Dinucleoside polyphosphates act as 5’-RNA caps in Escherichia coli

Abstract

Dinucleoside polyphosphates (Np_nNs), discovered more than 50 years ago,¹ are pleiotropic molecules present in almost all types of cells.² It has been shown that their intracellular concentration can under stress conditions increase from the μM to mM range^2,3. However, the cellular roles and mechanisms of action of Np_nNs are still speculative^4,5. They have never been considered as part of the RNA, even though they have similar chemical structures as already known RNA caps, such as the nicotinamide adenine dinucleotide (NAD)^6–8 and 7-methylguanylate cap⁹. Here, we show that both methylated and non-methylated Np_nNs serve as RNA caps in Escherichia coli (E. coli). Np_nNs are excellent substrates for T7 and E. coli RNA polymerases (RNAP) and efficiently initiate transcription. Further, we demonstrate that the E. coli decapping enzyme RNA 5’ pyrophosphohydrolase (RppH) is able to remove the Np_nNs-cap from the RNA. RppH was, however, not able to cleave the methylated forms of the Np_nN-caps, suggesting that the methylation adds an additional layer to the RNA stability regulation. Our work introduces an original perspective on the chemical structure of RNA in prokaryotes and the function of RNA caps. This is the first evidence that small molecules like Np_nNs can act in cells via their incorporation into RNA and influence the cellular metabolism.