Figure 6.

Increased synthesis of monophosphorylated C-initiated transcripts in E. coli cells lacking cytidylate kinase activity. (A) Effect of cytidylate kinase on the percentage of efp 5′ ends that were monophosphorylated in E. coli cells lacking RppH. The phosphorylation state of the 5′ end of efp mRNA (top) and efp-C1A mRNA (bottom) in an isogenic set of E. coli strains was analyzed by PABLO, as in Figure 1, to quantify 5′ termini that were monophosphorylated (MonoP). A bent arrow leads from each RNA to its ligation product. PPase, pyrophosphatase. Representative experiments are shown. (B) Difference in the decay rate of monophosphorylated versus diphosphoryated/triphosphoryated efp mRNA in E. coli cells lacking both cytidylate kinase activity and RppH. Total cellular RNA extracted at time intervals after inhibiting transcription with rifampicin was analyzed by PABLO, as in Figure 1, to differentiate between efp 5′ ends that were monophosphorylated (ligated) or diphosphoryated/triphosphoryated (unligated), and the resulting band intensities were plotted semilogarithmically and analyzed by linear regression. Representative experiments are shown.