The oxidoreductase PYROXD1 uses NAD(P)+ as an antioxidant to sustain tRNA ligase activity in pre-tRNA splicing and unfolded protein response

• Published in: Molecular cell 2021-06, Vol.81 (12), p.2520-2532.e16
• Main Authors: Asanović, Igor, Strandback, Emilia, Kroupova, Alena, Pasajlic, Djurdja, Meinhart, Anton, Tsung-Pin, Pai, Djokovic, Nemanja, Anrather, Dorothea, Schuetz, Thomas, Suskiewicz, Marcin Józef, Sillamaa, Sirelin, Köcher, Thomas, Beveridge, Rebecca, Nikolic, Katarina, Schleiffer, Alexander, Jinek, Martin, Hartl, Markus, Clausen, Tim, Penninger, Josef, Macheroux, Peter, Weitzer, Stefan, Martinez, Javier
• Format: Article
• Language: English
• Subjects: Animals; Antioxidants - physiology; Catalytic Domain; copper; Female; HeLa Cells; Humans; Male; Medicin och hälsovetenskap; metalloenzyme; Mice; Mice, Inbred C57BL; Mice, Knockout; myopathy; NAD - metabolism; NADH; NADP - metabolism; NADPH; Oxidation-Reduction; oxidative stress; oxidoreductase; Oxidoreductases - metabolism; Oxidoreductases Acting on Sulfur Group Donors - metabolism; Oxidoreductases Acting on Sulfur Group Donors - physiology; pre-tRNA splicing; PYROXD1; RNA Ligase (ATP) - chemistry; RNA Ligase (ATP) - genetics; RNA Ligase (ATP) - metabolism; RNA Splicing - genetics; RNA Splicing - physiology; RNA, Transfer - metabolism; RtcB; tRNA ligase complex; Unfolded Protein Response - physiology; UPR; X-Box Binding Protein 1 - metabolism
• ISSN: 1097-2765; 1097-4164; 1097-4164
• DOI: 10.1016/j.molcel.2021.04.007

The tRNA ligase complex (tRNA-LC) splices precursor tRNAs (pre-tRNA), and Xbp1-mRNA during the unfolded protein response (UPR). In aerobic conditions, a cysteine residue bound to two metal ions in its ancient, catalytic subunit RTCB could make the tRNA-LC susceptible to oxidative inactivation. Here, we confirm this hypothesis and reveal a co-evolutionary association between the tRNA-LC and PYROXD1, a conserved and essential oxidoreductase. We reveal that PYROXD1 preserves the activity of the mammalian tRNA-LC in pre-tRNA splicing and UPR. PYROXD1 binds the tRNA-LC in the presence of NAD(P)H and converts RTCB-bound NAD(P)H into NAD(P)+, a typical oxidative co-enzyme. However, NAD(P)+ here acts as an antioxidant and protects the tRNA-LC from oxidative inactivation, which is dependent on copper ions. Genetic variants of PYROXD1 that cause human myopathies only partially support tRNA-LC activity. Thus, we establish the tRNA-LC as an oxidation-sensitive metalloenzyme, safeguarded by the flavoprotein PYROXD1 through an unexpected redox mechanism. [Display omitted] •The RNA ligase RTCB is susceptible to copper-dependent oxidative inactivation•The flavoprotein PYROXD1 uses NAD(P)+ to safeguard mammalian RTCB against oxidation•Depletion of PYROXD1 impairs pre-tRNA splicing and the unfolded protein response•Myopathy-causing genetic variants of PYROXD1 fail to preserve the activity of RTCB How did enzymes from Earth’s ancient anaerobic history, such as the RNA ligase RTCB, adapt to modern, aerobic environments? Asanovic et al. revealed an unexpected solution: RTCB co-evolved with a dedicated oxidoreductase, PYROXD1, which is linked to myopathies in humans, and paradoxically uses the principal prooxidative cofactor of the cell, NAD(P)+, as the “private” antioxidative protector of RTCB.