LAT (linker for activation of T cells) is a transmembrane adaptor protein that plays an essential role in TCR-mediated signaling and thymocyte development. Because LAT-deficient mice have an early block in thymocyte development, we utilized an inducible system to delete LAT in primary T cells to study LAT function in T cell activation, homeostasis, and survival. Deletion of LAT caused primary T cells to become unresponsive to stimulation from the TCR and impaired T cell homeostatic proliferation and long term survival. Furthermore, deletion of LAT led to reduced expression of Foxp3, CTLA-4, and CD25 in T reg cells and impaired their function. Consequently, mice with LAT deleted developed a lymphoproliferative syndrome similar to that in LATY136F mice, although less severe. Our data implicate that LAT has positive and negative roles in the regulation of mature T cells.Upon engagement of the TCR (T cell receptor), 3 LAT (linker for activation of T cells) is phosphorylated at multiple tyrosine residues by ZAP-70 kinase (1) and functions as a protein scaffold to assemble a large membrane-tethered signalosome (1-3). LAT binding of Gads leads to the membrane recruitment of SLP-76, which in turn interacts with Itk, as well as Vav1 and PLC-␥1. Through its interaction with LAT and SLP-76, PLC-␥1 is also brought to the plasma membrane. Upon activation by ZAP-70 and Itk, PLC-␥1 hydrolyzes phosphatidylinositol 4,5-bisphosphate into the secondary messengers inositol trisphosphate and diacylglycerol (4, 5). Inositol trisphosphate induces Ca 2ϩ mobilization, whereas diacylglycerol binds to RasGRP1 to activate the Ras-MAPK pathway (6, 7). LAT also contributes to Ras-MAPK activation through recruitment of the Grb2-Sos complex to the plasma membrane to activate Ras (1, 8). Initiation of these signaling cascades eventually leads to activation of transcription factors that regulate the genes critical for T cell proliferation and effector functions.The essential role of LAT in T cell activation was initially demonstrated in LAT-deficient Jurkat T cells. Although the proximal signals upstream of LAT phosphorylation remain intact in these cells, TCR-mediated calcium mobilization, Ras-MAPK activation, and NFAT activation are all abolished (9, 10). LAT function during T cell development has also been well characterized. Thymocyte development in LAT Ϫ/Ϫ mice is completely blocked at the CD25 ϩ CD44 Ϫ DN3 stage, indicating an absolute requirement for LAT in pre-TCR signaling (11). Our recent studies using LAT conditional knock-out mice show that LAT is also required during thymocyte development at the DP to SP transition, as positive selection in LAT-deficient DP thymocytes is markedly compromised in these mice. Consequently, the maturation of SP thymocytes is severely blocked (12). In addition, a point mutation at LAT tyrosine 136 (Y136F), which specifically abolishes the LAT-PLC-␥1 interaction, renders both positive and negative thymic selection processes defective in the LATY136F knock-in mice (13), highlighting the importance of LAT-m...

Background: RasGRP4 is one member of the RasGRP family and is highly expressed in mast cells. Results: Fc⑀RI signaling, mast cell function, and thymocyte development are severely impaired by RasGRP1 and RasGRP4 deficiency. Conclusion: This study indicated that the RasGRP family is important in mast cells and T cells.Significance: This work demonstrates immune cells employ multiple members of the RasGRP family to activate the Ras-Erk pathway.

Linker for activation of T cells (LAT) is a transmembrane adaptor protein that is essential to bridge T cell receptor (TCR) engagement to downstream signaling events. The indispensable role of LAT in thymocyte development and T cell activation has been well characterized; however, the function of LAT in cytotoxic-T-lymphocyte (CTL) cytotoxicity remains unknown. We show here that LAT-deficient CTLs failed to upregulate FasL and produce gamma interferon after engagement with target cells and had impaired granule-mediated killing. We further dissected the effect of the LAT deletion on each step of granule exocytosis. LAT deficiency led to altered synapse formation, subsequently causing unstable T cell-antigen-presenting cell (APC) conjugates. Microtubule organizing center polarization and granule reorientation were also impaired by LAT deficiency, leading to reduced granule delivery. Despite these defects, granule release was still observed in LAT-deficient CTLs due to residual calcium flux and phospholipase C (PLC) activity. Our data demonstrated that LAT-mediated signaling intricately regulates CTL cytotoxicity at multiple steps.