Inhibitory effect of the transcription factor encoded by the mi mutant allele in cultured mast cells of mice

The mi locus of mice encodes a transcription factor of the basic-helix-loop -helix-leucine zipper protein family (MITF). The MITF encoded by the mutant mi allele (mi-MITF) deletes 1 of 4 consecutive arginines in the basic doma in. The mice of mi/mi genotype express mi-MITF, whereas the mice of tg/tg g enotype have a transgene at the 5' flanking region of the mi gene and do no t express any MITF. To investigate the function of mi-MITF in cultured mast cells (CMCs), we took two approaches. First, mRNA obtained from mi/mi CMCs or tg/tg CMCs was subtracted from complementary (c) DNA library of normal (+/+) CMCs, and the (+/+-mi/mi) and (+/+-tg/tg) subtraction libraries were obtained. When the number of clones that hybridized more efficiently with /+ CMC cDNA probe than with mi/mi or tg/tg CMC cDNA probe was compared usin g Southern analysis, the number was larger in the (+/+-mi/mi) library than in the (+/+-tg/tg) library. Second, we compared mRNA expression of six gene s between mi/mi and tg/tg CMCs by Northern analysis. The transcription of t hree genes encoding mouse mast cell proteases was impaired in both mi/mi an d tg/tg CMCs. On the other hand, the transcription of three genes encoding c-kit receptor, tryptophan hydroxylase, and granzyme B was markedly reduced in mi/mi CMCs, but the reduction was significantly smaller in tg/tg CMCs, These results indicated the inhibitory effect of mi-MITF on the transactiva tion of particular genes in CMCs. (C) 1999 by The American Society of Hemat ology.