Mammalian ALKBH8 Possesses tRNA Methyltransferase Activity Required for the Biogenesis of Multiple Wobble Uridine Modifications Implicated in Translational Decoding
Abstract
Mammalian ALKBH8 Possesses tRNA Methyltransferase Activity Required for the Biogenesis of Multiple Wobble Uridine Modifications Implicated in Translational Decoding ▿ Lene Songe-Møller 1 , § , Erwin van den Born 2 , § , Vibeke Leihne 2 , § , Cathrine B. Vågbø 3 , § , Terese Kristoffersen 1 , Hans E. Krokan 3 , Finn Kirpekar 4 , Pål Ø. Falnes 1 , 2 , # , * and Arne Klungland 1 , 5 , # , * 1 Center for Molecular Biology and Neuroscience and Institute of Medical Microbiology, Oslo University Hospital, Oslo, Norway 2 Department of Molecular Biosciences, University of Oslo, P.O. Box 1041 Blindern, N-0316 Oslo, Norway 3 Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology, N-7489 Trondheim, Norway 4 Department of Biochemistry and Molecular Biology, University of Southern Denmark, 5230 Odense M, Denmark 5 Institute of Basic Medical Sciences, University of Oslo, P.O. Box 1018 Blindern, N-0315 Oslo, Norway ABSTRACT Uridines in the wobble position of tRNA are almost invariably modified. Modifications can increase the efficiency of codon reading, but they also prevent mistranslation by limiting wobbling. In mammals, several tRNAs have 5-methoxycarbonylmethyluridine (mcm 5 U) or derivatives thereof in the wobble position. Through analysis of tRNA from Alkbh8 −/− mice, we show here that ALKBH8 is a tRNA methyltransferase required for the final step in the biogenesis of mcm 5 U. We also demonstrate that the interaction of ALKBH8 with a small accessory protein, TRM112, is required to form a functional tRNA methyltransferase. Furthermore, prior ALKBH8-mediated methylation is a prerequisite for the thiolation and 2′- O -ribose methylation that form 5-methoxycarbonylmethyl-2-thiouridine (mcm 5 s 2 U) and 5-methoxycarbonylmethyl-2′- O -methyluridine (mcm 5 Um), respectively. Despite the complete loss of all of these uridine modifications, Alkbh8 −/− mice appear normal. However, the selenocysteine-specific tRNA (tRNA Sec ) is aberrantly modified in the Alkbh8 −/− mice, and for the selenoprotein Gpx1, we indeed observed reduced recoding of the UGA stop codon to selenocysteine.