LFA-1 Regulates CD8 ⁺ T Cell Activation and Immune Signal Network.

Abstract 1641

Poster Board I-667

LFA-1 regulates T cell activation and signal transduction through the immunological synapse. TCR stimulation rapidly activates LFA-1, which provides unique LFA-1-dependent signals to promote T cell activation. We found LFA-1 directly participates in Erk1/2 signaling upon TCR stimulation in CD8⁺ T cells. The presence of LFA-1, not ligand binding, is required for the TCR-mediated Erk1/2 signal pathway. LFA-1-KO T cells have defects in sustained Erk1/2 signaling and TCR/CD3 clustering, which subsequently prevents MTOC re-orientation, cell-cycle progression and mitosis. LFA-1 regulates the TCR-mediated Erk1/2 signal pathway in the context of immunological synapse for recruitment and amplification of Erk1/2 signal. In addition, LFA-1 ligation with ICAM-1 generates an additional Erk1/2 signal, which synergizes with the existing TCR-mediated Erk1/2 signal to enhance T cell activation. We demonstrated that the function of LFA-1 is to enhance TCR signaling through the immunological synapse and deliver distinct signal in CD8⁺ T cell activation.

Based on our results, we proposed a model of TCR-mediated and LFA-1-mediated Erk1/2 signal pathways in CD8⁺ T cell activation. However, the detailed molecular pathways that regulate these processes and global impact on immune functions are poorly defined. With the launching of The Immunological Genome Project, we have generated data with CD8⁺ T cell expression array to explore and understand the LFA-1 and TCR signaling network. GeneChip hybridization and analysis CD8⁺ T cells from C57BL/6 mice and LFA-1-KO mice were collected before and after stimulation. Microarray experiments were carried out using the “ Mouse Whole Genome Oligo Microarray Kit” from Agilent. Differentially expressed gene lists between samples were considered significant if their p values were <0.0001 and their fold-change >1.8. The results are summarized below: 1) 641 genes were up-regulated and 174 were down-regulated in unstimulated LFA-1-KO CD8⁺ T cells comparing to these in unstimulated WT CD8⁺ T cells; 2)1036 genes were up-regulated and 406 were down-regulated in activated LFA-1-KO CD8⁺ T cells comparing to these in activated WT CD8⁺ T cells after 1 hours stimulation with unstimulated CD8⁺ T cells as control; 3)636 genes were up-regulated and 354 were down-regulated in activated LFA-1-KO CD8⁺ T cells comparing to these in activated WT CD8⁺ T cells after 8 hours stimulation with unstimulated CD8⁺ T cells as control.

To estimate which pathways are significantly enriched among the genes that are differentially expressed, we used a database IPA (Ingenuity Pathways Analysis) software. Signaling pathways enriched by the genes differentially expressed in activated LFA-1-KO CD8⁺ T cells vs WT CD8⁺ T cells after 8 hours stimulation generated the most highly enriched differentially expressed genes in signaling pathways displayed below: 1) Pattern Recognition Receptors in Recognition of Bacteria and Viruses; 2) Activation of IRF by Cytosolic Pattern Recognition Receptors; 3) Aryl Hydrocarbon Receptor Signaling; 4) IL-10 Signaling; p38 MAPK Signaling; 5) LPS/IL-1 Mediated Inhibition of RXR Function; Cell Cycle: G2/M DNA Damage Checkpoint Regulation; 6) Notch Signaling; Natural Killer Cell Signaling; 7) Cell Cycle: G1/S Checkpoint Regulation and Chemokine Signaling. Furthermore, we examined the biological functions enriched by the genes differentially expressed in activated LFA-1-KO CD8⁺ T cells vs WT CD8⁺ T cells. The LFA-1-KO CD8⁺ T cells vs WT CD8⁺ T cells after 8 hours stimulation generated the most highly enriched differentially expressed genes in following biological functions: Immunological Disease; Antigen Presentation; Cell-mediated Immune Response; Humoral Immune Response; Inflammatory Response; Cellular Development; Cell-To-Cell Signaling and Interaction; Hematological System Development and Function; Immune Cell Trafficking; Cellular Growth and Proliferation; Inflammatory Disease; Cellular Movement; Hematopoiesis Cell Death; Hematological Disease; Lymphoid Tissue Structure and Development; Infectious Disease; Tissue Development and Cancer. Our results indicate that LFA-1 plays an important role in the immune signal network and has a global impact on immune system.