Akt2-and ETS1-dependent IP3 receptor 2 expression in dendritic cell migration

Abstract

Background/Aims: The protein kinase Akt2/PKBβ is a known regulator of macrophage and dendritic cell (DC) migration. The mechanisms linking Akt2 activity to migration remained, however, elusive. DC migration is governed by Ca 2+ signaling. We thus explored whether Akt2 regulates DC Ca 2+ signaling. Methods: DCs were derived from bone marrow of Akt2-deficient mice (akt2 -/- ) and their wild type littermates (akt2 +/+ ). DC maturation was induced by lipopolysaccharides (LPS) and evaluated by flow cytometry. Cytosolic Ca 2+ concentration was determined by Fura-2 fluorescence, channel activity by whole cell recording, transcript levels by RT-PCR, migration utilizing transwells. Results: Upon maturation, chemokine CCL21 stimulated migration of akt2 +/+ but not akt2 -/- DCs. CCL21-induced increase in cytosolic Ca 2+ concentration, thapsigargin-induced release of Ca 2+ from intracellular stores with subsequent store-operated Ca 2+ entry (SOCE), ATP-induced inositol 1,4,5-trisphosphate (IP 3 )-dependent Ca 2+ release as well as Ca 2+ release-activated Ca 2+ (CRAC) channel activity were all significantly lower in mature akt2 -/- than in mature akt2 +/+ DCs. Transcript levels of IP 3 receptor IP 3 R2 and of IP 3 R2 regulating transcription factor ETS1 were significantly higher in akt2 +/+ than in akt2 -/- DCs prior to maturation and were upregulated by LPS stimulation (1h) in akt2 +/+ and to a lower extent in akt2 -/- DCs. Following maturation, protein abundance of IP 3 R2 and ETS1 were similarly higher in akt2 +/+ than in akt2 -/- DCs. The IP 3 R inhibitor Xestospongin C significantly decreased CCL21-induced migration of akt2 +/+ DCs and abrogated the differences between genotypes. Finally, knock-down of ETS1 with siRNA decreased IP 3 R2 mRNA abundance, thapsigargin-and ATP-induced Ca 2+ release, SOCE and CRAC channel activation, as well as DC migration. Conclusion: Akt2 upregulates DC migration at least in part by ETS1-dependent stimulation of IP 3 R2 transcription. © 2014 S. Karger AG, Basel.