Fig. 1. Pleiohomeotic binding sites are required but not sufficient for MCP silencing function. (A) The basic structure of the reporter construct used to assay the silencing activity of MCP fragments (not drawn to scale). The MCP element was replaced with mutant or deletion fragments discussed in the text. (B) Diagram of the MCP fragments. The top line shows the wild-type MCP822 element which is a SalI-XbaI fragment. The numbers represent nucleotide residues corresponding to those indicated in Fig. 7. Open circles denote locations of the putative PHO binding sites. 5MPHO, 4MPHO, PHO62 and PHO604 contain mutations of PHO binding consensus sequence at sites indicated by X. MCP1 is a fragment containing sequences between nucleotides 573 and 675. In the adjacent table, the ‘# lines’ column shows the total number of independent transformant lines carrying the constructs and examined for lacZ expression in the wing and haltere discs; the ‘silenced’ column indicates the number of lines in which expression of lacZ is silenced in the wing disc (ps4 and ps5) and in the anterior compartment of the haltere disc (ps5) (Fig. 2A); the ‘derepressed’ column indicates the number of lines in which lacZ is derepressed in the wing or haltere discs. (C) In vitro binding of PHO protein to the putative PHO604 binding site in MCP. Radioactively labeled oligonucleotide PHO604 (Fig. 7) was incubated with in vitro-synthesized PHO protein (Materials and Methods). Arrow indicates the PHO-oligo complexes. Unlabeled competitors were PHO604, ADF (non-specific sequence control) and MPHO604 (PHO604 site mutated; see Fig. 7).
