DNA-SEQUENCE REQUIREMENTS FOR TRANSCRIPTIONAL INITIATOR ACTIVITY IN MAMMALIAN-CELLS

A transcriptional initiator (Inr) for mammalian RNA polymerase II can be defined as a DNA sequence element that overlaps a transcription sta rt site and is sufficient for (i) determining the start site location in a promoter that lacks a TATA box and (ii) enhancing the strength of a promoter that contains a TATA box. We have prepared synthetic promo ters containing random nucleotides downstream of Sp1 binding sites to determine the range of DNA sequences that convey Inr activity. Numerou s sequences behaved as functional Inrs in an in vitro transcription as say, but the Inr activities varied dramatically. An examination of the functional elements revealed loose but consistent sequence requiremen ts, with the approximate consensus sequence Py Py A+1 N T/A Py Py. Mos t importantly, almost every functional Inr that has been described fit s into the consensus sequence that we have defined. Although several p roteins have been reported to bind to specific Inrs, manipulation of t hose elements failed to correlate protein binding with Inr activity. T he simplest model to explain these results is that all or most Inrs ar e recognized by a universal binding protein, similar to the functional recognition of all TATA sequences by the same TATA-binding protein. T he previously reported proteins that bind near specific Inr elements m ay augment the strength of an Inr or may impart transcriptional regula tion through an Inr.