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Complementation analysis of the murine scid cell line
M Z Zdzienicka1, W Jongmans, M Oshimura
1Department of Radiation Genetics and Chemical Mutagenesis, University of Leiden, The Netherlands.
Abstract:
It has been shown that several X-ray-sensitive Chinese hamster cell mutants defective in repair of DNA double-strand breaks (DSBs) are also impaired in the process of V(D)J recombination. The hamster mutants with this phenotype represent three distinct complementation groups, represented by the xrs series, XR-1 and V-3. The murine scid cell line also shows the same phenotype, and therefore we examined whether the scid mutant represents a new complementation group or belongs to one of the existing groups. Scid cells were fused with hamster cell mutants representing the three complementation groups. Hybrids between V-3 and scid cells were only partially complemented for X-ray sensitivity, whereas hybrids derived from fusions with the other mutants were resistant to X rays. These results suggest that V-3 and scid cells are defective in the same gene. To confirm this finding, a single human chromosome 8, which is known to carry the scid gene, was introduced into V-3 cells by microcell-mediated chromosome transfer. Nine hybrid clones derived from V-3 and carrying human chromosome 8 were obtained, and seven were found to be partially complemented for X-ray sensitivity. When human chromosome 8 was introduced into scid cells, seven of eight hybrid clones became resistant to X rays. The results indicate that the defective genes in V-3 and scid are both localized on human chromosome 8. This supports the results from the fusion analysis that V-3 and scid cells are defective in the same gene.
Insights
Chinese hamster cell mutants and murine scid cells share defects in DNA double-strand break repair and V(D)J recombination. These studies suggest the V-3 mutant and scid cells are defective in the same gene on human chromosome 8.
Area of Science:
- Genetics
- Molecular Biology
- Cell Biology
Background:
- Several X-ray-sensitive Chinese hamster cell mutants exhibit defects in DNA double-strand break (DSB) repair and V(D)J recombination.
- These mutants fall into three complementation groups: xrs series, XR-1, and V-3.
- The murine severe combined immunodeficiency (scid) cell line shares this phenotype, prompting investigation into its genetic relationship with the hamster mutants.
Purpose of the Study:
- To determine if the murine scid mutation represents a new complementation group or belongs to an existing one.
- To investigate the genetic locus of the defect in V-3 and scid cells.
Main Methods:
- Cell fusion experiments were conducted between scid cells and hamster mutants from the three complementation groups.
- Microcell-mediated chromosome transfer was used to introduce human chromosome 8 into V-3 cells.
- X-ray sensitivity was assessed in hybrid cell lines to evaluate complementation.
Main Results:
- Fusion of V-3 and scid cells resulted in partial complementation of X-ray sensitivity, suggesting a defect in the same gene.
- Introduction of human chromosome 8 into V-3 cells partially complemented X-ray sensitivity in most hybrid clones.
- Introduction of human chromosome 8 into scid cells conferred X-ray resistance in the majority of hybrid clones.
Conclusions:
- The V-3 hamster mutant and the scid mutation are defective in the same gene.
- The defective gene in both V-3 and scid cells is located on human chromosome 8.
- These findings support the genetic linkage between V-3 and scid mutations.
