Abstract
Transcription in all living organisms is accomplished by multi-subunit RNA polymerases (msRNAPs). msRNAPs are highly conserved in evolution and invariably share a ∼400 kDa five-subunit catalytic core. Here we characterize a hypothetical ∼100 kDa single-chain protein, YonO, encoded by the SPβ prophage of Bacillus subtilis. YonO shares very distant homology with msRNAPs, but no homology with single-subunit polymerases. We show that despite homology to only a few amino acids of msRNAP, and the absence of most of the conserved domains, YonO is a highly processive DNA-dependent RNA polymerase. We demonstrate that YonO is a bona fide RNAP of the SPβ bacteriophage that specifically transcribes its late genes, and thus represents a novel type of bacteriophage RNAPs. YonO and related proteins present in various bacteria and bacteriophages have diverged from msRNAPs before the Last Universal Common Ancestor, and, thus, may resemble the single-subunit ancestor of all msRNAPs.
Highlights
Transcription in all living organisms is accomplished by multi-subunit RNA polymerases. msRNAPs are highly conserved in evolution and invariably share a B400 kDa five-subunit catalytic core
The msRNAP evolved before the Last Universal Common Ancestor, and already had a 5-subunit (2a, b, b0, o in bacterial nomenclature) catalytic core including most of the domains that are believed to be essential for its functions[1,2,3,4]
We found that YonO readily formed stable complexes with the RNA–DNA hybrid, a property expected from RNAP (Fig. 1b)
Summary
Transcription in all living organisms is accomplished by multi-subunit RNA polymerases (msRNAPs). msRNAPs are highly conserved in evolution and invariably share a B400 kDa five-subunit catalytic core. Bioinformatic and, later, structural analyses revealed a group of single-subunit proteins that are very distant relatives of msRNAP and which must have diverged from the msRNAP branch far before the LUCA This group includes the eukaryotic RNA-dependent RNAPs involved in post-transcriptional gene silencing, and several hypothetical proteins present in the prophages of some firmicutes, and in the main genomes of many firmicutes and cyanobacteria[3,7,8,9]. Structural predictions and structural analysis have indicated the presence of these domains in YonO-like proteins and eukaryotic RNA-dependent RNAP, respectively[3,7,8,9,10] These domains carry several absolutely conserved amino acids that participate in binding of the catalytic metal and incoming nucleotides, and are believed to be the most ancient domains of msRNAPs3,7–9 (Fig. 1a). We studied YonO, both in vivo and in vitro, and show that it is a new type of specific and highly processive DNA-dependent RNAP and a bona fide RNAP of the SPb bacteriophage of B. subtilis
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