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Molecular and Cellular Biology, November 2003, p. 8295-8305, Vol. 23, No. 22
0270-7306/03/$08.00+0     DOI: 10.1128/MCB.23.22.8295-8305.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

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

Keizo Nishikawa,1,2 Makoto Kobayashi,1,2* Atsuko Masumi,3 Susan E. Lyons,4 Brant M. Weinstein,5 P. Paul Liu,6 and Masayuki Yamamoto1,2

Center for Tsukuba Advanced Research Alliance and Institute of Basic Medical Sciences, University of Tsukuba, Tsukuba 305-8577,1 Yamamoto Environmental Response Project, ERATO, Japan Science and Technology Corporation, Tsukuba, Ibaraki 300-2635,2 Department of Safety Research on Biologics, National Institute of Infectious Disease, Musashimurayama 208-0011, Japan,3 Department of Internal Medicine, University of Michigan, Ann Arbor, Michigan 48109,4 National Institute of Child Health and Human Development,5 National Human Genome Research Institute, National Institutes of Health, Bethesda, Maryland 208926

Received 20 March 2003/ Returned for modification 24 April 2003/ Accepted 6 August 2003

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.

* Corresponding author. Mailing address: Center for TARA, University of Tsukuba, Tsukuba 305-8577, Japan. Phone: 81-29-847-5195. Fax: 81-29-847-9195. E-mail: kobayash{at}tara.tsukuba.ac.jp.

Molecular and Cellular Biology, November 2003, p. 8295-8305, Vol. 23, No. 22
0022-538X/03/$08.00+0     DOI: 10.1128/MCB.23.22.8295-8305.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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