Dinucleoside polyphosphates act as 5'-RNA caps in bacteria

It has been more than 50 years since the discovery of dinucleoside polyphosphates (Np_nNs) and yet their roles and mechanisms of action remain unclear. Here, we show that both methylated and non-methylated Np_nNs serve as RNA caps in Escherichia coli. Np_nNs are excellent substrates for T7 and E. coli RNA polymerases (RNAPs) and efficiently initiate transcription. We demonstrate, that the E. coli enzymes RNA 5'-pyrophosphohydrolase (RppH) and bis(5'-nucleosyl)-tetraphosphatase (ApaH) are able to remove the Np_nN-caps from RNA. ApaH is able to cleave all Np_nN-caps, while RppH is unable to cleave the methylated forms suggesting that the methylation adds an additional layer to RNA stability regulation. Our work introduces a different perspective on the chemical structure of RNA in prokaryotes and on the role of RNA caps. We bring evidence that small molecules, such as Np_nNs are incorporated into RNA and may thus influence the cellular metabolism and RNA turnover.