Transcription of the human c-Src promoter is dependent on Sp1, a novel pyrimidine binding factor SPy, and can be inhibited by triplex-forming oligonucleotides

The tyrosine kinase pp60(C-src) has been implicated in the regulation of nu merous normal physiological processes as well the development of several hu man cancers. However, the mechanisms regulating its expression have not bee n addressed. In the present study, we report the presence of two Sp1/Sp3 bi nding sites and three polypurine:polypyrimidine (Pu:Py) tracts in the c-Src promoter that are essential for controlling expression. We demonstrate tha t Sp1, but not Sp3, is capable of activating the c-Src promoter and that Sp 3 is also capable of inhibiting Sp1-mediated transactivation. The presence of multiple Pu:Py tracts conferred S1 sensitivity on plasmids in vitro, sug gesting they are capable of adopting non B-DNA conformations. These tracts specifically bind a nuclear factor we named SPy (Src pyrimidine binding fac tor), which demonstrates both novel double- and single-stranded binding spe cificities. Mutations eliminating SPy binding compromised Src transcription al activity, especially in concert with additional mutations affecting Sp1 binding, suggesting the two factors may cooperate in regulating c-Src expre ssion. Finally, we demonstrate that tripler-forming oligonucleotides design ed to target both Sp1 and Spy binding sites can down-regulate c-Src express ion in vitro, suggesting a potential therapeutic approach to controlling c- Src expression in diseases where aberrant expression or activity has been d ocumented.