Abstract
RtcB enzymes are novel RNA ligases that join 2',3'-cyclic phosphate and 5'-OH ends. The phylogenetic distribution of RtcB points to its candidacy as a tRNA splicing/repair enzyme. Here we show that Escherichia coli RtcB is competent and sufficient for tRNA splicing in vivo by virtue of its ability to complement growth of yeast cells that lack the endogenous "healing/sealing-type" tRNA ligase Trl1. RtcB also protects yeast trl1Δ cells against a fungal ribotoxin that incises the anticodon loop of cellular tRNAs. Moreover, RtcB can replace Trl1 as the catalyst of HAC1 mRNA splicing during the unfolded protein response. Thus, RtcB is a bona fide RNA repair enzyme with broad physiological actions. Biochemical analysis of RtcB highlights the uniqueness of its active site and catalytic mechanism. Our findings draw attention to tRNA ligase as a promising drug target.
Highlights
Escherichia coli RtcB exemplifies a new family of RNA ligases that directly seal 2Ј,3Ј-cyclic phosphate and 5Ј-OH ends [1,2,3]
RtcB homologs purified from archaeal and mammalian cells can seal broken tRNA halves and are thereby imputed to be the catalysts of archaeal and mammalian tRNA splicing [2, 3]. This scenario is complicated by the existence of a yeast/plant-like tRNA splicing pathway in mammalian cells (8 –12) and of analogous yeast-like RNA repair enzymes in many archaeal taxa [13,14,15,16]
We interrogated the tRNA repair function of E. coli RtcB in vivo by complementation of a lethal deletion of the TRL1 gene encoding the tRNA ligase of Saccharomyces cerevisiae [7]
Summary
Escherichia coli RtcB exemplifies a new family of RNA ligases that directly seal 2Ј,3Ј-cyclic phosphate and 5Ј-OH ends [1,2,3]. Yeast and plants rely on a different mechanism of tRNA splicing in which the broken 3Ј and 5Ј ends are healed (converted to a 3Ј-OH/2Ј-PO4 and 5Ј-PO4, respectively) and sealed by a classical ATPdependent RNA ligase [7] (see Fig. 1).
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