Molecular Determinants of Mg2+ and Ca2+ Permeability and pH Sensitivity in TRPM6 and TRPM7

Abstract

The channel kinases TRPM6 and TRPM7 have recently been discovered to play important roles in Mg 2؉ and Ca 2؉ homeostasis, which is critical to both human health and cell viability. However, the molecular basis underlying these channels' unique Mg 2؉ and Ca 2؉ permeability and pH sensitivity remains unknown. Here we have created a series of amino acid substitutions in the putative pore of TRPM7 to evaluate the origin of the permeability of the channel and its regulation by pH. Two mutants of TRPM7, E1047Q and E1052Q, produced dramatic changes in channel properties. The I-V relations of E1052Q and E1047Q were significantly different from WT TRPM7, with the inward currents of 8-and 12-fold larger than TRPM7, respectively. permeation, rendering TRPM7 a monovalent selective channel. In addition, the ability of protons to potentiate inward currents was lost in E1047Q, indicating that E1047 is critical to Ca 2؉ and Mg 2؉ permeability of TRPM7, and its pH sensitivity. Mutation of the corresponding residues in the pore of TRPM6, E1024Q and E1029Q, produced nearly identical changes to the channel properties of TRPM6. Our results indicate that these two glutamates are key determinants of both channels' divalent selectivity and pH sensitivity. These findings reveal the molecular mechanisms underpinning physiological/pathological functions of TRPM6 and TRPM7, and will extend our understanding of the pore structures of TRPM channels.TRPM6 and TRPM7 belong to the TRP channel superfamily (1-5) and are distinguished from other known ion channels by virtue of having both ion channel and protein kinase activities (6 -11). In addition, TRPM6 and TRPM7 uniquely exhibit strong outward rectification, permeation to Ca 2ϩ , Mg 2ϩ , monovalent cations, and a wide array of trace metals (6 -8, 11, 12 (8,20,21,23), whereas TRPM7 is ubiquitously expressed, with highest expression in the kidney and heart (5, 6). In addition to these channels' regulation of Mg 2ϩ homeostasis, several studies have suggested multiple cellular and physiology functions for TRPM7, including anoxic neuronal death (24), cell adhesion and actomyosin contractility (25, 26), and skeletogenesis (27). Although the mechanisms by which TRPM6 and TRPM7 exert their physiological and/or pathological functions are not yet completely understood, it is clear that permeation of Ca 2ϩ and Mg 2ϩ contributes substantially to the known functions of these channels (7, 20 -22, 24, 25, 27). Moreover, a recent study demonstrated that the sensitivity of TRPM7 to external pH may contribute to controlling neurotransmitter release (28). Therefore, it is essential to understand the molecular mechanisms underlying the Ca 2ϩ and Mg 2ϩ permeability of TRPM6 and TRPM7, as well as their sensitivities to changes in pH.The aim of the present study was to identify the amino acid residues that determine Mg 2ϩ and Ca 2ϩ permeation of TRPM6 and TRPM7. We previously demonstrated that external protons significantly enhance TRPM6 and TRPM7 inward currents (11,19) by decreasing the divalent affinity to t...

Mechanism of Urinary Calcium Regulation by Urinary Magnesium and pH

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Hypomagnesemia Tetany in Cattle

“…A fundamental relationship cannot be inferred from these studies, but they obviously indicate the significance of pH and Mg solubility in the rumen relating to Mg absorption. Besides, the putative Mg channel (TRPM7) exhibits a strong relationship to pH and is activated a pH of <7.0 in patch clamp studies [58].…”

New TRPM6 missense mutations linked to hypomagnesemia with secondary hypocalcemia

“…In contrast, the Q 1663 R variant (SNP rs55679040) displayed current amplitudes comparable to WT (Figure 2). Q 1663 R biophysical properties do not differ from those of WT TRPM6 To investigate whether the Q 1663 R sequence variant showed different functional behaviour with respect to WT TRPM6, we examined divalent-mediated block as previously shown by Li et al 28 and the response to epidermal growth factor (EGF) application.…”