Deficient perforin gene expression as a cause of natural killer defect in mutant mice of mi/mi Genotype

The mi locus of mice encodes a bHLH-Zip-type transcription factor, MITF. Mice of mi/mi genotype express a mutant form of MITF (mi-MITF), whereas mice of tg/tg genotype have a transgene in the 5′ flanking region of the mi gene and do not express any MITF. Spleen cells of mi/mi mice but not of tg/tg mice are defective in natural killer (NK) activity. We compared mi/mi spleen cells with tg/tg spleen cells to know the mechanism of the NK defect of the former. Although the proportion of cells expressing an NK cell marker, NK1.1, was comparable in mi/mi and tg/tg spleen cells, that of large granular lymphocytes was significantly decreased in mi/mi spleen cells. The difference between mi/mi and tg/tg spleen cells was reproducible in the culture continuously stimulated by interleukin-2. The cultured spleen cells of mi/mi but not of tg/tg mice were defective in the perforin gene expression. In the CTLL-2 cytotoxic T cells, transactivation of the perforin gene promoter was enhanced by wild-type (+) MITF but suppressed by mi-MITF through a 9-mer NF-P motif, which has been established as a strong cis-acting element. Neither +− nor mi-MITF bound the NF-P motif. Instead, either form of MITF interacted with the NF-P-binding nuclear protein obtained from tg/tg cultured spleen cells. The NF-P-binding protein was located mainly in the nucleus of +/+ and tg/tg cultured spleen cells, whereas it remained in the cytoplasm of the mi/mi cells. In addition to the defective nuclear localization, mi-MITF may also inhibit the translocation of the association partner, NF-P-binding protein. This appeared to result in the poor transactivation of the perforin gene and defective NK activity of mi/mi spleen cells.