The main objective of this study is to determine the mechanism by which human NK cells activated with cytokines undergo apoptosis when they encounter activating ligands on target cells. We observed that human primary NK cells that were stimulated in vitro with the cytokines IL-2 and IL-15 showed extensive potential to undergo apoptosis when their activation receptors were cross linked. In order to study this ?activation induced cell death? in NK cells further, we drew incite from our earlier microarray study. Our microarray data on genes that are modulated upon stimulation of human NK cells in culture with IL-2 showed that the anti-apoptotic molecule, TOSO was down regulated in cells cultured with IL-2 by 5 fold. TOSO, also called as Fas Apoptotic Inhibitory Molecule 3 (FAIM3) is a member of immunoglobulin gene superfamily and it inhibits Fas mediated apoptosis by binding to FADD at its C-terminus and blocking further activation of the cell death pathway. TOSO is expressed mainly in certain classes of immune cells. However, the role of TOSO in the physiological context of NK cell activation and their function is not well known. This prompted us to look into the role of TOSO in activation induced cell death in NK cells. We also confirmed the fact that TOSO levels were down regulated and quantitated the relative levels of TOSO expression using real time PCR assays. Further, we found that human NK cells when stimulated with IL-2 expressed high levels of the death receptor Fas. In order to study the involvement of TOSO in this phenomenon, we are in the process of cloning the human TOSO gene and over expressing it in primary human NK cells. We are also planning to establish a cell line model for stable expression of TOSO.
