Background:

Akt2/PKBβ is known to be required for macrophage and dendritic cell (DC) migration, though the involved mechanisms remained undefined. On the other hand, DC migration is governed by Ca²⁺ signalling. In the present project, we addressed the mechanisms of Akt2-dependent DC migration by studying the possible involvement of Akt2 in Ca²⁺ signalling.

Methods/Results:

DCs were derived from the bone marrow of Akt2-deficient mice ( akt2^-/- ) and their wild type littermates ( akt2^+/+ ) and their maturation was induced by lipopolysaccharides (LPS). Chemokine CCL21 (25 ng/ml)-induced migration of akt2^-/- DCs was impaired compared to akt2^+/+ DCs. In Ca²⁺ imaging experiments, CCL21 (75 ng/ml)-triggered increase in cytosolic Ca²⁺ concentrations ([Ca²⁺]_i) was also significantly diminished in akt2^-/- DCs. Moreover, release of Ca²⁺ from intracellular stores induced by thapsigargin (1 µM) and the following Ca²⁺ entry were reduced in akt2^-/- DCs. Stimulation of P2Y receptors with ATP (100 µM) results in IP₃-dependent Ca²⁺ release, which was also significantly decreased in akt2^-/- DCs. Analysis of the expression of IP₃ receptor isoforms in akt2^+/+ DCs and akt2^-/- DCs by RT-PCR and western blot revealed a significantly impaired transcript and protein abundance of IP₃R2 in akt2^-/-DCs. Expression of transcription factor ETS1, which is known to upregulate the IP₃R2 transcription, was significantly lower in akt2^-/- DCs than in akt2^+/+ DCs. Finally, DC migration was sensitive to the IP3R inhibitor Xestospongin C, which significantly decreased CCL21-induced migration of akt2^+/+ DCs and abrogated the differences between genotypes.

Conclusions:

Akt2 upregulates IP₃R2 transcription in DCs, presumably by stimulating the expression of ETS-1 and this effect may underlie the reduced migration of Akt2-deficient DCs.