Nucleic Acids Research, Vol 25, Issue 23 4730-4739, Copyright © 1997 by Oxford University Press
MC Justice, BP Hogan and AK Vershon
The homeodomain (HD) is a conserved sequence-specific DNA-binding motif
found in many eukaryotic transcriptional regulatory proteins. Despite the
wealth of in vitro data on the mechanism HD proteins use to bind DNA,
comparatively little is known about the roles of individual residues in
these domains in vivo . The Saccharomyces cerevisiae Pho2 protein contains
a HD that shares significant sequence identity with the Drosophila
Engrailed protein. We have used the co-crystal structure of Engrailed as a
model to predict how Pho2 might contact DNA and have examined how
individual residues of the Pho2 HD contribute to transcriptional activation
in vivo and to DNA binding in vitro. Our results demonstrate that Pho2 and
Engrailed share many similar DNA- binding characteristics. However, our
results also show that some highly conserved residues, which contact the
DNA in many HD structures, make relatively small contributions to the
DNA-binding affinity and in vivo activity of the Pho2 protein. We also show
that the N-terminal arm of the Pho2 HD is a critical component in
determining the DNA-binding specificity of the protein and that the
requirements for residues in the N-terminal arm are promoter-dependent for
Pho2 transcriptional activation and DNA binding.
ARTICLES
Homeodomain-DNA interactions of the Pho2 protein are promoter-dependent
Waksman Institute and Department of Molecular Biology and Biochemistry, Rutgers University, Piscataway, NJ 08855, USA.
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