RNA polymerase I passage through nucleosomes depends on its lobe binding subunits

Abstract

RNA polymerase I (Pol I) is a highly efficient enzyme specialized to synthesize most of the ribosomal RNA. After nucleosome deposition at each round of replication the Pol I transcription machinery has to deal with nucleosomal barriers. It was suggested that Pol I-associated factors facilitate chromatin transcription, but it is not known whether Pol I has an intrinsic capacity to transcribe through nucleosomes. Here we used in vitro transcription assays to study purified Pol I of the yeast S. cerevisiae and Pol I mutants in comparison to Pol II and Pol III to pass a nucleosome. Under identical conditions, purified Pol I and Pol III, but not Pol II, were able to transcribe nucleosomal templates. Pol I mutants lacking either the heterodimeric subunit Rpa34.5/Rpa49 or the C-terminal part of the specific subunit Rpa12.2 showed a lower processivity on naked DNA templates, which was even more reduced in the presence of a nucleosome. The contribution of Pol I specific subunit domains to efficient passage through nucleosomes in context with transcription rate and processivity is discussed.