Sequences 3' of immunoglobulin heavy chain genes influence their expression

Abstract

The function of sequences 3' of Ig genes in controlling their expression has been investigated by analyzing mutants of Ig-producing cells and by gene transfection experiments. A mutant of an IgA producing myeloma was isolated whose steady-state level of heavy chain mRNA and protein was decreased. Analysis of the mutant showed it had deleted at least 4 kb of DNA immediately 3' of the alpha gene and introduced at least 5 kb of non-Ig sequence in its place. Examination of nuclear RNA showed no accumulation of aberrant transcripts or altered processing patterns. Instead, the transcription rate of heavy chain in the mutant was approximately 1/7 of that in its parent. This mutant suggests that sequences 3' of Ig genes facilitate their transcription; alternatively, the non-Ig sequences may act to depress transcription. A complete gamma 2b heavy chain gene containing both the secreted and membrane exons was transfected into lymphoid cells. The ratio of membrane/secreted Ig mRNA produced by the transfectants was found to reflect the phenotype of the recipient cell; myelomas made mostly secreted mRNA while lymphomas made only a slight excess of secreted mRNA. When a heavy chain was used with a deletion of sequences within the IVS between the secreted and membrane exons which left the AATAA poly A addition signal intact, but removed the site of cleavage and polyadenylation, the processing ratio was altered so that predominantly membrane Ig was produced in transfected myeloma cells. The alteration in processing could be a result of the deletion of the normal site for poly A addition; alternatively it could be a result of the deletion of another sequence which is recognized by processing enzymes. A 13 bp sequence (GTCCTGGTTCTTT), was found to be highly conserved both in position and sequence in human and mouse gamma chain genes. When a gene with a deletion which left the poly A addition site and the conserved sequence intact was used for transfection, the processing pattern was found to be identical to that of a wild type heavy chain gene.