Yeast transcription factor IID participates in cell-free transcription of a mammalian ribosomal protein TATA-less promoter

Abstract

We analysed transcription of the gene for the ribosomal protein (rp) L32 of the mouse, which is transcribed in mouse L1210 nuclear extracts in vitro. The rpL32 gene lacks a canonical TATA box. Hence it has been suggested that this gene has an alternative transcription pathway not requiring transcription factor IID (TFIID). Selective inactivation of TFIID in nuclear extract completely abolished the transcription of rpL32 in vitro. Selective inactivation was restored by the addition of cloned and purified yeast TFIID (yTFIID), indicating that this TATA-less rpL32 promoter utilizes TFIID for its transcription initiation. Furthermore, addition of an oligonucleotide-containing TATA sequence interfered with the rpL32 transcription and this was overcome by the addition of yTFIID. To further examine the stage of involvement of TFIID in rpL32 transcription, TATA oligonucleotide was added to nuclear extract before and after the formation of the transcription complex. The results reveal that TFIID associates with the pre-initiation complex and that this complex is largely resistant to added TATA oligonucleotide. Our results show, for the first time, that the TATA-less rpL32 gene utilizes TFIID for transcription initiation.