THE US-1 ELEMENT FROM THE GENE ENCODING RAT POLY(ADP-RIBOSE) POLYMERASE BINDS THE TRANSCRIPTION FACTOR-SP1

By comparing the upstream DNA sequence of the rat and human genes enco ding poly(ADP-ribose) polymerase (PARP), we have defined a 16-bp conse rved region and designated it as US-1 for 'upstream sequence 1'. This element is homologous to the recently described binding site for the t ranscription factor Sp1 in the promoter sequence of the mouse p12 gene which encodes a protease inhibitor. Analyses in gel mobility shift as says revealed that a nuclear protein, produced by all tissue-culture c ells tested, specifically binds the US-1 element. The pattern of shift ed DNA . protein complexes obtained was strikingly similar to that for Sp1, which is supported by the positive displacement of these complex es by an oligomer containing the Sp1 binding site in gel shift competi tion experiments. Replacement of the Sp1 binding site from the basal p romoter of the mouse p12 gene by the rPARP US-1 element did not result in any significant variations in the level of expression of the chlor amphenicol acetyltransferase (CAT) reporter gene upon transient transf ection of tissue-culture cells. However, when point mutations are intr oduced in the US-1 element in a similar substitution experiment, a sig nificant reduction in CAT gene expression could be observed. These dat a are consistent with Sp1 interacting with the US1 element. Results fr om DNase I footprinting experiments clearly indicated that purified Sp 1 not only binds to the US-1 element but also to four other closely lo cated cis-acting sites scattered in the promoter of the rat PARP gene, therefore suggesting that Spl is likely to modulate strongly the expr ession of that gene in different tissues.