RAG-1 and RAG-2 are not sufficient to direct all phases of immunoglobulin gene rearrangement in pre-B-cell lines.
To probe the factors controlling immunoglobulin heavy-chain gene rearrangement, we analyzed Abelson virus-transformed pre-B-cell lines that fail to undergo VH-to-DJH joining at an appreciable frequency. Despite this feature, some of these cell lines (rechi) rearrange an extrachromosomal recombination substrate at levels normal for transformed pre-B cells. Others (reclo) rearrange these substrates at levels characteristic of nonlymphoid hematopoietic cells. The DJH rearrangements from a representative rechi cell line were aberrant, suggesting that these cells probably fail to complete heavy-chain gene assembly because some of the necessary cis-acting signals are missing. In contrast, both DJH rearrangements from a reclo cell line appeared normal in structure, indicating that trans-acting factors necessary for recombination might be missing. Introduction of the RAG-1 and RAG-2 genes, genes encoding two such factors, failed to confer a rechi phenotype to these cells. However, fusion of the reclo cells to a rechi cell line generated a high frequency of rechi hybrids. In addition, most of the hybrids rearranged the endogenous kappa light-chain locus. Neither the rechi phenotype nor kappa-chain rearrangement correlated with levels of RAG-1 and RAG-2 expression in all of the hybrids. Thus, both gene transfer and cell fusion experiments indicate that RAG-1 and RAG-2 are not sufficient to activate immunoglobulin gene recombination in at least some pre-B-cell lines. In addition, the fusion experiments suggest that two gene products in addition to RAG-1 and RAG-2 may be required for kappa-gene rearrangement.
Wang
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LC
Department of Pathology, Tufts University School of Medicine, Boston, Massachusetts 02111.
Rosenberg
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N
D016475:3T3 Cells; D000818:Animals; D001402:B-Lymphocytes / Q000378:metabolism*; D001483:Base Sequence; D015139:Blotting, Southern; D002461:Cell Line, Transformed; D004247:DNA; D004254:DNA Nucleotidyltransferases / Q000378:metabolism; D004268:DNA-Binding Proteins; D015322:Gene Rearrangement, B-Lymphocyte; D018398:Homeodomain Proteins; D007143:Immunoglobulin Heavy Chains / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D011506:Proteins / Q000378:metabolism*; D045567:VDJ Recombinases
D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D007143:Immunoglobulin Heavy Chains; D011506:Proteins; C499308:Rag2 protein, mouse; C064655:V(D)J recombination activating protein 2; C064658:RAG-1 protein; D004247:DNA; D004254:DNA Nucleotidyltransferases; D045567:VDJ Recombinases