Functional analysis of a 1.6 kb fragment of the vg Polycomb response element (PRE). (A) Score plot showing PRE prediction at the vestigial (vg) locus according to Ringrose et al. . (B) vg locus, showing coding exons (black,) non-coding exons (grey), enhancers, (blue) and downstream PRE (green). BE = boundary enhancer [73, 74]; EE = predicted embryonic muscle enhancer ; QE = quadrant enhancer [73, 74]. (C) DNA motifs in a vg PRE fragment of 1.6 kb. The grey box corresponds to the highest-scoring region shown in (A). ZESTE, GAGA factor/Pipsqueak (GAF/PSQ) and Pleiohomeotic/Pleihomeotic-like (PHO/PHOL) DNA-binding motifs are shown. In addition, the GTGT motif, found to be enriched in many PREs [8, 11] is enriched in the high scoring region and sequences flanking this region. (D) The 1.6 kb vg PRE shows responsiveness to Polycomb. Eye colors of 5-day-old adult male flies are shown. Flies are homozygous for transgenes on chromosome II as indicated above each panel, and heterozygous on chromosome III for either a balancer chromosome (top; TM3,Sb/+) or the PcXL5mutation (bottom;PcXL5/+). The presence of the PcXL5mutation had no effect on the control construct lacking the PRE at site 1 (no PRE, left) or sites 2 and 3 (not shown), but led to a loss of mw repression in flies carrying the vg PRE at all three sites.