A human neoplastic B cell line SSK41 that expresses IgM on its surface switches spontaneously to IgG-producing cells. The SSK41 line contains a single immunoglobulin heavy-chain locus, the constant region (C) genes of which retain the germline configuration. The IgG-producing SSK41 line was purified by sorting, and shown to have undergone S-S recombination with deletion of the C mu gene. This line produced secretory and membrane-bound forms of gamma-chain mRNA. From cDNA libraries of a mixed population of IgM+/IgG+ SSK41 cells, we have isolated cDNA clones encoding the mature membrane-bound and secretory forms of the mu and gamma 1 heavy chains, all of which share the same variable region sequence. cDNA clones containing the mature gamma 3 chain were identified as well. We also isolated cDNA clones containing C gamma 1 and C gamma 3 sterile transcripts from the SSK41 line. These sterile transcripts contained additional exon sequences designated 'I' which were localized upstream of the C gamma 1 and C gamma 3 switch regions and homologous to murine counterparts. The I sequences were precisely spliced to the 5' ends of the corresponding C gamma exon sequences. These features of germline CH transcripts, i.e. the isotype specificity to class switching, location of exons, and sequences per se, are highly conserved between man and mouse.

We have studied immunoglobulin doubleisotype expression in, a transgenic mouse (TG.SA) in which expression of the endogenous immunoglobulin heavy chain locus is almost completely excluded by a nonallelic rearranged human ,u transgene. By flow-cytometric analyses, we have shown that a small, but significant, portion (about 4%) of transgenic spleen cells expresses human ,u (transgene) and mouse y (endogenous) chains when cultured in vitro with bacterial lipopolysaccharide and interleukin 4. By using amplification of cDNA by the polymerase chain reaction, followed by cloning and sequencing of the amplified cDNA fragment, we have demonstrated expression of trans-mRNA consisting of the transgenic variable and endogenous constant (yl) region sequences. Such trans-mRNA could be produced by either switch recombination or trans-splicing between the transgene and endogenous sterile yl-gene transcripts. These results indicate that trans-splicing might be a possible mechanism for the immunoglobulin double-isotype expression in normal B lymphocytes that have not rearranged the second expressed constant region gene.Ce genes (12, 13). As the existence of such sterile transcripts from a CH gene seemed to correlate well with switching to that particular CH isotype, the sterile transcripts were thought to be indicative of opening of the chromatin structure of the CH genes that make the putative switch recombinase system accessible (12-15).The accessibility model (12-15) thus considered the sterile transcript useless by itself. However, there remains the possibility that the sterile transcript could be utilized as a substrate of the trans-splicing reaction for the double-isotype expression (P. Sideras, T.-R.M., A.S., and T.H., unpublished data). To test this hypothesis, we took advantage of a human immunoglobulin transgenic mouse (TG.SA) in which a majority of B lymphocytes expresses human transgenic u chain and, by allelic exclusion, does not completely rearrange the endogenous immunoglobulin heavy chain locus (16). In this paper, we report that the transgenic VHDJH exon can be expressed in conjunction with the endogenous mouse C.1 gene. These results could accommodate either a transsplicing or a DNA recombinant mechanism.l During the course of B-lymphocyte differentiation, progeny of a single B lymphocyte can switch the expressed immunoglobulin isotype from IgM to IgG or other classes of immunoglobulin without changing the antigen specificity determined by the V region sequence (where V, D, J, and C indicate variable, diversity, joining, and constant regions of immunoglobulin). This phenomenon, known as immunoglobulin class switching, is accompanied by DNA rearrangement that takes place between S regions located 5' to each CH gene

In three lymphoma cell lines carrying t(14;18), named FL-18, FL-218, and FL-318, the genomic organization of IgH gene was compared with the expression of bcl-2 gene; the t(14;18) of the FL-18 cells occurred downstream from the major breakpoint cluster region (mbr) of a bcl-2 gene, and that of the FL-218 and FL-318 cells within the mbr. The FL- 318 expressed the normal-sized bcl-2 transcript of 8.5-kb mRNA having the noncoding region 3 to the mbr, which was found in the FL-18, and the FL-218 lacking the intact bcl-2 gene did not. This finding suggests that in t(14;18)-positive lymphoma having the breakpoint within the mbr, transcription of the nontranslocated bcl-2 allele is not necessarily silent. In addition, the FL-218 and FL-318 expressed aberrant bcl-2 transcripts and heterogenous IgH transcripts lacking the VH region, and the bcl-2 transcripts each comigrated with parts of the sterile IgH mRNAs. The FL-318, which did not exhibit switch recombination on either IgH allele, contained abundant amounts of l gamma mRNAs, a prerequisite for the recombination into the C gamma locus. One of the I-mRNA species comigrated with the aberrant bcl-2 transcript. The FL-18 and FL-218 lacking the I gamma mRNAs had completed switch recombination of both IgH alleles. This result raises a possibility that deregulated bcl-2 transcription caused by t(14;18) is capable of playing a role in class switch recombination of IgH gene.

No abstract