Hydrogen peroxide removes TRPM4 current desensitization conferring increased vulnerability to necrotic cell death.
Necrosis is associated with an increase in plasma membrane permeability, cell swelling, and loss of membrane integrity with subsequent release of cytoplasmic constituents. Severe redox imbalance by overproduction of reactive oxygen species is one of the main causes of necrosis. Here we demonstrate that H(2)O(2) induces a sustained activity of TRPM4, a Ca(2+)-activated, Ca(2+)-impermeant nonselective cation channel resulting in an increased vulnerability to cell death. In HEK 293 cells overexpressing TRPM4, H(2)O(2) was found to eliminate in a dose-dependent manner TRPM4 desensitization. Site-directed mutagenesis experiments revealed that the Cys(1093) residue is crucial for the H(2)O(2)-mediated loss of desensitization. In HeLa cells, which endogenously express TRPM4, H(2)O(2) elicited necrosis as well as apoptosis. H(2)O(2)-mediated necrosis but not apoptosis was abolished by replacement of external Na(+) ions with sucrose or the non-permeant cation N-methyl-d-glucamine and by knocking down TRPM4 with a shRNA directed against TRPM4. Conversely, transient overexpression of TRPM4 in HeLa cells in which TRPM4 was previously silenced re-established vulnerability to H(2)O(2)-induced necrotic cell death. In addition, HeLa cells exposed to H(2)O(2) displayed an irreversible loss of membrane potential, which was prevented by TRPM4 knockdown.
Simon
Felipe
F
Centro de Estudios Moleculares de la Célula and Instituto de Ciencias Biomédicas, Facultad de Medicina, Universidad de Chile, Av Independencia 1027, Independencia 838-0543, Santiago, Chile.
Leiva-Salcedo
Elías
E
Armisén
Ricardo
R
Riveros
Ana
A
Cerda
Oscar
O
Varela
Diego
D
Eguiguren
Ana Luisa
AL
Olivero
Pablo
P
Stutzin
Andrés
A
D020816:Amino Acid Motifs; D017209:Apoptosis; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism*; D008564:Membrane Potentials; D009336:Necrosis / Q000235:genetics / Q000378:metabolism*; D050053:TRPM Cation Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D050053:TRPM Cation Channels; C437888:TRPM4 protein, human; D006861:Hydrogen Peroxide