Structural and mechanistic basis of reiterative transcription initiation

Abstract

Significance In standard transcription initiation, RNA polymerase (RNAP) synthesizes an initial RNA product complementary to the DNA template. In an alternative pathway of transcription initiation, termed “reiterative transcription initiation,” observed at promoters containing homopolymeric sequences at the transcription start site, RNAP synthesizes an initial RNA product having a 5′ sequence containing a variable number, up to tens to hundreds, of nucleotides noncomplementary to the DNA template. Here, using crystallography, cryoelectron microscopy, photocrosslinking, and single-molecule DNA nanomanipulation, we show that RNA extension in reiterative transcription initiation involves slipping of RNA relative to DNA within a short RNA–DNA hybrid and generates RNA that exits RNAP through a previously unobserved RNA path (“alternative RNA path”) and previously unobserved RNA exit (“alternative RNA exit”).