The molecular basis of a heavy-chain, disease-like gamma 2B immunoglobulin protein has been investigated in a murine model system. DNA sequencing of the expressed gamma 2b gene in the mutant showed a deletion of 99 nucleotides around the 3' end of the CH?1? domain, which removed the splice site and neighboring DNA. The sequence deleted in the mutant is flanked by short direct repeats of the octameric sequence CCAGCCAG in the wild-type gene. In the mutant, one copy of the repeat in addition to the sequences between the repeats has been lost. This explains why no CH?1? sequences are found in the mRNA. A deletion between repeated pentanucleotides has also been characterized in another mutant. The effect of direct repeats on deletion formation will be examined as well as the mechanisms by which the deletions arose. The metabolism of the mutant mRNAs produced by the altered genes is being studied by measurements of the rates of synthesis and degradation. The differential expression of two forms of mRNA, coding for proteins with either secretory or membrane anchoring carboxyl-termini, from a single gamma 2b gene is being investigated. The immunoglobulin gene has been transfected, in collaboration with Dr. S. L. Morrison, into cell lines fixed at various stages of B-cell development. We find that the transfected gene is differentially expressed in the various cell lines. Constructs with deletions of portions of the immunoglobulin genes have been shown to differentially produce membrane- or secretory-specific mRNAs with sequences 3' of the secretory polyadenylation site important for correct 3' end formation. Equimolar transcription of the entire gamma 2b gene, secretory plus membrane exons, has been observed. In isolated nuclei, the secretory polyadenylstion site is preferentially used by cells producing primarily secretory mRNA. Selective use of alternate polyadenylation sites can, therefore, modulate the production of the two forms of mRNA from one gene. (AB)
