The mechanism of DNA replication primer synthesis by RNA polymerase

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Abstract
Accurate DNA replication is vital to reproduction in all living organisms. Three papers in this issue and a new Web Focus ( http://tinyurl.com/e3ecg ) present answers to long-standing questions about what goes on at DNA replication forks to ensure this accuracy. Heller and Marians throw light on the fact that even heavily damaged DNA is replicated at high speed. They find that bacterial replication restart systems can prime both leading and lagging DNA strands via DnaG primase. This contradicts the accepted view that leading-strand synthesis is necessarily continuous, and may force a re-evaluation of models for initiation of chromosome replication. Zenkin et al. tackled the mystery of how a short transcript synthesized by RNA polymerase can serve as a primer for DNA replication. The answer lies in a previously unknown transcription elongation complex that may also link DNA replication and transcription machineries. And Lee et al. tackled the matter of how the very different processes taking place on leading and lagging DNA strands are synchronized. As primer synthesis proceeds, DNA primase acts as a molecular brake on the leading-strand polymerase during slow enzymatic steps on the lagging strand. RNA primers for DNA replication are usually synthesized by specialized enzymes, the primases1. However, some replication systems have evolved to use cellular DNA-dependent RNA polymerase for primer synthesis1,2. The main requirement for the replication primer, an exposed RNA 3′ end annealed to the DNA template, is not compatible with known conformations of the transcription elongation complex3, raising a question of how the priming is achieved. Here we show that a previously unrecognized kind of transcription complex is formed during RNA polymerase-catalysed synthesis of the M13 bacteriophage replication primer. The complex contains an overextended RNA–DNA hybrid bound in the RNA-polymerase trough that is normally occupied by downstream double-stranded DNA, thus leaving the 3′ end of the RNA available for interaction with DNA polymerase. Transcription complexes with similar topology may prime the replication of other bacterial mobile elements and may regulate transcription elongation under conditions that favour the formation of an extended RNA–DNA hybrid.