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Involvement of transcription factor encoded by the mi locus in the
expression of c-kit receptor tyrosine kinase in cultured mast cells of mice
T Tsujimura, E Morii, M Nozaki, K Hashimoto, Y Moriyama, K Takebayashi, T Kondo, Y Kanakura and Y Kitamura
Department of Pathology, Medical School, Osaka University, Suita, Japan.
The mi locus of mice encodes a member of the basic-helix-loop-helix-
leucine zipper (bHLH-Zip) protein family of transcription factors
(hereafter called MITF). Cultured mast cells of mi/mi genotype (mi/mi CMCs)
did not normally respond to stem cell factor (SCF), a ligand for the c-kit
receptor tyrosine kinase. The poor response of mi/mi CMCs to SCF was
attributed to the deficient expression of c-kit both the mRNA and protein
levels. The purpose of the present study is to investigate the effect of
MITF on the transcription of the c-kit gene. First, we introduced cDNA
encoding normal (+) MITF or mutant (mi) MITF into mi/mi CMCs using the
retroviral vector. Overexpression of (+)-MITF but not mi- MITF normalized
the expression of the c-kit and the poor response of mi/mi CMCs to SCF,
indicating the involvement of (+)-MITF in the c-kit gene transactivation.
Second, we analyzed the promoter of the c-kit gene. Three CANNTG motifs
recognized by bHLH-Zip-type transcription factors were conserved between
the mouse and human c-kit promoters. Among these three CANNTG motifs, only
the CACCTG motif (nt -356 to - 351) was specifically bound by (+)-MITF.
When the luciferase gene under the control of the c-kit promoter was
contransfected into NIH/3T3 fibroblasts with cDNA encoding (+)-MITF or
mi-MITF, the luciferase activity significantly increased only when (+)-MITF
cDNA was cotransfected. The deletion of the promoter region containing the
CACCTG motif or the mutation of the CACCTG to CTCCAG abolished the
transactivation effect of (+)-MITF, indicating that (+)-MITF transactivated
the c-kit gene through the CACCTG motif. When the luciferase gene under the
control of the c-kit promoter was introduced into the FMA3 mastocytoma and
FEC-P1 myeloid cell lines, remarkable luciferase activity was observed only
in FMA3 cells. Thus, the involvement of (+)-MITF in the c-kit
transactivation appeared to be specific to the mast cell lineage.
Volume 88,
Issue 4,
pp. 1225-1233,
08/15/1996
Copyright © 1996 by The American Society of Hematology

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