I.29 lymphoma cells express a nonmutated VH gene before and after H chain switch
UMass Chan Affiliations
Department of Molecular Genetics and MicrobiologyDocument Type
Journal ArticlePublication Date
1988-03-01Keywords
Amino Acid SequenceAnimals
B-Lymphocytes
Base Sequence
Cell Line
Clone Cells
Cloning, Molecular
Germ Cells
Hybrid Cells
Immunoglobulin A
Immunoglobulin Heavy Chains
Immunoglobulin M
Immunoglobulin Switch Region
Immunoglobulin Variable Region
Liver
Lymphoma
Mice
Molecular Sequence Data
Mutation
Life Sciences
Medicine and Health Sciences
Women's Studies
Metadata
Show full item recordAbstract
The I.29 B cell lymphoma consists of IgM+ and IgA+ cells which express the same germ-line VH gene. IgA+ cells of the I.29 lymphoma were derived from the IgM+ cells by a typical H chain switch recombination event. The IgM+ cells can be induced with LPS to undergo H chain switching in culture. It has been proposed that the somatic hypermutation process is activated during H chain switch, since V genes expressed in IgG+ and IgA+ cells have more frequently undergone mutation than those expressed in IgM+ cells. We have investigated this question by sequencing VH genes expressed before and after H chain switch in the I.29 lymphoma. We have also sequenced the germ-line VH gene corresponding to the gene expressed by I.29 cells to determine whether the VH gene expressed in the IgM+ cells had already undergone somatic mutation. Our results indicate that somatic mutation was not activated in the precursor cell for the I.29 lymphoma, nor during isotype switch in I.29 cells. It is possible that cells of the I.29 lymphoma, or their precursor, have not received the signal which induces somatic mutation, or that I.29 cells belong to a subset of B cells that cannot be induced to undergo any (or much) somatic mutation.Source
J Immunol. 1988 Mar 1;140(5):1676-84.Permanent Link to this Item
http://hdl.handle.net/20.500.14038/50706PubMed ID
3126234Related Resources
Link to article in PubMedCollections
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Induction of immunoglobulin isotype switching in cultured I.29 B lymphoma cells. Characterization of the accompanying rearrangements of heavy chain genesStavnezer, Janet; Sirlin, S.; Abbott, J. (1985-03-01)The murine B cell lymphoma I.29 contains cells expressing surface IgM or IgA with identical heavy chain variable regions (9, 25, and D. Klein and J. Stavnezer, unpublished data). Purified IgM+ cells from the lymphoma have been adapted to culture and induced to switch to IgA, IgE, or IgG2 by treatment with lipopolysaccharide (LPS) or by treatment with a monoclonal anti-I.29 antiidiotype plus LPS. Clones of IgM+ cells have been obtained and induced to switch. Under optimal conditions, 30% of the cells in the culture expressed IgA 8 d after the inducers were added, and by 15 d 90% of the cells were IgA+. In actively switching cultures, up to 50% of the cells whose cytoplasm stained positively with anti-IgA stained simultaneously with anti-IgM, which indicates that the appearance of IgA+ cells in the cultures was due to isotype switching and not to clonal outgrowth. Examination by Southern blotting experiments of the Ig heavy chain genes in I.29 cells before and after switching revealed that isotype switching was accompanied by DNA recombinations that occurred within or immediately 5' to the tandemly repeated switch sequences. Within 3 d after the addition of inducers of switching, the nonexpressed chromosome underwent a variety of deletions or expansions within the S mu region, and a portion of the S alpha regions had undergone a 0.9-kb deletion. In cultures that contained at least 12% IgA+ cells, rearranged, expressed alpha genes, produced by recombination between the S mu region within the expressed mu gene and the S alpha region, were detected.
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Overexpression of BSAP/Pax-5 inhibits switching to IgA and enhances switching to IgE in the I.29 mu B cell lineQiu, G.; Stavnezer, Janet (1998-09-15)B cell-specific activator protein (BSAP)/Pax-5 is a paired domain DNA-binding protein expressed in the developing nervous system, testis, and in all B lineage cells, except terminally differentiated plasma cells. BSAP regulates transcription of several genes expressed in B cells and also the activity of the 3' IgH enhancer. As it has binding sites within or 5' to the switch regions of nearly all Ig heavy chain C region genes and also is known to increase transcription of the germline epsilon RNA, BSAP has been hypothesized to be involved in regulation of Ab class switch recombination. To directly examine the effects of BSAP on isotype switching, we use a tetracycline-regulated expression system to overexpress BSAP in the surface IgM+ I.29 mu B cell line, a mouse cell line that can be induced to undergo class switch recombination. We find that overexpression of BSAP inhibits switching to IgA in I.29 mu cells stimulated with LPS + TGF-beta 1 + nicotinamide, but enhances switching to IgE in cells stimulated with LPS + IL-4 + nicotinamide. Parallel to its effects on switching, overexpression of BSAP inhibits germline alpha RNA expression and the transcriptional activity of the germline alpha promoter, while enhancing activity of the germline epsilon promoter. Proliferation of I.29 mu cells is not affected in this system. The possible mechanisms and significance of the effect of BSAP on isotype switching are discussed.
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Rearrangements and deletions of immunoglobulin heavy chain genes in the double-producing B cell lymphoma I.29Stavnezer, Janet; Marcu, K. B.; Sirlin, S.; Alhadeff, B.; Hammerling, U. (1982-08-01)The B cell lymphoma I.29 consists of a mixture of cells expressing membrane-bound immunoglobulin M (IgM) (lambda) and IgA (lambda) of identical idiotypes. Whereas most of the cells express either IgM or IgA alone, 1 to 5% of the cells in this tumor express IgM and IgA simultaneously within the cytoplasm and on the cell membrane (R. Sitia et al., J. Immunol. 127:1388-1394, 1981; R. Sitia, unpublished data). When IgM+ cells are purified from the lymphoma and passaged in mice or cultured, a portion of the cells convert to IgA+. These properties suggest that some cells of the I.29 lymphoma may undergo immunoglobulin heavy chain switching, although it is also possible that the mixed population was derived by a prior switching event in a clone of cells. We performed Southern blotting experiments on genomic DNAs isolated from populations of I.29 cells containing variable proportions of IgM+ and IgA+ cells and on a number of cell lines derived from the lymphoma. The results were consistent with the deletion model for heavy chain switching, as the IgM+ cells contained rearranged mu genes and alpha genes in the germ line configuration on both the expressed and nonexpressed heavy chain chromosomes, whereas the IgA+ cells had deleted both mu genes and contained one rearranged and one germ line alpha gene. In addition, segments of DNA located within the intervening sequence 5' to the mu gene, near the site of switch recombination, were deleted from both the expressed and the nonexpressed chromosomes. Although mu genes were deleted from both chromosomes in the IgA+ cells, the sites of DNA recombination differed on the two chromosomes. On the expressed chromosome, Smu sequences were recombined with S alpha sequences, whereas on the nonexpressed chromosome, Smu sequences were recombined with S gamma 3 sequences.