RECOGNITION OF A HUMAN ARREST SITE IS CONSERVED BETWEEN RNA-POLYMERASE-II AND PROKARYOTIC RNA-POLYMERASES

DNA sequences that arrest transcription by either eukaryotic RNA polym erase II or Escherichia coil RNA polymerase have been identified previ ously. Elongation factors SH and GreB are RNA polymerase-binding prote ins that enable readthrough of arrest sites by these enzymes, respecti vely. This functional similarity has led to general models of elongati on applicable to both eukaryotic and prokaryotic enzymes. Here we have transcribed with phage and bacterial RNA polymerases, a human DNA seq uence previously defined as an arrest site for RNA polymerase II. The phage and bacterial enzymes both respond efficiently to the arrest sig nal in vitro at limiting levels of nucleoside triphosphates. The E. co li polymerase remains in a template-engaged complex for many hours, ca n be isolated, and is potentially active. The enzyme displays a relati vely slow first-order loss of elongation competence as it dwells at th e arrest site. Bacterial RNA polymerase arrested at the human site is reactivated by GreB in the same way that RNA polymerase II arrested at this site is stimulated by SII, Very efficient readthrough can be ach ieved by phage, bacterial, and eukaryotic RNA polymerases in the absen ce of elongation factors if 5-Br-UTP is substituted for UTP. These fin dings provide additional and direct evidence for functional similarity between prokaryotic and eukaryotic transcription elongation and readt hrough mechanisms.