Abstract
Ribotoxins cleave essential RNAs for cell killing, and RNA repair neutralizes the damage inflicted by ribotoxins for cell survival. Here we report a new bacterial RNA repair complex that performs RNA repair linked to immunity. This new RNA repair complex is a 270-kDa heterohexamer composed of three proteins—Pnkp1, Rnl and Hen1—that are required to repair ribotoxin-cleaved RNA in vitro. The crystal structure of the complex reveals the molecular architecture of the heterohexamer as two rhomboid-shaped ring structures of Pnkp1–Rnl–Hen1 heterotrimer fused at the Pnkp1 dimer interface. The four active sites required for RNA repair are located on the inner rim of each ring. The architecture and the locations of the active sites of the Pnkp1–Rnl–Hen1 heterohexamer suggest an ordered series of repair reactions at the broken RNA ends that confer immunity to recurrent damage.
Highlights
Ribotoxins cleave essential RNAs for cell killing, and RNA repair neutralizes the damage inflicted by ribotoxins for cell survival
Building on the study by Martin and Shuman[21], we discovered a bacterial RNA repair complex comprised of two proteins named Pnkp and Hen[1], which form a heterotetramer in vitro[22] (Supplementary Fig. 2b)
In addition to the three enzymatic activities seen in the T4 RNA repair system, the Pnkp–Hen[1] RNA repair complex possesses a fourth enzymatic activity in Hen[1], which carries out methylation of the 20-OH group that is responsible for the RNA cleavage (Supplementary Fig. 1)
Summary
Ribotoxins cleave essential RNAs for cell killing, and RNA repair neutralizes the damage inflicted by ribotoxins for cell survival. Bioinformatic analysis of 2,181 prokaryotic genomes revealed an estimated B10,000 toxins belonging to toxin-antitoxin systems[9], and a similar number of secreted toxins were predicted in the form of polymorphic toxins[8] These reasons suggest that ribotoxins might constitute the biggest percentage of toxins in living organisms[10,11]. Building on the study by Martin and Shuman[21], we discovered a bacterial RNA repair complex comprised of two proteins named Pnkp and Hen[1], which form a heterotetramer in vitro[22] (Supplementary Fig. 2b). Our study revealed that the N-terminal domain of Hen[1] activates the ligase activity of Pnkp by locking the insertion domain of Pnkp into a conformation that allows access to the ligase pocket[24] Such an arrangement for Hen1-dependent RNA repair guarantees Hen[1] the opportunity to carry out 20-O-methylation during RNA repair
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