Mice reconstituted with DNA polymerase beta-deficient fetal liver cells are able to mount a T cell-dependent immune response and mutate their Ig genes normally - PubMed (original) (raw)

Mice reconstituted with DNA polymerase beta-deficient fetal liver cells are able to mount a T cell-dependent immune response and mutate their Ig genes normally

G Esposito et al. Proc Natl Acad Sci U S A. 2000.

Abstract

The ubiquitously expressed, error-prone DNA polymerase beta (polbeta) plays a role in base excision repair, and the involvement of this molecule in the nonhomologous end joining (NHEJ) process of DNA repair has recently been demonstrated in yeast. Polbeta-deficient mice are not viable, and studies on conditional mutants revealed a competitive disadvantage of polbeta(-/-) vs. wild-type cells. We show here that polbeta-deficient mice survive up to day 18.5 postcoitum, but die perinatally; a circumstance that allowed the investigation of a potential role of polbeta in lymphocyte development by transfer of fetal liver cells (FLC) derived from polbeta(-/-) embryos into lethally irradiated hosts. FLC transfers using mutant cells lead to an almost normal reconstitution of the lymphocyte compartment, indicating that polbeta-deficiency does not prevent V(D)J recombination, which is known to employ factors of the NHEJ pathway. Mice reconstituted with polbeta(-/-) FLC mount a normal T cell-dependent immune response against the hapten (4-hydroxy-3-nitrophenyl) acetyl (NP). Moreover, germinal center B cells from NP-immunized reconstituted mice show normal levels and patterns of somatic point mutations in their rearranged antibody genes, demonstrating that polbeta is not critically involved in somatic hypermutation.

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Figures

Figure 1

Figure 1

polβ−/− embryos (Right) at day 18.5 p.c. are growth retarded compared with their polβ+/− (Center) and polβ+/+ (Left) littermates. Homozygous mutant embryos appeared to be cyanotic.

Figure 2

Figure 2

Lethally irradiated recipient 129/Sv mice were reconstituted with 106 FLC prepared from day 18.5 p.c. polβ+/− (●) and polβ−/− (○) embryos. One and two months after reconstitution, the total number of thymocytes, splenocytes, and femur bone marrow cells (A) was determined as well as the number of splenic B220+ B cells, CD4+ T cells, and CD8+ T cells (B).

Figure 3

Figure 3

Pattern of nucleotide exchanges in the V186.2 rearrangements of GC B cells derived from a wild-type C57BL/6 mouse and from mice reconstituted with wild-type and polβ-deficient day 18.5 p.c. FLC, respectively. n, the number of mutations; shared mutations in clonally related sequences were counted only once. The G to T exchange in codon 33, which represents a result of selection for NP-binding, was not considered to avoid skewing of the analysis (see text). Ts., transitions; Tv., transversions; Ts./Tv., the transitions over transversions ratio.

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