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Mg2+ Homeostasis: The Mg2+nificent TRPM Chanzymes
Current Biology, Vol. 13, R799–R801, October 14, 2003, ©2003 Elsevier Science Ltd. All rights reserved. DOI 10.1016/j.cub.2003.09.048
Mg2+ Homeostasis: The Mg2+nificent TRPM Chanzymes Craig Montell
TRPM6 and TRPM7 are distinct from all other ion channels in that they are composed of linked channel and protein kinase domains. Recent studies demonstrate that these ‘chanzymes’ are essential for Mg2+ homeostasis, which is critical for human health and cell viability.
The surprises emerging from examination of the TRP family of cation channels show no signs of abating. From analyses of the originally identified member of the TRP superfamily, which functions in Drosophila phototransduction [1], it was expected that the mammalian TRP channels would be activated through pathways coupled to phospholipase C (PLC) and be permeable primarily to Ca2+ and Na+. While mammalian members of the ‘classical or canonical’ TRP subfamily (TRPC) largely conform to these general properties, it turns out that there are three additional subfamilies of TRP channels, TRPV, TRPM and TRPN, which display a much greater diversity of biophysical features and physiological roles than initially envisioned [2]. The largest of the TRP subfamilies, the TRPMs, is especially unusual in that it includes three members which consist of a TRP channel linked to a carboxy-terminal enzyme domain. One of these ‘chanzymes’, referred to as TRPM2, includes an ADP-ribose pyrophosphatase [3,4], while two others, TRPM6 and TRPM7, contain an atypical protein kinase fused to the TRPM channel region (Figure 1) [5–7]. The combination of channel and enzyme domains in these proteins is unique among known ion channels, and raises intriguing questions concerning the functions of the enzyme domains and physiological roles of the chanzymes. Among the three chanzymes, TRPM7 has received the most scrutiny. Nevertheless, the precise function of the kinase domain of TRPM7 remains controversial. A series of recent studies has highlighted the critical role of TRPM6 and TRPM7 in Mg2+ homeostasis and in human health and disease. Mg2+ has long been known to be essential for the activities of myriad enzymes that function in a host of biological processes [8]. At least some of the Mg2+ that enters cells does so through passive mechanisms. Whether there exist ion channels that are required to promote regulated influx of Mg2+ across the plasma membrane has been unclear. According to some studies, Mg2+ slowly enters cells, abrogating the necessity for rapid and regulated influx [8]. Other reports demonstrate that agonists, such as insulin and IGF-1, promote an immediate rise in cellular Mg2+ influx [9]. Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA. E-mail: [email protected]
Dispatch
The discovery that TRPM7 conducts Mg2+ and is required for cell viability suggested that the TRPM7dependent Mg2+ influx is essential. But TRPM7 is not a Mg2+-selective channel; rather, it conducts Ca2+, as well as trace and toxic metals such as Zn2+, Ni2+, Ba2+ and Co2+ [5,6,10]. Nevertheless, strong support for the view that TRPM7 functions in Mg2+ homeostasis has come from the recent demonstration by Schmitz et al. [11] that supplementing the cell-culture medium with Mg2+ rescues the growth arrest and cell lethality that otherwise results from inducibly knocking out TRPM7 expression. Addition of high levels of other cations is ineffective in substituting for the loss of TRPM7. The ability of supplemental Mg2+ to complement the lack of TRPM7 is presumably the result of increased Mg2+ entry through the slow and passive mode of Mg2+ influx. The demonstration that TRPM7 is critical for Mg2+ homeostasis raises the possibility that mutation of TRPM7 may cause human disease as a result of reduced cellular Mg2+ levels. While such an association has not been established for TRPM7, it has recently been shown that mutations in the highly related chanzyme TRPM6 cause hypomagnesemia with secondary hypocalcemia [12,13]. Individuals with this disease display seizures and muscle spasms during infancy, which may lead to death if untreated. Interestingly, the symptoms associated with TRPM6 mutations are ameliorated by supplementation with high doses of Mg2+, consistent with the observation that the cell lethality caused by knocking out TRPM7 is suppressed by addition of high levels of Mg2+ to the culture medium. Whether TRPM6 and TRPM7 functionally interact in vivo has not been addressed, but this seems quite plausible. Previous studies [14–16] of Drosophila and mammalian TRPC proteins established that TRP channels are capable of forming heteromultimeric assemblies, which may alter the biophysical properties of the channels. Such interactions are favored among those channels that are the most related [15]. While TRPM7 is expressed widely, its RNA is found at relatively high levels in the kidneys [6], a tissue that also expresses comparatively high concentrations of TRPM6 RNA [12,13]. A key question concerns the nature of the mechanisms underlying the activation and regulation of TRPM7. In particular, what is the function of the atypical protein kinase domain? According to one report [6], activation of the TRPM7 channel requires ATP, a functional kinase domain and autophosphorylation. It has also been reported that the TRPM7 current is suppressed by increasing concentrations of Mg2+, leading to the suggestion that the stimulatory effect of ATP is a consequence of a drop in free Mg2+ [5]. Several recent observations suggest that the kinase domain is not required for activation. As the free Mg2+ levels are increased, the kinase activity increases, but the channel
Dispatch R800
Mg2+ Extracellular
P
Intracellular
Kinase domain
N
raised from 30 to 1000 µM [18]. While a Mg2+-activated PLC has not been described in other cell types, it may be present in a variety of other cell types including other lymphoid-derived cell lines, such as Jurkat T cells and rat basophilic leukemia cells. These latter cell lines are attractive candidates for expressing a Mg2+-activated PLC, as they contain an endogenous TRPM7-like current, referred to as a Mg2+-inhibited cation channel (MIC) [19,20]. Many additional questions remain concerning TRPM6 and TRPM7. The in vivo substrate for their kinase domains remains to be identified. Furthermore, the physiological mode of activation of TRPM7 is still unresolved. At present, the ion selectivity or any other biophysical properties of TRPM6-dependent currents have not been described. It is intriguing to speculate that, if TRPM6 conducts toxic metals as effectively as TRPM7, those individuals with TRPM6 mutations may display an elevated resistance to exposure to toxic metals such as Ni2+ and Co2+. References
C Current Biology
Figure 1. Domain organization of TRPM6 and TRPM7. The following domains are indicated: transmembrane segments in red, the atypical protein kinase domain in green, the TRP domain [2] in blue and the pore loop (P).
activity is diminished rather than increased [11]. Furthermore, Schmitz et al. [11] found that TRPM7 channels containing point mutations that disrupt the kinase activity are still active, though they render the channel less sensitive to Mg2+ inhibition. Further evidence that these kinase-defective mutants retain channel function in vivo is that their introduction into TRPM7-deficient cells fully rescues the growth arrest and lethality. Conversely, a truncated form of TRPM7, lacking the kinase domain, displays an increased sensitivity to Mg2+ suppression and fails to complement the cell lethality resulting from knockout of TRPM7. However, deletion of the kinase domain does not prevent activation. The basis for the divergent effects on Mg2+ sensitivity resulting from point mutations in or deletion of the kinase domain is unresolved. Nevertheless, the work of Schmitz et al. [11] indicates that the kinase domain is not required for channel activation, but influences sensitivity of the channel to Mg2+ inhibition. In addition to Mg2+ inhibition, TRPM7 has also been shown to be inactivated by hydrolysis of the phosphoinositide lipid PIP2 [17]. This latter finding is intriguing, given that TRPM7 was discovered by one group on the basis of binding to PLC, which cleaves PIP2 [6]. Thus, the question arises as to whether there is a link between the two seemingly disparate modes of channel inhibition: an increase in Mg2+ and hydrolysis of PIP2. An attractive possibility, which needs to be investigated, is that Mg2+ activates the PLC that associates with TRPM7. Such a possibility is strengthened by a study in B lymphocytes demonstrating the presence of phosphoinositide-specific PLC which is activated as the Mg2+ concentration is
1. Montell, C. and Rubin, G.M. (1989). Molecular characterization of the Drosophila trp locus: a putative integral membrane protein required for phototransduction. Neuron 2, 1313-1323. 2. Montell, C. (2001). Physiology, phylogeny and functions of the TRP superfamily of cation channels. Science’s STKE, http://stke.sciencemag.org/cgi/content/full/OC_sigtrans;2001/90/re1. 3. Perraud, A.L., Fleig, A., Dunn, C.A., Bagley, L.A., Launa, P., Schmitz, C., Stokes, A.J., Zhu, Q., Bessman, M.J., Penner, R., et al. (2001). ADP-ribose gating of the calcium-permeable LTRPC2 channel revealed by Nudix motif homology. Nature 411, 595-599. 4. Sano, Y., Inamura, K., Miyake, A., Mochizuki, S., Yokoi, H., Matsushime, H. and Furuichi, K. (2001). Immunocyte Ca2+ influx system mediated by LTRPC2. Science 293, 1327-1330. 5. Nadler, M.J., Hermosura, M.C., Inabe, K., Perraud, A.L., Zhu, Q., Stokes, A.J., Kurosaki, T., Kinet, J.P., Penner, R., Scharenberg, A.M. and Fleig, A. (2001). LTRPC7 is a Mg.ATP-regulated divalent cation channel required for cell viability. Nature 411, 590-595. 6. Runnels, L.W., Yue, L. and Clapham, D.E. (2001). TRP-PLIK, a bifunctional protein with kinase and ion channel activities. Science 291, 1043-1047. 7. Montell, C., Birnbaumer, L. and Flockerzi, V. (2002). The TRP channels, a remarkably functional family. Cell 108, 595-598. 8. Romani, A.M. and Scarpa, A. (2000). Regulation of cellular magnesium. Front. Biosci. 5, D720-D734. 9. Takaya, J., Higashino, H., Miyazaki, R. and Kobayashi, Y. (1998). Effects of insulin and insulin-like growth factor-1 on intracellular magnesium of platelets. Exp. Mol. Pathol. 65, 104-109. 10. Monteilh-Zoller, M.K., Hermosura, M.C., Nadler, M.J., Scharenberg, A.M., Penner, R. and Fleig, A. (2003). TRPM7 provides an ion channel mechanism for cellular entry of trace metal ions. J. Gen. Physiol. 121, 49-60. 11. Schmitz, C., Perraud, A.L., Johnson, C.O., Inabe, K., Smith, M.K., Penner, R., Kurosaki, T., Fleig, A. and Scharenberg, A.M. (2003). Regulation of vertebrate cellular Mg2+ homeostasis by TRPM7. Cell 114, 191-200. 12. Schlingmann, K.P., Weber, S., Peters, M., Niemann Nejsum, L., Vitzthum, H., Klingel, K., Kratz, M., Haddad, E., Ristoff, E., Dinour, D., et al. (2002). Hypomagnesemia with secondary hypocalcemia is caused by mutations in TRPM6, a new member of the TRPM gene family. Nat. Genet. 31, 166-170. 13. Walder, R.Y., Landau, D., Meyer, P., Shalev, H., Tsolia, M., Borochowitz, Z., Boettger, M.B., Beck, G.E., Englehardt R.K., Carmi, R. and Sheffield, V.C. (2002). Mutation of TRPM6 causes familial hypomagnesemia with secondary hypocalcemia. Nat. Genet. 31, 171174. 14. Xu, X-ZS., Li, H-S., Guggino, WB. and Montell, C. (1997). Coassembly of TRP and TRPL produces a distinct store-operated conductance. Cell 89, 1155-1164. 15. Hofmann, T., Schaefer, M., Schultz, G. and Gudermann, T. (2002). Subunit composition of mammalian transient receptor potential channels in living cells. Proc. Natl. Acad. Sci. USA 99, 7461-7466. 16. Strübing, C., Krapivinsky, G., Krapivinsky, L. and Clapham, D.E. (2001). TRPC1 and TRPC5 form a novel cation channel in mammalian brain. Neuron 29, 645-655.
Current Biology R801
17. 18.
19.
20.
Runnels, L.W., Yue, L. and Clapham, D.E. (2002). The TRPM7 channel is inactivated by PIP2 hydrolysis. Nat. Cell Biol. 4, 329-336. Chien, M.M. and Cambier, J.C. (1990). Divalent cation regulation of phosphoinositide metabolism. Naturally occurring B lymphoblasts contain a Mg2+-regulated phosphatidylinositol-specific phospholipase C. J. Biol. Chem. 265, 9201-9207. Kozak, J.A., Kerschbaum, H.H. and Cahalan, M.D. (2002). Distinct properties of CRAC and MIC channels in RBL cells. J. Gen. Physiol. 120, 221-235. Prakriya, M. and Lewis, R.S. (2002). Separation and characterization of currents through store-operated CRAC channels and Mg2+-inhibited cation (MIC) channels. J. Gen. Physiol. 119, 487-507.
Mg2+ Homeostasis: The Mg2+nificent TRPM Chanzymes Craig Montell
TRPM6 and TRPM7 are distinct from all other ion channels in that they are composed of linked channel and protein kinase domains. Recent studies demonstrate that these ‘chanzymes’ are essential for Mg2+ homeostasis, which is critical for human health and cell viability.
The surprises emerging from examination of the TRP family of cation channels show no signs of abating. From analyses of the originally identified member of the TRP superfamily, which functions in Drosophila phototransduction [1], it was expected that the mammalian TRP channels would be activated through pathways coupled to phospholipase C (PLC) and be permeable primarily to Ca2+ and Na+. While mammalian members of the ‘classical or canonical’ TRP subfamily (TRPC) largely conform to these general properties, it turns out that there are three additional subfamilies of TRP channels, TRPV, TRPM and TRPN, which display a much greater diversity of biophysical features and physiological roles than initially envisioned [2]. The largest of the TRP subfamilies, the TRPMs, is especially unusual in that it includes three members which consist of a TRP channel linked to a carboxy-terminal enzyme domain. One of these ‘chanzymes’, referred to as TRPM2, includes an ADP-ribose pyrophosphatase [3,4], while two others, TRPM6 and TRPM7, contain an atypical protein kinase fused to the TRPM channel region (Figure 1) [5–7]. The combination of channel and enzyme domains in these proteins is unique among known ion channels, and raises intriguing questions concerning the functions of the enzyme domains and physiological roles of the chanzymes. Among the three chanzymes, TRPM7 has received the most scrutiny. Nevertheless, the precise function of the kinase domain of TRPM7 remains controversial. A series of recent studies has highlighted the critical role of TRPM6 and TRPM7 in Mg2+ homeostasis and in human health and disease. Mg2+ has long been known to be essential for the activities of myriad enzymes that function in a host of biological processes [8]. At least some of the Mg2+ that enters cells does so through passive mechanisms. Whether there exist ion channels that are required to promote regulated influx of Mg2+ across the plasma membrane has been unclear. According to some studies, Mg2+ slowly enters cells, abrogating the necessity for rapid and regulated influx [8]. Other reports demonstrate that agonists, such as insulin and IGF-1, promote an immediate rise in cellular Mg2+ influx [9]. Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA. E-mail: [email protected]
Dispatch
The discovery that TRPM7 conducts Mg2+ and is required for cell viability suggested that the TRPM7dependent Mg2+ influx is essential. But TRPM7 is not a Mg2+-selective channel; rather, it conducts Ca2+, as well as trace and toxic metals such as Zn2+, Ni2+, Ba2+ and Co2+ [5,6,10]. Nevertheless, strong support for the view that TRPM7 functions in Mg2+ homeostasis has come from the recent demonstration by Schmitz et al. [11] that supplementing the cell-culture medium with Mg2+ rescues the growth arrest and cell lethality that otherwise results from inducibly knocking out TRPM7 expression. Addition of high levels of other cations is ineffective in substituting for the loss of TRPM7. The ability of supplemental Mg2+ to complement the lack of TRPM7 is presumably the result of increased Mg2+ entry through the slow and passive mode of Mg2+ influx. The demonstration that TRPM7 is critical for Mg2+ homeostasis raises the possibility that mutation of TRPM7 may cause human disease as a result of reduced cellular Mg2+ levels. While such an association has not been established for TRPM7, it has recently been shown that mutations in the highly related chanzyme TRPM6 cause hypomagnesemia with secondary hypocalcemia [12,13]. Individuals with this disease display seizures and muscle spasms during infancy, which may lead to death if untreated. Interestingly, the symptoms associated with TRPM6 mutations are ameliorated by supplementation with high doses of Mg2+, consistent with the observation that the cell lethality caused by knocking out TRPM7 is suppressed by addition of high levels of Mg2+ to the culture medium. Whether TRPM6 and TRPM7 functionally interact in vivo has not been addressed, but this seems quite plausible. Previous studies [14–16] of Drosophila and mammalian TRPC proteins established that TRP channels are capable of forming heteromultimeric assemblies, which may alter the biophysical properties of the channels. Such interactions are favored among those channels that are the most related [15]. While TRPM7 is expressed widely, its RNA is found at relatively high levels in the kidneys [6], a tissue that also expresses comparatively high concentrations of TRPM6 RNA [12,13]. A key question concerns the nature of the mechanisms underlying the activation and regulation of TRPM7. In particular, what is the function of the atypical protein kinase domain? According to one report [6], activation of the TRPM7 channel requires ATP, a functional kinase domain and autophosphorylation. It has also been reported that the TRPM7 current is suppressed by increasing concentrations of Mg2+, leading to the suggestion that the stimulatory effect of ATP is a consequence of a drop in free Mg2+ [5]. Several recent observations suggest that the kinase domain is not required for activation. As the free Mg2+ levels are increased, the kinase activity increases, but the channel
Dispatch R800
Mg2+ Extracellular
P
Intracellular
Kinase domain
N
raised from 30 to 1000 µM [18]. While a Mg2+-activated PLC has not been described in other cell types, it may be present in a variety of other cell types including other lymphoid-derived cell lines, such as Jurkat T cells and rat basophilic leukemia cells. These latter cell lines are attractive candidates for expressing a Mg2+-activated PLC, as they contain an endogenous TRPM7-like current, referred to as a Mg2+-inhibited cation channel (MIC) [19,20]. Many additional questions remain concerning TRPM6 and TRPM7. The in vivo substrate for their kinase domains remains to be identified. Furthermore, the physiological mode of activation of TRPM7 is still unresolved. At present, the ion selectivity or any other biophysical properties of TRPM6-dependent currents have not been described. It is intriguing to speculate that, if TRPM6 conducts toxic metals as effectively as TRPM7, those individuals with TRPM6 mutations may display an elevated resistance to exposure to toxic metals such as Ni2+ and Co2+. References
C Current Biology
Figure 1. Domain organization of TRPM6 and TRPM7. The following domains are indicated: transmembrane segments in red, the atypical protein kinase domain in green, the TRP domain [2] in blue and the pore loop (P).
activity is diminished rather than increased [11]. Furthermore, Schmitz et al. [11] found that TRPM7 channels containing point mutations that disrupt the kinase activity are still active, though they render the channel less sensitive to Mg2+ inhibition. Further evidence that these kinase-defective mutants retain channel function in vivo is that their introduction into TRPM7-deficient cells fully rescues the growth arrest and lethality. Conversely, a truncated form of TRPM7, lacking the kinase domain, displays an increased sensitivity to Mg2+ suppression and fails to complement the cell lethality resulting from knockout of TRPM7. However, deletion of the kinase domain does not prevent activation. The basis for the divergent effects on Mg2+ sensitivity resulting from point mutations in or deletion of the kinase domain is unresolved. Nevertheless, the work of Schmitz et al. [11] indicates that the kinase domain is not required for channel activation, but influences sensitivity of the channel to Mg2+ inhibition. In addition to Mg2+ inhibition, TRPM7 has also been shown to be inactivated by hydrolysis of the phosphoinositide lipid PIP2 [17]. This latter finding is intriguing, given that TRPM7 was discovered by one group on the basis of binding to PLC, which cleaves PIP2 [6]. Thus, the question arises as to whether there is a link between the two seemingly disparate modes of channel inhibition: an increase in Mg2+ and hydrolysis of PIP2. An attractive possibility, which needs to be investigated, is that Mg2+ activates the PLC that associates with TRPM7. Such a possibility is strengthened by a study in B lymphocytes demonstrating the presence of phosphoinositide-specific PLC which is activated as the Mg2+ concentration is
1. Montell, C. and Rubin, G.M. (1989). Molecular characterization of the Drosophila trp locus: a putative integral membrane protein required for phototransduction. Neuron 2, 1313-1323. 2. Montell, C. (2001). Physiology, phylogeny and functions of the TRP superfamily of cation channels. Science’s STKE, http://stke.sciencemag.org/cgi/content/full/OC_sigtrans;2001/90/re1. 3. Perraud, A.L., Fleig, A., Dunn, C.A., Bagley, L.A., Launa, P., Schmitz, C., Stokes, A.J., Zhu, Q., Bessman, M.J., Penner, R., et al. (2001). ADP-ribose gating of the calcium-permeable LTRPC2 channel revealed by Nudix motif homology. Nature 411, 595-599. 4. Sano, Y., Inamura, K., Miyake, A., Mochizuki, S., Yokoi, H., Matsushime, H. and Furuichi, K. (2001). Immunocyte Ca2+ influx system mediated by LTRPC2. Science 293, 1327-1330. 5. Nadler, M.J., Hermosura, M.C., Inabe, K., Perraud, A.L., Zhu, Q., Stokes, A.J., Kurosaki, T., Kinet, J.P., Penner, R., Scharenberg, A.M. and Fleig, A. (2001). LTRPC7 is a Mg.ATP-regulated divalent cation channel required for cell viability. Nature 411, 590-595. 6. Runnels, L.W., Yue, L. and Clapham, D.E. (2001). TRP-PLIK, a bifunctional protein with kinase and ion channel activities. Science 291, 1043-1047. 7. Montell, C., Birnbaumer, L. and Flockerzi, V. (2002). The TRP channels, a remarkably functional family. Cell 108, 595-598. 8. Romani, A.M. and Scarpa, A. (2000). Regulation of cellular magnesium. Front. Biosci. 5, D720-D734. 9. Takaya, J., Higashino, H., Miyazaki, R. and Kobayashi, Y. (1998). Effects of insulin and insulin-like growth factor-1 on intracellular magnesium of platelets. Exp. Mol. Pathol. 65, 104-109. 10. Monteilh-Zoller, M.K., Hermosura, M.C., Nadler, M.J., Scharenberg, A.M., Penner, R. and Fleig, A. (2003). TRPM7 provides an ion channel mechanism for cellular entry of trace metal ions. J. Gen. Physiol. 121, 49-60. 11. Schmitz, C., Perraud, A.L., Johnson, C.O., Inabe, K., Smith, M.K., Penner, R., Kurosaki, T., Fleig, A. and Scharenberg, A.M. (2003). Regulation of vertebrate cellular Mg2+ homeostasis by TRPM7. Cell 114, 191-200. 12. Schlingmann, K.P., Weber, S., Peters, M., Niemann Nejsum, L., Vitzthum, H., Klingel, K., Kratz, M., Haddad, E., Ristoff, E., Dinour, D., et al. (2002). Hypomagnesemia with secondary hypocalcemia is caused by mutations in TRPM6, a new member of the TRPM gene family. Nat. Genet. 31, 166-170. 13. Walder, R.Y., Landau, D., Meyer, P., Shalev, H., Tsolia, M., Borochowitz, Z., Boettger, M.B., Beck, G.E., Englehardt R.K., Carmi, R. and Sheffield, V.C. (2002). Mutation of TRPM6 causes familial hypomagnesemia with secondary hypocalcemia. Nat. Genet. 31, 171174. 14. Xu, X-ZS., Li, H-S., Guggino, WB. and Montell, C. (1997). Coassembly of TRP and TRPL produces a distinct store-operated conductance. Cell 89, 1155-1164. 15. Hofmann, T., Schaefer, M., Schultz, G. and Gudermann, T. (2002). Subunit composition of mammalian transient receptor potential channels in living cells. Proc. Natl. Acad. Sci. USA 99, 7461-7466. 16. Strübing, C., Krapivinsky, G., Krapivinsky, L. and Clapham, D.E. (2001). TRPC1 and TRPC5 form a novel cation channel in mammalian brain. Neuron 29, 645-655.
Current Biology R801
17. 18.
19.
20.
Runnels, L.W., Yue, L. and Clapham, D.E. (2002). The TRPM7 channel is inactivated by PIP2 hydrolysis. Nat. Cell Biol. 4, 329-336. Chien, M.M. and Cambier, J.C. (1990). Divalent cation regulation of phosphoinositide metabolism. Naturally occurring B lymphoblasts contain a Mg2+-regulated phosphatidylinositol-specific phospholipase C. J. Biol. Chem. 265, 9201-9207. Kozak, J.A., Kerschbaum, H.H. and Cahalan, M.D. (2002). Distinct properties of CRAC and MIC channels in RBL cells. J. Gen. Physiol. 120, 221-235. Prakriya, M. and Lewis, R.S. (2002). Separation and characterization of currents through store-operated CRAC channels and Mg2+-inhibited cation (MIC) channels. J. Gen. Physiol. 119, 487-507.