We have recently identified and cloned a novel 120/130 kDa protein, termed FYB that is restricted in its expression to cells of hematopoietic origin. FYB is a substrate in TcRzeta/CD3 generated signaling pathways and expression of FYB in T-cells enhances IL-2 expression in response to TcRzeta/CD3 and TcRzeta/CD3-CD28 stimulation. FYB is physically associated in a complex with SLP-76 and the NCK adaptor molecule. FYB binds to SLP-76 via the SH2 domain of SLP-76, suggesting that it functions downstream of the ZAP-70/Vav/SLP-76 pathway that regulates NFAT activity and IL-2 expression. In turn, the association with NCK suggests a functional interaction with signaling cascades generated by the protooncogenes p21rac and cdc42, members of the Rho family of small GTPases. Signals generated by p21rac/cdc-42 are believed to participate in the activation of the AP-1 transcription factor, which regulates IL-2 promoter activity. Formation of a AP-1/NFAT complex in turn is essential for maximal IL-2 expression. Thus, FYB appears to be ideally suited to provide a bridge between the primary activation signals generated when T-cells encounter antigen, and those initiated by co-receptors, such as CD28. In addition, given that FYB is prominently phosphorylated in thymocytes the molecule is likely to play a key role in the selection mechanism in the thymus.
In Aims 1 and 2 we will attempt to investigate at the molecular and functional levels the role that the interactions of p120/130 with SLP-76 and NCK play in the activation of T-cells, IL-2 expression and IL-2 promoter activity. Lastly, in Aim 3 we propose to generate p120/130 deficient knock-out mice for an analysis of p120/130 in T-cell activation and hematopoiesis, in particular, thymic development. Given that most proteins with a similar restricted pattern of expression to cells of hematopoietic origin (e.g. lck, vav, RAG-2, etc) have been shown to control key steps in differentiation, this approach is also likely to provide important information on the role of p123/130 I one of the major tyrosine phosphorylation cascade in T-cell development and other stages of hematopoietic differentiation.
