Transfer and expression of three cloned human non-HLA-A,B,C class I major histocompatibility complex genes in mutant lymphoblastoid cells.

Abstract

The HLA-A, -B, and -C class I human histocompatibility antigens and the genes that encode them have been isolated and characterized. Apparently complete class I non-HLA-A,B,C genes have been identified on HindIII-generated 5.4-kilobase (kb), 6.0-kb, and 6.2-kb DNA fragments derived from lymphoblastoid cell line (LCL) 721. We studied the expressibility of these genes by subcloning them into the nonintegrating pHeBo vector and transferring the chimeric plasmids into mutant LCL 721.221. This mutant was derived from LCL 721 by means of immunoselections following .gamma.-ray mutagenesis that eliminated expressions of the HLA-A,-B, and -C .alpha. chains. The HLA-A,B,C-null phenotype of mutant 721.221 made it possible to monitor the expression of class I genes transferred into it by assaying cell surface binding of monoclonal antibodies BBM.1 and W6/32, which recognize .beta.2-microglobin and HLA class I .alpha.-chain epitopes, respectively. Increased binding of BBM.1 and W6/32 was clearly observed in transferents containing the class I gene of the 6.0-kb DNA fragment but not in transferents containing the class I genes of the 5.4- and 6.2-kb DNA fragments. However, one-dimensional gel electrophoresis of BBM.1 and W6/32 immunoprecipitates made with [35S]methionine-labeled cell lysates showed that transfer of each non-HLA-A,B,C class I gene into 721.211 resulted in the appearance of an .alpha. chain that coprecipitated with .beta.2-microglobulin. The three previously unreported .alpha. chains differed from each other in size and were smaller than HLA-A,-B, and -C .alpha. chains. These observations clearly show that these three cloned, nonallelic, non-HLA-A,B,C class I genes encode .alpha. chains that can be expressed in human cells.