Mechanistic basis for oxidative stress protection of the human tRNA ligase complex by the oxidoreductase PYROXD1

Loeff, Luuk; Kroupova, Alena; Asanović, Igor; Boneberg, Franziska M; Pfleiderer, Moritz M.; Ferdigg, Andrè; Ackle, Fabian; Martı́nez, Javier; Jinek, Martin

doi:10.1101/2023.04.06.535761

Cited by 4 publications

(1 citation statement)

References 38 publications

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“…This study demonstrates that the redox sensitivity of the tRNA-LC is a tightly regulated process that involves several factors and impacts pre-tRNA splicing, UPR and ultimately, protein biosynthesis. PYROXD1 has been shown to protect RTCB from oxidation ( Asanovic et al 2021 ) and, recently, to dissociate from RTCB before substrate binding ( Loeff et al 2023 ). Such dissociation offers an opportunity for ROS to oxidize RTCB.…”

Section: Resultsmentioning

confidence: 99%

Thioredoxin regulates the redox state and the activity of the human tRNA ligase complex

Jaksch,

Irnstorfer,

Kalman

et al. 2023

RNA

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The mammalian tRNA ligase complex (tRNA-LC) catalyzes the splicing of intron-containing pre-tRNAs in the nucleus and the splicing ofXBP1mRNA during the unfolded protein response (UPR) in the cytoplasm. We recently reported that the tRNA-LC coevolved with PYROXD1, an essential oxidoreductase that protects the catalytic cysteine of RTCB, the catalytic subunit of the tRNA-LC, against aerobic oxidation. In this study, we show that the oxidoreductase Thioredoxin (TRX) preserves the enzymatic activity of RTCB under otherwise inhibiting concentrations of oxidants. TRX physically interacts with oxidized RTCB, and reduces and reactivates RTCB through the action of its redox-active cysteine pair. We further show that TRX interacts with RTCB at late stages of UPR. Since the interaction requires oxidative conditions, our findings suggest that prolonged UPR generates reactive oxygen species. Thus, our results support a functional role for TRX in securing and repairing the active site of the tRNA-LC, thereby allowing pre-tRNA splicing and UPR to occur when cells encounter mild, but still inhibitory levels of reactive oxygen species.

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Section: Resultsmentioning

confidence: 99%

Thioredoxin regulates the redox state and the activity of the human tRNA ligase complex

Jaksch,

Irnstorfer,

Kalman

et al. 2023

RNA

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Structural and mechanistic insights into activation of the human RNA ligase RTCB by Archease

Gerber,

Morales Guzmán,

Worf

et al. 2024

Nat Commun

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RNA ligases of the RTCB-type play an essential role in tRNA splicing, the unfolded protein response and RNA repair. RTCB is the catalytic subunit of the pentameric human tRNA ligase complex. RNA ligation by the tRNA ligase complex requires GTP-dependent activation of RTCB. This active site guanylylation reaction relies on the activation factor Archease. The mechanistic interplay between both proteins has remained unknown. Here, we report a biochemical and structural analysis of the human RTCB-Archease complex in the pre- and post-activation state. Archease reaches into the active site of RTCB and promotes the formation of a covalent RTCB-GMP intermediate through coordination of GTP and metal ions. During the activation reaction, Archease prevents futile RNA substrate binding to RTCB. Moreover, monomer structures of Archease and RTCB reveal additional states within the RNA ligation mechanism. Taken together, we present structural snapshots along the reaction cycle of the human tRNA ligase.

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Thioredoxin regulates the redox state and the activity of the human tRNA ligase complex

Jaksch,

Irnstorfer,

Kalman

et al. 2023

Preprint

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The mammalian tRNA ligase complex (tRNA-LC) catalyzes the splicing of intron-containing pre-tRNAs in the nucleus and the splicing of XBP1 mRNA during the unfolded protein response (UPR) in the cytoplasm. We recently reported that the tRNA-LC co-evolved with PYROXD1, an essential oxidoreductase that protects the catalytic cysteine of RTCB, the catalytic subunit of the tRNA-LC, against aerobic oxidation. In this study we show that the oxidoreductase Thioredoxin (TRX) preserves the enzymatic activity of RTCB under otherwise inhibiting concentrations of oxidants. TRX physically interacts with oxidized RTCB, and reduces and re-activates RTCB through the action of its redox-active cysteine pair. We further show that TRX interacts with RTCB at late stages of UPR. Since the interaction requires oxidative conditions, our findings suggest that prolonged UPR generates reactive oxygen species. Thus, our results support a functional role for TRX in securing and repairing the active site of the tRNA-LC, thereby allowing pre-tRNA splicing and UPR to occur when cells encounter mild, but still inhibitory levels of reactive oxygen species.

show abstract

Mechanistic basis for oxidative stress protection of the human tRNA ligase complex by the oxidoreductase PYROXD1

Cited by 4 publications

References 38 publications

Thioredoxin regulates the redox state and the activity of the human tRNA ligase complex

Thioredoxin regulates the redox state and the activity of the human tRNA ligase complex

Structural and mechanistic insights into activation of the human RNA ligase RTCB by Archease

Thioredoxin regulates the redox state and the activity of the human tRNA ligase complex

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