Formation of coding joints in V(D)J recombination-inducible severe combined immune deficient pre-B cell lines.
Characterization of the severe combined immune deficient (scid) defect in the recombination process has provided many insights into the underlying mechanisms of variable (diversity) joining recombination. By using recombination-inducible scid pre-B cell lines transformed with the temperature-sensitive Abelson-murine leukemia virus, we show that large quantities of recombination intermediates can be generated, and their resolution can be followed during further cell culture. In this study, we demonstrate that the ability of these scid pre-B cell lines to resolve coding ends depends on the cell culture temperature. At the nonpermissive temperature of 39 degreesC, scid pre-B cell lines fail to form coding joints and contain mostly unresolved hairpin-coding ends. Once the cell culture is returned to the permissive temperature of 33 degreesC, these same cells make a significant amount of coding joints concomitant with the disappearance of hairpin-coding ends. Thus, the scid cells are capable of resolving coding ends under certain culture conditions. However, the majority of the recovered coding joints contains extensive deletions, indicating that the temperature-dependent resolution of coding ends is still scid-like. Our results suggest that the inability of scid cells to promptly nick hairpin-coding ends may lead to aberrant joining in these cells.
Chang
Y
Y
Department of Microbiology, Molecular and Cellular Biology Program, Arizona State University, Tempe, AZ 85287-2701, USA. yung.chang@asu.edu
Brown
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ML
D000818:Animals; D001402:B-Lymphocytes / Q000276:immunology*; D002460:Cell Line; D006412:Hematopoietic Stem Cells / Q000276:immunology*; D007133:Immunoglobulin Joining Region / Q000235:genetics*; D007135:Immunoglobulin Variable Region / Q000235:genetics*; D051379:Mice; D016513:Mice, SCID; D008822:Mice, Transgenic; D009690:Nucleic Acid Conformation; D011995:Recombination, Genetic; D016511:Severe Combined Immunodeficiency / Q000276:immunology*
D007133:Immunoglobulin Joining Region; D007135:Immunoglobulin Variable Region