Similar to Drosophila, murine Polycomb-group (Pc-G) genes regulate anterior-posterior patterning of segmented axial structures by transcriptional repression of homeotic gene expression. eed (embryonic ectoderm development) is a recently isolated member of this group. It encodes a 441-amino acid protein with five WD motifs highly homologous to Drosophila ESC (Extra Sex Combs). Functional conservation between eed and esc has been inferred from posterior homeotic transformations along the axial skeleton in mice carrying a hypomorphic eed allele. However, eed is acting earlier and more globally during development than predicted from esc function. It has been hypothesized that additional developmental function(s) of Eed in mammals may reside in the amino terminus, the region of greatest sequence divergence between Eed and ESC. Outlined here are a series of experiments aimed toward understanding how eed exerts a broad range of biological effects. Embryo manipulation through chimera production, epiblast orthotopic transplants and transplantation of the early gastrula organizer will address whether the mutant epiblast is capable of producing and/or responding to axial organizing signals. Lineage analysis will address whether a fate map can be established for the wild-type blastocyst stage embryo and whether eed alters this clonal fate. Despite the ubiquitous presence of eed expression, the phenotype so far described in ice does not predict global alterations of Hox gene expression. Detailed temporal and spatial analyses of Hox genes in mutant embryos will address mechanisms of initiation and maintenance of Hox expression boundaries. Ubiquitous and tissue specific transgenic experiments both in mice and flies will address functional differences between the species as well as regional differences within each species. Absence of direct DNA binding and indirect evidence of chromatin changes associated with Pc-G protein-mediated repression prompted the hypothesis that Pc-G proteins form distinct complexes that function epigenetically by modifying higher-order chromatin structure. A biochemical and genetic approach in fly and mouse is proposed to begin to identify members of the Eed Pc-G complex.
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