The serum- and glucocorticoid-inducible kinase 1 (SGK1) influences platelet calcium signaling and function by regulation of Orai1 expression in megakaryocytes.
Platelets are activated on increase of cytosolic Ca2+ activity ([Ca2+](i)), accomplished by store-operated Ca2+ entry (SOCE) involving the pore-forming ion channel subunit Orai1. Here, we show, for the first time, that the serum- and glucocorticoid-inducible kinase 1 (SGK1) is expressed in platelets and megakaryocytes. SOCE and agonist-induced [Ca2+](i) increase are significantly blunted in platelets from SGK1 knockout mice (sgk1(-/-)). Similarly, Ca2+ -dependent degranulation, integrin α(IIb)β3 activation, phosphatidylserine exposure, aggregation, and in vitro thrombus formation were significantly impaired in sgk1(-/-) platelets, whereas tail bleeding time was not significantly enhanced. Platelet and megakaryocyte Orai1 transcript levels and membrane protein abundance were significantly reduced in sgk1(-/-) mice. In human megakaryoblastic cells (MEG-01), transfection with constitutively active (S422D)SGK1 but not with inactive (K127N)SGK1 significantly enhanced Orai1 expression and SOCE, while effects reversed by the SGK1 inhibitor GSK650394 (1μM). Transfection of MEG-01 cells with (S422D)SGK1 significantly increased phosphorylation of IκB kinase α/β and IκBα resulting in nuclear translocation of NF-κB subunit p65. Treatment of (S422D)SGK1-transfected MEG-01 cells with the IκB kinase inhibitor BMS-345541 (10μM) abolished SGK1-induced increase of Orai1 expression and SOCE. The present observations unravel SGK1 as novel regulator of platelet function, effective at least in part by NF-κB-dependent transcriptional up-regulation of Orai1 in megakaryocytes and increasing platelet SOCE.
Department of Physiology, University of Tübingen, Tübingen, Germany.
Oliver
O
Borst
Eva-Maria
EM
Schmidt
Patrick
P
Münzer
Tanja
T
Schönberger
Syeda T
ST
Towhid
Margitta
M
Elvers
Christina
C
Leibrock
Evi
E
Schmid
Anja
A
Eylenstein
Dietmar
D
Kuhl
Andreas E
AE
May
Meinrad
M
Gawaz
Florian
F
Lang
D000818:Animals; D001760:Bleeding Time; D001792:Blood Platelets / Q000378:metabolism*; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000235:genetics / Q000378:metabolism*; D020013:Calcium Signaling; D002478:Cells, Cultured; D005260:Female; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006801:Humans; D017874:Immediate-Early Proteins / Q000502:physiology*; D007124:Immunoenzyme Techniques; D007947:Leukemia, Megakaryoblastic, Acute / Q000378:metabolism / Q000473:pathology; D008297:Male; D008533:Megakaryocytes / Q000166:cytology / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism; D000071740:ORAI1 Protein; D010766:Phosphorylation; D010974:Platelet Aggregation; D017346:Protein Serine-Threonine Kinases / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013927:Thrombosis / Q000209:etiology / Q000378:metabolism / Q000473:pathology; D000099184:Serum-Glucocorticoid Regulated Kinases
D002118:Calcium; D015220:Calcium Channels; D017874:Immediate-Early Proteins; D016328:NF-kappa B; D000071740:ORAI1 Protein; D017346:Protein Serine-Threonine Kinases; D012333:RNA, Messenger; D000099184:Serum-Glucocorticoid Regulated Kinases; C520430:Orai1 protein, mouse