The major goal of this proposal is to understand the molecular mechanism by which CD4+ T lymphocytes are restricted to antigen presenting cells bearing class II major histocompatibility complex (MHC) antigens. It is evident that both CD4 and CD8 are involved in the selection of T cells in the thymus. Further, these accessory molecules can act to enhance the avidity of low affinity T cell receptor responses and may also be involved in signalling. Using a cell-binding assay, we have shown that B lymphocytes which express MHC class II antigens bind to monolayers of adherent cells which express high levels of CD4. However, the molecular mechanism by which the class II molecule interacts with CD4 on the T cell has not been defined. Our approach to further characterization of the CD4-class II interaction will be as follows: (1) Identification of residues in the human DRbeta 1 molecule important for attachment to CD4. Several experimental approaches will be used. Initially, we will test a panel of EBV-transformed B cells homozygous for the HLA=DR1 allele in the cell binding assay to look for natural polymorphisms which might affect binding to CD4. This will be followed by site-directed mutagenesis of the DR 1beta polypeptide. Mutant molecules will be re- expressed in class II negative murine B cells for analysis of CD4 binding. In addition, recognition of mutant class II molecules will be analyzed using class II DR1 restricted influenza-specific helper cell clones. (2) Identification of cellular signals mediating adhesion and de- adhesion. The kinetics of adhesion and de-adhesion in the cell binding assay will be studies. The role of p56 lck in class II antigen binding to CD4 will also be analyzed by transfection and co-expression of normal and mutant lck genes in a CD4 transfectant in Chinese hamster ovary (CHO) cells which binds class II-bearing lymphocytes.