SYNERGISTIC INTERACTIONS BETWEEN 2 DISTINCT ELEMENTS OF A REGULATED SPLICING ENHANCER

Regulated alternative splicing of doublesex (dsx) pre-mRNA requires a splicing enhancer designated the dsx repeat element (dsxRE) that conta ins six copies of a 13-nucleotide repeat sequence. Previous studies ha ve shown that the activity of the dsxRE requires the splicing regulato rs Transformer (Tra) and Transformer 2 (Tra2), and one or more members of the SR family of general splicing factors. In this paper we identi fy a purine-rich enhancer (PRE) sequence within the dsxRE, and show th at this element functionally synergizes with the repeat sequences. In vitro binding studies show that the PRE is required for specific bindi ng of Tra2 to the dsxRE, and that Tra and SR proteins bind cooperative ly to the dsxRE in the presence or absence of the PRE. Thus positive c ontrol of dsx pre-mRNA splicing requires the Tra- and Tra2-dependent a ssembly of a multiprotein complex on at least two distinct enhancer el ements.