Detailed examination of Mg 2 + and pH sensitivity of human TRPM 7 1 channels

24 TRPM7 channel-kinase is a protein highly expressed in cells of hematopoietic lineage, 25 such as lymphocytes. Studies performed in native and heterologous expression 26 systems have shown that TRPM7 forms non-selective cation channels functional in the 27 plasma membrane and activated upon depletion of cellular Mg. In addition to internal 28 Mg, cytosolic pH and the phospholipid PI(4,5)P2 are potent physiological regulators of 29 this channel: protons inhibit, while PI(4,5)P2 is required for TRPM7 channel activity. 30 These channels are also inhibited from inside by other metal cations and polyamines. 31 While the regulation of TRPM7 channels by internal metal ions, acidic pH and PI(4,5)P2 32 is voltage-independent, extracellular metal cations and polyamines block voltage33 dependently at micromolar concentrations and appear to occupy a distinct blocking site. 34 In the current study we investigated Mgiand pHi-dependence of native TRPM7 35 currents using whole-cell patch clamp electrophysiology in human Jurkat T lymphocytes 36 and HEK293 cells. 37 Our main findings are: 1) Mg inhibition involves not one but two separate sites 38 of high (~10 μM) and low (~165 μM) affinity; 2) while sharing certain characteristics with 39 Mg inhibition, protons most likely inhibit through one inhibitory site, corresponding to 40 the low affinity Mg site, with an estimated IC50 of pH 6.3. Additionally, we present data 41 on amplitude distribution of pre-activated TRPM7 currents in Jurkat T lymphocytes in 42 the absence of prior Mg or proton depletion. 43