Somatic cell hybridization of human T- and B- lymphoblastoid cell lines (LCL) induces the expression of silent HLA genes encoded by the T-LCL partner. By using as fusion partners regulatory mutant B-LCL lacking class I or class II antigens on the cell surface, a minimum of two trans-acting genes regulating class I antigen expression and two trans-acting genes regulating class II antigen expression have been identified. Three of these genes act by regulating levels of specific mRNA. The fourth facilitates the assembly of HLA class I glycoprotein-beta2 microglobulin dimers. The objectives of this proposal are to characterize these regulatory genes, and other HLA regulatory genes which may be identified in additional mutant B-LCL. Genes will be isolated by molecular cloning techniques. They will be identified by subtractive hybridization approaches, or by secondary mutagenesis, using retroviral insertion, of hybrids of T-LCL with mutant B-LCL. Restoration of class I or class II antigen expression in the appropriate negative variant cell line upon transfection of a cloned gene will provide conclusive evidence of its regulatory function. Identification and structural characterization of genes which mediate the pre- and post- translational control of HLA antigen expression may ultimately contribute to our understanding of the regulation of in vivo immune responses.