An eh1-Like Motif in Odd-skipped Mediates Recruitment of Groucho and Repression In Vivo

Abstract

Drosophila Groucho, like its vertebrate Transducin-like Enhancer-of-split homologues, is a corepressor that silences gene expression in numerous developmental settings. Groucho itself does not bind DNA but is recruited to target promoters by associating with a large number of DNA-binding negative transcriptional regulators. These repressors tether Groucho via short conserved polypeptide sequences, of which two have been defined. First, WRPW and related tetrapeptide motifs have been well characterized in several repressors. Second, a motif termed Engrailed homology 1 (eh1) has been found predominantly in homeodomain-containing transcription factors. Here we describe a yeast two-hybrid screen that uncovered physical interactions between Groucho and transcription factors, containing eh1 motifs, with different types of DNA-binding domains. We show that one of these, the zinc finger protein Odd-skipped, requires its eh1-like sequence for repressing specific target genes in segmentation. Comparison between diverse eh1 motifs reveals a bias for the phosphoacceptor amino acids serine and threonine at a fixed position, and a mutational analysis of Odd-skipped indicates that these residues are critical for efficient interactions with Groucho and for repression in vivo. Our data suggest that phosphorylation of these phosphomeric residues, if it occurs, will down-regulate Groucho binding and therefore repression, providing a mechanism for posttranslational control of Groucho-mediated repression.

ACKNOWLEDGMENTS

We thank members of our laboratory and of the Biochemistry Department for continued help and encouragement during this project. In particular, we thank Einat Cinnamon, Peleg Hasson, James Jaynes, Gerardo Jiménez, Oded Meyuhas, and Joel Yisraeli for enjoyable discussions and for insightful comments on the manuscript; Keren Cohen for fly food; and Steve Cohen, Doug Coulter, Ryan Green, Rachel Hoang, James Jaynes, Tommy Kaplan, Judith Lengyel, Hanah Margalit, Bill McGinnis, Tally Naveh-Many, Marcus Noll, Jim Skeath, Eric Wieschaus, and Ernst Wimmer for reagents and fly stocks.

This work was supported by grants from the Israel Science Foundation (501/04), the Israel Cancer Research Fund, the Lejwa Fund for Biochemistry, the United States-Israel Binational Science Foundation (96-108), and the Jan M. and Eugenia Król Charitable Foundation to Z.P. and grants from NIH (EY015718) and PSCoR (NT10005) to A.P.B. R.E.G. is most grateful for support by a Yishayahu Horowiz Foundation Ph.D. scholarship. Z.P. is a Braun Lecturer.