foundation-Flanders (G.0856.13N to J.V.), the Research Council of the KU Leuven (OT/13/113 to J.V. and T.D. and PF-TRPLe to T.V.) and by the Planckaert-De Waele fund (to J.V.). K.D.C. and K.H. are funded by the FWO Belgium. None of the authors have a conflict of interest.
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Pharmacology of Vanilloid Transient Receptor Potential Cation Channels
Molecular Pharmacology, 2009Co-Authors: Joris Vriens, Giovanni Appendino, Bernd NiliusAbstract:Depending on their primary structure, the 28 mammalian transient receptor potential (TRP) cation channels identified so far can be sorted into 6 subfamilies: TRPC (“Canonical”), TRPV (“Vanilloid”), TRPM (“Melastatin”), TRPP (“Polycystin”), TRPML (“Mucolipin”), and TRPA (“Ankyrin”). The TRPV subfamily (vanilloid receptors) comprises channels critically involved in nociception and thermosensing (TRPV1, TRPV2, TRPV3, and TRPV4), whereas TRPV5 and TRPV6 are involved in renal Ca 2 absorption/reabsorption. Apart from TRPV1, the pharmacology of these channels is still insufficiently known. Furthermore, only few small-molecule ligands for non-TRPV1 vanilloid receptors have been identified, and little is known of their endogenous ligands, resulting in a substantial “orphan” state for these channels. In this review, we summarize the pharmacological properties of members of the TRPV subfamily, highlighting the critical issues and challenges facing their “deorphanization” and clinical exploitation.
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molecular determinants of permeation through the cation channel trpv4
Journal of Biological Chemistry, 2002Co-Authors: Thomas Voets, Joris Vriens, Annelies Janssens, Jean Prenen, Hiroyuki Watanabe, Ulrich Wissenbach, Matthias Bodding, Guy DroogmansAbstract:TRPM6 and its closest relative TRPM7 are members of the Transient Receptor Potential Melastatin (TRPM) subfamily of cation channels and are known to be Mg 2+ permeable. By aligning the sequence of the putative TRPM6 pore with the pore sequences of the other subfamily members, we located in the loop between the fifth and the sixth transmembrane domain, a stretch of amino acids residues, 1028 GEIDVC 1033 , as the potential selectivity filter. Two negatively charged residues, E 1024 (conserved in TRPM6, TRPM7, TRPM1 and TRPM3) and D 1031 (conserved along the entire TRPM subfamily), were identified as important determinants of cation permeation through TRPM6, because neutralization of both residues into an alanine resulted in non-functional channels. Neutralization of E 1029 (conserved in TRPM6, TRPM7, TRPM4 and TRPM5) resulted in channels with increased conductance for Ba 2+ and Zn 2+ , decreased ruthenium red sensitivity and larger pore diameter compared to wild-type TRPM6. Changing the residue I 1030 into methionine, resulted in channels with lower conductance for Ni 2+ , decreased sensitivity to ruthenium red block and reduced pore diameter. Thus, these data demonstrate that amino acid residues E 1024 ,I 1030 and D 1031 are important for channel function and that subtle amino acid variation in the pore region accounts for TRPM6 permeation properties.
