Regulation of the transcription factor GATA-2.

GATA-2, a member of the GATA family of transcription factors, has been shown by gene ablation to be critical for the proliferation of hematopoietic progenitors. Mice null for Gata2 have severe deficits in primitive and definitive hematopoietis, resulting in early embryonic lethality. Previously, it has been demonstrated that a Yeast Artificial Chromosome (YAC) containing 271 kbp of the Gata2 genomic locus is able to rescue the hematopoietic defect of Gata2 null mice. However, these compound heterozygotes (Gata2-/-: YAC) died at birth due to hydroureteronephrosis, suggesting that the YAC lacks regulatory elements required for appropriate expression of GATA-2 in the developing urogenital system, and demonstrating that GATA-2 plays a critical role in urogenital development. Given the widespread expression of Gata2 in a number of different tissues in development, one important question is how expression of this gene is activated and extinguished at specific times and tissues during development. To understand how Gata2 expression is regulated, we attempted to identify a number of the tissue-specific enhancers required for Gata2 expression. First, we identified one enhancer that is sufficient to recapitulate Gata2 midbrain expression. Second, we demonstrated the existence of an endothelial enhancer for Gata2 and identified critical upstream regulators of its activity. Third, we developed a BAC transgenic approach to identify distant urogenital enhancers that were missing from the rescuing YAC transgene. Using this approach, we identified three different urogenital enhancers of Gata2, and demonstrated that two confer spatially-distinct expression patterns in the urogenital mesenchyme. Finally, using Cre/Lox recombination, we created a single transgene spanning more than 400 kbp of the Gata2 locus, which should recapitulate Gata2 expression in both the hematopoietic and urogenital systems, allowing for a complete rescue of GATA-2 deficiency. In summary, we have demonstrated the existence of more than five new tissue-specific enhancers in the Gata2 locus, scattered over more than 300 kbp, and deduced the presence of several others located over 700 kbp away from the structural gene. The data presented here demonstrate the intricate regulatory systems that have evolved to control the expression of genes that play critical roles in the development of multi-cellular organisms.PhDBiological SciencesCellular biologyGeneticsMolecular biologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/124454/2/3138197.pd