tailieunhanh - Báo cáo khoa học: Modulation of Ca2+ entry and plasma membrane potential by human TRPM4b

TRPM4b is a Ca 2+ -activated, voltage-dependent monovalent cation chan-nel that has been shown to act as a negative regulator of Ca 2+ entry and to be involved in the generation of oscillations of Ca 2+ influx in Jurkat T-lymphocytes. Transient overexpression of TRPM4b as an enhanced green fluorescence fusion protein in human embryonic kidney (HEK) cells resulted in its localization in the plasma membrane, as demonstrated by confocal fluorescence microscopy. | ễFEBS Journal Modulation of Ca2 entry and plasma membrane potential by human TRPM4b Ralf Fliegert1 Gunter Glassmeier2 Frederike Schmid1 Kerstin Cornils1 Selda Genisyuerek1 Angelika Harneit1 Jurgen R. Schwarz2 and Andreas H. Guse1 1 Calcium Signalling Group Institute of Biochemistry and Molecular Biology I Cellular SignalTransduction Center of ExperimentalMedicine University MedicalCenter Hamburg-Eppendorf Hamburg Germany 2 Institute of Applied Physiology Center of ExperimentalMedicine University MedicalCenter Hamburg-Eppendorf Hamburg Germany Keywords Ca2 calcium membrane potential TRPM4 TRP channel Correspondence A. H. Guse Institute of Biochemistry and Molecular Biology I Cellular Signal Transduction Center of Experimental Medicine University MedicalCenter Hamburg-Eppendorf Martinistr. 52 D-20246 Hamburg Germany Fax 49 40 42803-9880 Tel 49 40 42803 2828 E-mail guse@ Received 31 August 2006 revised 20 November 2006 accepted 22 November 2006 doi TRPM4b is a Ca2 -activated voltage-dependent monovalent cation channel that has been shown to act as a negative regulator of Ca2 entry and to be involved in the generation of oscillations of Ca2 influx in Jurkat T-lymphocytes. Transient overexpression of TRPM4b as an enhanced green fluorescence fusion protein in human embryonic kidney HEK cells resulted in its localization in the plasma membrane as demonstrated by confocal fluorescence microscopy. The functionality and plasma membrane localization of overexpressed TRPM4b was confirmed by induction of Ca2 -dependent inward and outward currents in whole cell patch clamp recordings. HEK-293 cells stably overexpressing TRPM4b showed higher ionomycin-activated Ca2 influx than wild-type cells. In addition analysis of the membrane potential using the potentiometric dye bis- 1 3-dibutylbarbituric acid -trimethine oxonol and by current clamp experiments in the perforated patch configuration revealed a faster initial depolarization .