Self-Association of Gata1 Enhances Transcriptional Activity In Vivo in Zebra Fish Embryos

Abstract

Gata1 is a prototype transcription factor that regulates hematopoiesis, yet the molecular mechanisms by which Gata1 transactivates its target genes in vivo remain unclear. We previously showed, in transgenic zebra fish, that Gata1 autoregulates its own expression. In this study, we characterized the molecular mechanisms for this autoregulation by using mutations in the Gata1 protein which impair autoregulation. Of the tested mutations, replacement of six lysine residues with alanine (Gata1KA6), which inhibited self-association activity of Gata1, reduced the Gata1-dependent induction of reporter gene expression driven by the zebra fish gata1 hematopoietic regulatory domain (gata1 HRD). Furthermore, overexpression of wild-type Gata1 but not Gata1KA6 rescued the expression of Gata1 downstream genes in vlad tepes, a germ line gata1 mutant fish. Interestingly, both GATA sites in the double GATA motif in gata1 HRD were critical for the promoter activity and for binding of the self-associated Gata1 complex, whereas only the 3′-GATA site was required for Gata1 monomer binding. These results thus provide the first in vivo evidence that the ability of Gata1 to self-associate critically contributes to the autoregulation of the gata1 gene.

ACKNOWLEDGMENTS

We thank Toshiko Arai, Ayako Hayashi, and Toyoko Kinoshita for help with fish maintenance and Norio Suzuki, Yaeko Takagi, Hitoshi Osanai, and Takafumi Suzuki for help and discussion. We also thank Igor B. Dawid, Kazuhiko Igarashi, Kinuko Ohneda, and Ritsuko Shimizu for critical reading of the manuscript and Kyosuke Nagata and Yasutake Katoh for the gift of E. coli strains.

This work was supported by grants-in-aid from the NOVARTIS Foundation (Japan) for the Promotion of Science, the Japan Society for Promotion of Sciences (JSPS-RFTF), the Japan Science and Technology Corporation (ERATO), the Naito Foundation, and the Ministry of Education, Science, Sports and Culture of Japan.