Ca2+ but not H2O2 modulates GRE-element activation by the human mineralocorticoid receptor in HEK cells.
The mineralocorticcoid receptor (MR) plays an important role in salt and water homeostasis as well as during cardiovascular and renal fibrosis but little is known regarding its modulation by other signaling pathways. To investigate a possible modulation under controlled conditions we used human embryonic kidney (HEK) cells (devoid of endogenous MR) transfected with the human MR and measured transactivation with a GRE-SEAP-reporter construct. MR was compared to the glucocorticoid receptor (GR) as well as to MR lacking the N-terminal domains AB (MR(CDEF)). Chelation of cytosolic Ca2+ enhanced MR activity and SGK1-expression, whereas elevation of cytosolic Ca2+ with ionomycin or thapsigargin reduced MR activity. GR activity was not affected by ionomycin or thapsigargin. MR(CDEF) activity was not affected by chelation or elevation of cytosolic Ca2+. Inhibition of ERK1/2 activation by U0126 or activation of PKA by cAMP, previously shown to modulate MR and GR activity, did not affect MR(CDEF) activity either. H2O2<500micromol/l did not affect basal nor hormone-induced reporter activity. Higher concentrations exerted the same relative inhibitory effect on GRE-SEAP-activity under basal conditions as in the presence of aldosterone-stimulated MR and elicited cytotoxic effects. Our data indicate that the genomic function of MR can be modulated by cytosolic Ca2+, PKA and ERK1/2 via an interaction with the AB-domain. H2O2 seems not to affect relative MR activity directly under our experimental conditions.
Pfau
Anja
A
Physiologisches Institut, Universität Würzburg, Röntgenring 9, 97070 Würzburg, Germany.
Grossmann
Claudia
C
Freudinger
Ruth
R
Mildenberger
Sigrid
S
Benesic
Andreas
A
Gekle
Michael
M
D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects* / Q000502:physiology; D002460:Cell Line; D002614:Chelating Agents / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004622:Embryo, Mammalian / Q000166:cytology / Q000378:metabolism; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D016877:Oxidants / Q000494:pharmacology*; D011494:Protein Kinases / Q000378:metabolism; D011965:Receptors, Glucocorticoid / Q000096:biosynthesis; D018161:Receptors, Mineralocorticoid / Q000096:biosynthesis*; D020218:Response Elements / Q000502:physiology*
D002614:Chelating Agents; D004791:Enzyme Inhibitors; D007476:Ionophores; D016877:Oxidants; D011965:Receptors, Glucocorticoid; D018161:Receptors, Mineralocorticoid; D015759:Ionomycin; D006861:Hydrogen Peroxide; D011494:Protein Kinases; D002118:Calcium