The role of the Escherichia coli RNA pyrophosphohydrolase (RppH) in RNA metabolism
Bowden, Katherine Elaine
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RNA turnover in Escherichia coli was originally thought to initiate through the action of ribonucleases. It was not until the discovery of the “decapping” enzyme RNA pyrophosphohydrolase (RppH) in Escherichia coli that this hypothesis was called into question. Prior to the experiments described here, RppH had only been implicated in mRNA decay and hybrid jamming. Here, we have analyzed the role of RppH in tRNA maturation and called into question the original idea that the catalytic activity of RppH was required to promote endonucleolytic cleavages by RNase E. We have shown that this is actually not the case for tRNA processing. In contrast, RppH activity is required for the 5’-maturation of certain tRNAs by RNase P. In addition, we saw that this effect was regulated by RNase PH, a 3’→5’ exoribonuclease. After analysis of tRNA maturation, we examined the role of RppH on regulation of the entire transcriptome. Previous microarray analysis has been limited by the use of probes for only open-reading frames (ORFs). Here, through the use of a tiling microarray, which provides probes across the entire transcriptome, we have discovered that RppH is involved in regulation of the flagellar gene regulatory network. Based on these findings, we have shown that E. coli carrying a rppHΔ754 mutation is hypermotile and restores motility to the nonmotile ΔapaH mutant strain. This work has attempted to uncover processes that RppH has not been previously known to play a role. Although more insight has been gained, there still remains a question as to how RNase PH is involved in RppH-dependent regulation. In addition, it has become apparent that the activity of RppH is much more complex than originally thought.