Since LY2109761 TGF-beta inhibitor previous studies indicate that NKp30 plays a crucial role in triggering NK cell-mediated cytotoxicity and induces maturation of iDCs by engaging undefined ligands, we initially analyzed the expression pattern of BAT3 in iDCs. BAT3 was detectable in lysates of Perifosine purchase monocyte-derived iDCs, and released into the extracellular compartment in response to non-lethal heat shock. Moreover, a BAT3- specific ELISA demonstrated that extracellular BAT3 derived from iDCS was specifically recognized by NKp30-Ig, whereas no binding to NKp46-Ig was observed. These data show that iDCs secrete BAT3, that specifically binds to NKp30 and support previous reports suggesting a role for NKp30 but not for NKp46 in the NK-DC cross-talk. Laser Scanner Microscopy revealed that intracellular BAT3 was predominantly detectable in the cell nuclei and also on the cell membrane of heat shocked iDCs. Interestingly, colocalization with surface MHC class I molecules was observed. Taken together, the expression profile of BAT3 in iDCs is in line with a potential NKp30 ligand. The putative BAT3 promoter is glycine and cysteine rich and possesses heat shock elements at position -125 and within the first intron of the ubiquitin-like domain of BAT3. Therefore we investigated whether BAT3 expression was regulated on the transcriptional level. Real-time PCR revealed an increase of BAT3 mRNA in response to a non-lethal heat shock in tumor cells and iDCs. The enhanced mRNA expression in dendritic cells was reproducible, although the induction varied from donor to donor. A consistent correlation of the BAT3 protein release estimated in a BAT3-specific sandwich ELISA and the mRNA increase was demonstrated. We next analyzed the functional role of the NKp30-ligand BAT3 for iDC killing and maturation. A europium release assay with activated NK cells as effector cells and monocyte- derived iDCs as target cells was performed in the presence or absence of BAT3-masking antibodies. Addition of anti-BAT3 inhibited NK cell mediated cytotoxicity significantly when compared to a rabbit control serum ; Fig. 2A). Moreover, down regulation of BAT3 protein upon nucleofection of BAT3 siRNA into iDCs resulted in a decrease of cytotoxicity compared to the control siRNA. The maturation of DCs leads to high expression of MHC class-I molecules and thus protects from NK-mediated cytotoxicity. It is well known that mature DCs are less susceptible to lysis as compared to iDCs. However, subsequent blocking of MHCclass- I molecules on mature DCs with HLA-A, B, C specific mAb lead to efficient lysis by NK cells and this effect could be blocked by adding BAT3 antibodies. The NKp30-dependent NK-DC cross talk also results in iDC maturation.