Isolation of transcription factors that discriminate between different promoters recognized by RNA polymerase II

Abstract

A new fractionation procedure separates a wholecell HeLa extract into three components required for accurate in vitro transcription. One component (Spl) is a promoter-specific factor required for transcription of the SV40 early and late promoters, but not for transcription of other promoters we have tested. The second component (Sp2) is a general factor required for transcription of the SV40 promoters and a series of others, including the adeno-virus 2 major late promoter, the human β-globin promoter and the avian sarcoma virus promoter. The third component is a fraction containing the endogenous RNA polymerase II. When SV40 and adenovirus templates were present simultaneously in an in vitro transcription reaction, addition of the Spl factor stimulated SV40 early promoter transcription 40-fold, and inhibited adenovirus major late promoter transcription by 40%. This finding suggests that Spl is involved in promoter selection, and is not merely a general transcription stimulatory factor.