Inhibitory Effect of the Transcription Factor Encoded by themi Mutant Allele in Cultured Mast Cells of Mice

Abstract

AbstractThe 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 domain. The mice of mi/migenotype express mi-MITF, whereas the mice of tg/tggenotype have a transgene at the 5′ flanking region of themi gene and do not 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 ortg/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 using Southern analysis, the number was larger in the (+/+-mi/mi) library than in the (+/+-tg/tg) library. Second, we compared mRNA expression of six genes betweenmi/mi and tg/tg CMCs by Northern analysis. The transcription of three genes encoding mouse mast cell proteases was impaired in both mi/mi and tg/tg CMCs. On the other hand, the transcription of three genes encoding c-kit receptor, tryptophan hydroxylase, and granzyme B was markedly reduced inmi/mi CMCs, but the reduction was significantly smaller intg/tg CMCs. These results indicated the inhibitory effect ofmi-MITF on the transactivation of particular genes in CMCs.