Transcription of yeast DNA by homologous RNA polymerases I and II: selective transcription of ribosomal genes by RNA polymerase I.
Purified yeast DNA was transcribed by homologous RNA polymerases I and II and Escherichia coli RNA polymerase. Transcripts synthesized in vitro were analyzed by molecular hybridization with complementary DNA (cDNA) synthesized from yeast poly(A)-containing mRNA with viral reverse transcriptase and ribosomal DNA labeled in vitro by nick translation with E. coli DNA polymerase I. RNA synthesized by polymerase I and II in the presence of Mn2+ contained sequences complementary to cDNA and rDNA at a frequency consistent with random transcription of the template. Similarly, E. coli RNA polymerase synthesized an apparently random transcript in the presence of either Mn2+ or Mg2+. In contrast to these results, RNA polymerase I but not polymerase II transcripts were markedly enriched in sequences complementary to rDNA when transcription was carried out in the presence of Mg2+. The observed enrichment was 15-30-fold higher than observed for polymerase II or E. coli polymerase transcripts and is consistent with the transcript being comprised of 6-10% ribosomal sequences. These data strongly suggest that RNA polymerase I plays a critical role in selective transcription of ribosomal cistrons.
D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D004789:Enzyme Activation; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D008970:Molecular Weight; D009691:Nucleic Acid Denaturation; D009693:Nucleic Acid Hybridization; D012318:RNA Polymerase I / Q000302:isolation & purification / Q000378:metabolism*; D012319:RNA Polymerase II / Q000302:isolation & purification / Q000378:metabolism*; D012270:Ribosomes / Q000502:physiology*; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D013045:Species Specificity; D013698:Templates, Genetic; D014158:Transcription, Genetic
D008345:Manganese; D012319:RNA Polymerase II; D012321:DNA-Directed RNA Polymerases; D012318:RNA Polymerase I; D008274:Magnesium