A NOVEL 43 KD PROTEIN BINDS A CONSERVED MAMMALIAN CACCC MOTIF WITHIN THE DROSOPHILA RAS2 ROP BIDIRECTIONAL PROMOTER/

The Drosophila ras2 promoter is an authentic bidirectional promoter go verning the expression of both the Dras2 and rop genes by a single mec hanism. Characterisation of the Dras2/rop promoter has revealed that a unitary complex (M) interacts with two promoter sub-domains (regions A and B). Two distinct transcription factors (factors A and B),which m ake up the major complex (M), bind regions A and B, respectively. We h ave analyzed the putative CACCC element and AP-1-Iike sequence contain ed within region B (-41 to -20) of the Dras2/rop promoter. It was foun d that AP-1 is not involved in Dras2 expression as is the case for the human Ha-ras1 gene. The entire CACCC motif (-34 to -21) shares 83% ho mology with the conserved mammalian element. Detailed mutational analy sis has however revealed that the CACCC core sequence (-27 to -23) is vital for Dras2/rop recognition by factor B. The cytosine residues at positions -27, -25, -24 and -23 were observed to play a critical role in factor B recognition. Factor B has been purified as a 43 kD polypep tide as measured by SDS-PAGE and the relative mass was confirmed by ph oto-chemical crosslinking. Our findings are the first report of the co nservation of the mammalian CACCC motif in Drosophila.