MUTANT FREQUENCY AT THE H-2K CLASS-1 AND HPRT GENES IN T-LYMPHOCYTES FROM THE X-RAY-EXPOSED MOUSE

The frequency of H-2K(k) and HPRT-deficient T cells was measured in th e H-2K(b,k)D(d,k) genotype mouse 8-10 weeks after X-ray exposure at do ses up to 6 Gy to compare the mutant frequency (MF) of an autosomal ge ne with that of an X-chromosomal gene. H-2K mutants were enriched by m agnetic cell separation (MACS) using the H-2K(k)-specific monoclonal a ntibody H100.5/28 and were isolated by limiting dilution cloning. Fina lly, the mutant phenotype was verified by flow cytometric analysis in a representative number of clones. The frequency of HPRT-deficient T c ells rises from 2.5 X 10(-6) at 0 Gy to a maximum of 1.13 X 10(-4) at 4 Gy, and decreases to 2.9 X 10(-5) at 6 Gy. The H-2K(-) MF in the non -irradiated mouse was 8.4 X 10(-7). It increases with dose to a maximu m of 8.1 X 10(-6) at 4 Gy and declines to 3.3 X 10(-6) at 6 Gy. The H- 2K(-) MF measured depends on the monoclonal antibody used for the isol ation of mutants. In a pilot study with another H-2K(k)-specific monoc lonal antibody (11.4.1), the spontaneous MF was four times higher than in experiments with the H100.5/28 monoclonal antibody. The expression of other class 1 antigens was investigated in H-2K(-) clones. The H-2 D(d) antigen had also disappeared in six of 41 clones from irradiated animals. This gene is situated at a distance of 1500 kb from the K-loc us. The H-2K(b) antigen was present in every investigated clone. In th e discussion a model is presented that explains the shape of the dose- response curve of MF by selection against mutants in vivo systems unde r homeostasis. The results of the present investigation indicate that observed X-ray mutagenicity depends on many factors and that several g enes have to be explored before reliable risk estimates are possible.