Transcription of cloned Xenopus laevis ribosomal DNA microinjected into Xenopus oocytes, and the identification of an RNA polymerase I promoter

Abstract

Transcription of a cloned Xenopus laevis ribosomal DNA (rDNA) fragment, microinjected into Xenopus oocytes, is initiated at the in vivo 40S pre-ribosomal RNA (pre-rRNA) site (±2 bp) by RNA polymerase I. An X. laevis RNA polymerase I promoter has been mapped by studying the transcription of in vitro rDNA mutants in the oocyte system. The active promoter lies within the DNA segment beginning 145 bp upstream, and most probably ending 16 bp downstream, from the 40S pre-rRNA initiation site (−145 bp to +16 bp). Furthermore, active promoter elements lie more than 35 bp upstream from the initiation site (−35 bp). The X. laevis RNA polymerase I promoter therefore lies mainly upstream from the 40S pre-rRNA initiation site. Independent deletion of three adjacent rDNA segments lying between −61 and +16 bp reduces promoter activity by a factor of more than 16. The central of these “null” deletions removes an oligo(T) 6 motif at −27 bp that is in an analogous position to the Goldberg-Hogness (TATA) box of RNA polymerase II promoters.