| Summary: | Transcription, the first step of gene expression, is accomplished in all domain of life by the multisubunit RNA polymerase (RNAP). Accordingly, the RNAP is an ancient enzyme that is highly conserved in all cellular organisms. However, in-depth bioinformatics has led to the identification of proteins distantly related to the multisubunit RNAP, such as YonO and ORF6 RNAP. Whilst being putative single subunit RNAPs, YonO and ORF6 RNAP are completely unrelated to T7 RNAP. YonO, encoded by the Bacillus subtilis SPβ prophage, has incredibly little similarity to RNAP. Conversely, ORF6 RNAP belonging to the Kluyveromyces lactis killer system contains half of the conserved RNAP active centre. YonO and ORF6 RNAP are potentially novel RNAPs and their characterisation will give new insights into mechanisms of transcription as well as the biology of the organisms which they belong to. Despite lacking most of the conserved RNAP active centre, we have shown YonO is a very efficient DNA dependant RNAP. Striking, unlike all of its multisubunit relatives, YonO is able to initiate transcription on double stranded DNA without accessory factors (such as σ factors in bacteria). Furthermore, our work suggests YonO is expressed during induction of SPβ and functions to transcribe the SPβ late genes. This potentially makes YonO the first bacteriophage single subunit RNAP that resembles the multisubunit RNAP. On the other hand, ORF6 RNAP was very poor at polymerisation, with or without its putative accessory subunit. This suggests additional, currently unknown proteins are required for RNAP activity and potentially a new molecular mechanism of RNA polymerisation. YonO homologues exist in a wide range of bacteria including fermicutes and cyanobacteria. YonO represents a new class of RNAP and our discoveries potentially open up a new area of research as well as being potentially useful for biotechnology and synthetic biology. In contrast, transcription by ORF6 RNAP is more complex than previously thought, with alternative lines of investigation required to identify additional factors that contribute to ORF6 RNAP activity.
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