Np₄A alarmones function in bacteria as precursors to RNA caps

Stresses that increase the cellular concentration of dinucleoside tetraphosphates (Np₄Ns) have recently been shown to impact RNA degradation by inducing nucleoside tetraphosphate (Np₄) capping of bacterial transcripts. However, neither the mechanism by which such caps are acquired nor the function of Np₄Ns in bacteria is known. Here we report that promoter sequence changes upstream of the site of transcription initiation similarly affect both the efficiency with which Escherichia coli RNA polymerase incorporates dinucleoside polyphosphates at the 5' end of nascent transcripts in vitro and the percentage of transcripts that are Np₄-capped in E. coli, clear evidence for Np₄ cap acquisition by Np₄N incorporation during transcription initiation in bacterial cells. E. coli RNA polymerase initiates transcription more efficiently with Np₄As than with ATP, particularly when the coding strand nucleotide that immediately precedes the initiation site is a purine. Together, these findings indicate that Np₄Ns function in bacteria as precursors to Np₄ caps and that RNA polymerase has evolved a predilection for synthesizing capped RNA whenever such precursors are abundant.