title: The histone-like protein HU does not obstruct movement of T7 RNA polymerase in Escherichia coli cells but stimulates its activity.
abstract: In vivo, RNA polymerases (RNAPs) do not transcribe naked DNA but do transcribe protein-associated DNA. Studies with the model enzyme T7 RNAP have shown that, in eukaryotic cells or in vitro, nucleosomes can inhibit both transcription initiation and elongation. We examine here whether the presence of HU, one of the major histone-like proteins in Escherichia coli cells (the genuine milieu for T7 RNAP) affects its activity. An engineered lac operon fused to the T7 late promoter was introduced into the chromosome of T7 RNAP-producing strains that either overexpress HU or lack it. The flows of RNAP that enter and exit this operon were compared with regard to the content of HU. We found that the fraction of T7 RNAP molecules that do not reach the end of the lac operon (ca. 15%) is the same whether the host cells overexpressed HU or lacked it: thus, the enzyme either freely displaces HU or transcribes through it. However, in these cells, the transcript yield was increased when HU is overexpressed and decreased in the hup mutants, presumably reflecting changes in DNA supercoiling. Thus, in contrast to eukaryotic nucleosomes, HU does not impair T7 RNAP activity but has a stimulatory effect. Finally, our results suggest that HU can also influence mRNA stability in vivo.
authors.affiliation: Laboratoire de Physiologie Bactérienne (CNRS, UPR 9073), Institut de Biologie Physico-Chimique, 75005 Paris, France.
authors.forename: Pilar
authors.initials: P
authors.lastname: Morales
authors.forename: Josette
authors.initials: J
authors.lastname: Rouviere-Yaniv
authors.forename: Marc
authors.initials: M
authors.lastname: Dreyfus
mesh_terms: D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D017123:Bacteriophage T7 / Q000235:genetics* / Q000378:metabolism; D015152:Blotting, Northern; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D012321:DNA-Directed RNA Polymerases / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism*; D009154:Mutation; D012333:RNA, Messenger / Q000235:genetics; D014158:Transcription, Genetic; D014764:Viral Proteins
chemical_list: D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; C031908:H-NS protein, bacteria; D012333:RNA, Messenger; D014764:Viral Proteins; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases