Single-peptide DNA-dependent RNA polymerase homologous to multi-subunit RNA polymerase

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.

Multisubunit RNA Polymerases of Jumbo Bacteriophages

“…During the past two decades, extensive genome sequencing revealed genes coding for distant homologs of cellular RNAPs in the genomes of some eukaryotic viruses, bacteriophages, prophages and likely mobile elements located in the genomes of some Firmicutes and Cyanobacteria , and in fungal killer plasmids [ 16 , 17 , 18 , 19 , 20 , 21 , 22 ]. Some of these genes were shown to encode functional RNAPs [ 23 , 24 , 25 , 26 , 27 , 28 ], while the products of others remain uncharacterized. These partially characterized and non-characterized putative RNAPs are collectively referred to as “non-canonical RNAPs” since they are highly divergent from multisubunit RNAPs of cellular organisms.…”

A distinct lineage of Caudovirales that encodes a deeply branching multi-subunit RNA polymerase

“…β′-subunits 32 ; we were unable to identify any recognizable sequence homology of crAssphage RNAP to the COG0085 and COG0086 Hidden Markov Models (HMMs) we used in this study, however, indicating that the crAssphage enzyme is highly divergent and acquired independently from the mReC. Moreover, other phage have been found to encode a single-subunit YonO protein with similarity to the β′-subunit of RNAP 14 , although once again the highly divergent nature of these proteins hinders detailed phylogenetic analysis. It is not surprising that many divergent enzymes with either sequence or structural homology to RNAP subunits are present in the viral world considering the antiquity of this enzyme; indeed, structural homology has even been noted between RNAP subunits and eukaryotic RNA-dependent RNAP (RdRp) and archaeal replicative DNA polymerase 33 .…”

“…This interpretation must be made cautiously, however; while our concatenated β and β′ RNAP tree indicates that mReC RNAP forms a distinct clade separate from cellular lineages, the rooted β/β′ paralog tree is based on the alignment of highly divergent sequences and does not provide a definitive root. One may postulate an alternative scenario in which the mReC RNAP have an accelerated evolutionary rate that obfuscates phylogenetic analyses and potentially renders the resulting trees unreliable; indeed, other viruses encode YonO proteins or other homologs to RNAP subunits that have diverged considerably and cannot be robustly aligned to cellular homologs 13,14 . RNAP homologs from mReC still retain highly conserved regions that are readily alignable to cellular homologs, however ( Fig.…”

RNA polymerase active centre compensates for the absence of transcription proofreading factors in Cyanobacteria

“…Total RNA was isolated from mid-exponentially growing cultures of E. coli and Synechocystis sp PCC 6803 as described in (55). Quality of RNA was checked by BioAnalyser, sample preparation and sequencing were performed by Vertis Ltd, essentially as described in (24) coli strain and the reference genome were identified using samtools and bcftools (57).…”