Depletion of plasma membrane-associated phosphoinositides mimics inhibition of TRPM7 channels by cytosolic Mg, spermine, and pH.

Transient receptor potential cation channel subfamily M member 7 (TRPM7) is an ion channel/protein kinase belonging to the TRP melastatin and eEF2 kinase families. Under physiological conditions, most native TRPM7 channels are inhibited by cytoplasmic Mg, protons, and polyamines. Currents through these channels (I) are robustly potentiated when the cell interior is exchanged with low Mg-containing buffers. I is also potentiated by phosphatidyl inositol bisphosphate (PI(4,5)P) and suppressed by its hydrolysis. Here we characterized internal Mg- and pH-mediated inhibition of TRPM7 channels in HEK293 cells overexpressing WT voltage-sensing phospholipid phosphatase (VSP) or its catalytically inactive variant VSP-C363S. VSP-mediated depletion of membrane phosphoinositides significantly increased channel sensitivity to Mg and pH. Proton concentrations that were too low to inhibit I when the VSP-C363S variant was expressed (pH 8.2) became inhibitory in WT VSP-expressing cells. At pH 6.5, protons inhibited I both in WT and VSP C363S-expressing cells but with a faster time course in the WT VSP-expressing cells. Inhibition by 150 μm Mg was also significantly faster in the WT VSP-expressing cells. Cellular PI(4,5)P depletion increased the sensitivity of TRPM7 channels to the inhibitor 2-aminoethyl diphenyl borinate, which acidifies the cytosol. Single substitutions at Ser-1107 of TRPM7, reducing its sensitivity to Mg, also decreased its inhibition by spermine and acidic pH. Furthermore, these channel variants were markedly less sensitive to VSP-mediated PI(4,5)P depletion than the WT. We conclude that the internal Mg-, polyamine-, and pH-mediated inhibition of TRPM7 channels is not direct but, rather, reflects electrostatic screening and resultant disruption of PI(4,5)P-channel interactions.