A failure to express activating receptors, or vital parts of the signaling machinery activated by these receptors, would clarify this defect. Our microarray evaluation uncovered decreased expression of genes encoding the activating receptor NKp46 and various Ly49 receptors as well as proteins involved in signal transduction by these receptors. We confirmed the decreased expression of NKp46, Ly49D and Ly49H on BM and splenic mNK cells from Ets1 mice by movement cytometry. These activating NKRs have been also decreased on Ets1 mNK cells isolated from mixed BM chimeras indicating that this alteration was cell intrinsic. For that reason, the failure of Ets1 mNK cells to destroy NK cell targets might possibly be, in portion, a consequence of decreased expression of several activating NKRs. In contrast to NKp46, Ly49D and Ly49H the activating receptors NK1. 1 and NKG2D were expressed appropriately on BM and splenic mNK cells suggesting that these receptors are available for NK cell target recognition. To determine whether these receptors were functional, we examined the ability of NK1. 1 or NKG2D cross linking to induce degranulation, as measured by surface CD107a. As expected, cross linking of NK1.
1 or NKG2D resulted in elevated CD107a compared to IgG on Ets1 mNK cells. In contrast, NKG2D stimulation of Ets1 mNK cells didn’t induce CD107a above that observed with IgG, while CD107a was larger on IgG stimulated Ets1 mNK cells compared to Ets mNK cells. Cross linking of NK1. 1 on Ets1 mNK cells improved surface CD107a selleck chemical SB 525334 but the frequency of CD107a cells was lower than observed on Ets1 cells. In contrast, CD107a was efficiently induced by phorbol myristate acetate ionomycin in the two Ets1 and Ets1 mNK cells. These observations indicated that Ets1 mNK cells had been intrinsically defective within their ability to degranulate in response to activating NKR stimulation. In contrast, interferon production was not as severely impacted, while cross linking of NKG2D did not bring about a substantial accumulation of IFN at this time stage in Ets1 or Ets1 mNK cells. The reduced expression of several activating NKRs along with the impaired exocytosis function in Ets1 mNK cells is adequate to explain the decreased cytolytic perform of these cells.
On top of that on the ETS1 dependent genes we mentioned that many different genes connected to NK cell activation were improved in Ets1 mNK cells. Gzmb and Prf1 mRNA, encoding the cytolytic proteins Granzyme B and Perforin, had been greater as have been mRNAs encoding the serine protease inhibitors Serpinb6a and Serpinb9b. We confirmed that Gzmb mRNA was increased in Ets1 mNK cells by qPCR. Nfil3 mRNA, encoding Tubastatin A a cytokine responsive transcription element. was also increased in Ets1 mNK cells at the same time as in proNK cells. Interestingly, Ikzf2 mRNA, which encodes HELIOS, whose increased expression contributes to hyper responsiveness in No mice. was enhanced in Ets1 mNK cells.