INVOLVEMENT OF TRANSCRIPTION FACTOR ENCODED BY THE MOUSE MI LOCUS (MITF) IN EXPRESSION OF P75 RECEPTOR OF NERVE GROWTH-FACTOR IN CULTURED MAST-CELLS OF MICE

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). Cultured mast cells (CMCs) of mi/mi genotype showed a poor response to nerve growth factor (NGF). Addition of NGF to the suboptimal dose of interleukin-3 (IL-B);increased the plating efficien cy of normal (+/+) CMCs but nod: mi/mi CMCs. Although +/+ CMCs were be rberine sulfate-negative when cultured with IL-3, +/+ CMCs became berb erine sulfate-positive when cultured in the presence of both IL-3 and NGF, which suggested increased heparin content. In contrast, NGF did n ot influence the phenotype of mi/mi CMCs. The poor response of mi/mi C MCs to NGF was attributed to the deficient expression of p75 NGF recep tor. The purpose of the present study is to examine the effect of MITF on p75 gene transcription. Overexpression of +-MITF or mi-MITF was ob served in mi/mi CMCs to which cDNA encoding each type of MITF had been introduced using the retroviral vector. Overexpression of +-MITF but not of mi-MITF normalized the expression of p75 and the above-mentione d poor responses of mi/mi CMCs to NGF, indicating the involvement of -MITF in p75 gene transactivation. Then, we analyzed the promoter of t he p75 gene. Two CANNTG motifs recognized by bHLH-Zip-type transcripti on factors were conserved between the mouse and rat p75 promoters. One of these two CANNTG motifs was specifically bound by +-MITF. When the luciferase gene under the control of the p75 promoter was cotransfect ed into NIH/3T3 fibroblasts with cDNA encoding +-MITF or mi-MITF, luci ferase activity increased significantly only when +-MITF cDNA was cotr ansfected. The mutation of this CANNTG motif abolished the transactiva tion effect of +-MITF, indicating that +-MITF transactivated the p75 g ene, at least in part, through direct binding. (C) 1997 by The America n Society of Hematology.