Systematic method to obtain novel genes that are regulated by mi transcription factor: impaired expression of granzyme B and tryptophan hydroxylase in mi/mi cultured mast cells.
The mi locus encodes a member of the basic-helix-loop-helix-leucine zipper protein family of transcription factors (hereafter called MITF). We have reported that the expression of several genes was impaired in cultured mast cells (CMCs) of mi/mi genotype, and demonstrated the involvement of MITF in the transcription of these genes. To obtain new genes whose transcription may be regulated by MITF, we prepared a subtracted cDNA library using +/+ and mi/mi CMCs. We found two clones carrying the granzyme (Gr) B and tryptophan hydroxylase (TPH) cDNAs in the subtracted library. The expression of the Gr B and TPH genes decreased in mi/mi CMCs, and recovered to nearly normal level by the overexpression of normal (+) MITF but not of mutant (mi) MITF. The +-MITF bound three and one CANNTG motifs in the Gr B and TPH promoters, respectively, and transactivated these two genes, indicating the involvement of +-MITF in their expression. Because TPH is the rate-limiting enzyme for serotonin synthesis, we examined the serotonin content of +/+ and mi/mi CMCs. The serotonin content was significantly smaller in mi/mi CMCs than in +/+ CMCs. The introduction of +-MITF but not of mi-MITF normalized the serotonin content in mi/mi CMCs.
Ito
A
A
Department of Pathology, Medical School, Osaka University, Suita, Osaka 565, Japan.
Morii
E
E
Maeyama
K
K
Jippo
T
T
Kim
D K
DK
Lee
Y M
YM
Ogihara
H
H
Hashimoto
K
K
Kitamura
Y
Y
Nojima
H
H
D000818:Animals; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000235:genetics*; D005786:Gene Expression Regulation; D015723:Gene Library; D053804:Granzymes; D006640:Histidine Decarboxylase / Q000235:genetics; D008407:Mast Cells / Q000502:physiology; D051379:Mice; D008817:Mice, Mutant Strains; D051739:Microphthalmia-Associated Transcription Factor; D012697:Serine Endopeptidases / Q000235:genetics; D012701:Serotonin / Q000502:physiology; D014157:Transcription Factors / Q000235:genetics*; D053802:Tryptases; D014365:Tryptophan Hydroxylase / Q000235:genetics
D004268:DNA-Binding Proteins; D051739:Microphthalmia-Associated Transcription Factor; C497235:Mitf protein, mouse; C509190:Tpsb2 protein, mouse; D014157:Transcription Factors; D012701:Serotonin; D014365:Tryptophan Hydroxylase; D053804:Granzymes; C509214:Gzmb protein, mouse; D012697:Serine Endopeptidases; C509222:Tpsab1 protein, mouse; D053802:Tryptases; D006640:Histidine Decarboxylase