REGULATION OF MOUSE MAST-CELL PROTEASE-6 GENE-EXPRESSION BY TRANSCRIPTION FACTOR ENCODED BY THE MI LOCUS

The mi locus of mice encodes a member of the basic-helix-loop-helix-le ucine zipper (bHLH-Zip) protein family of transcription factors (herea fter called MITF). Because the expression of the mouse mast cell prote ase 6 (MMCP-6) gene is remarkably reduced in mast cells of mi/mi mutan t mice, we investigated the effect of MITF on the transcription of the MMCP-6 gene. First, we introduced the normal (+) MITF cDNA into mi/mi cultured mast cells using the retroviral vector. Overexpression of +- MITF but not mi-MITF normalized the expression of the MMCP-6 gene, ind icating the involvement of +-MITF in the MMCP-B gene transactivation. Second, we analyzed the promoter of the MMCP-6 gene by the transient c otransfection assay, The luciferase construct under the control of the MMCP-B promoter and the cDNA encoding +-MITF or mi-MITF were cotransf ected into NIH/3T3 fibroblasts. The coexpression of +-MITF but not mi- Miff increased the luciferase activity 10-fold. We found a CACATG and a CATCTG motif in the MMCP-6 promoter, both of which are generally rec ognized by bHLH-Zip-type transcription factors. We also found a GACCTG motif that was strongly bound by +-MITF. These three motifs were nece ssary for the 10-fold transactivation ability of the MMCP-B promoter b y +-MITF, Mutations of each motif significantly reduced the transactiv ation, suggesting that +-MITF directly transactivated the MMCP-6 gene through these three motifs. (C) 1996 by The American Society of Hemato logy.