Kti12, a PSTK-like tRNA dependent ATPase essential for tRNA modification by Elongator

Posttranscriptional RNA modifications occur in all domains of life. Modifications of anticodon bases are of particular importance for ribosomal decoding and proteome homeostasis. The Elongator complex modifies uridines in the wobble position and is highly conserved in eukaryotes. Despite recent insights into Elongator's architecture, the structure and function of its regulatory factor Kti12 have remained elusive. Here, we present the crystal structure of Kti12's nucleotide hydrolase domain trapped in a transition state of ATP hydrolysis. The structure reveals striking similarities to an O-phosphoseryl-tRNA kinase involved in the selenocysteine pathway. Both proteins employ similar mechanisms of tRNA binding and show tRNASec-dependent ATPase activity. In addition, we demonstrate that Kti12 binds directly to Elongator and that ATP hydrolysis is crucial for Elongator to maintain proper tRNA anticodon modification levels in vivo. In summary, our data reveal a hitherto uncharacterized link between two translational control pathways that regulate selenocysteine incorporation and affect ribosomal tRNA selection via specific tRNA modifications.</p

Structure of the Kti11/Kti13 heterodimer and its double role in modifications of tRNA and eukaryotic elongation factor 2

SummaryThe small, highly conserved Kti11 alias Dph3 protein encoded by the Kluyveromyces lactis killer toxin insensitive gene KTI11/DPH3 is involved in the diphthamide modification of eukaryotic elongation factor 2 and, together with Kti13, in Elongator-dependent tRNA wobble base modifications, thereby affecting the speed and accuracy of protein biosynthesis through two distinct mechanisms. We have solved the crystal structures of Saccharomyces cerevisiae Kti13 and the Kti11/Kti13 heterodimer at 2.4 and 2.9 Å resolution, respectively, and validated interacting residues through mutational analysis in vitro and in vivo. We show that metal coordination by Kti11 and its heterodimerization with Kti13 are essential for both translational control mechanisms. Our structural and functional analyses identify Kti13 as an additional component of the diphthamide modification pathway and provide insight into the molecular mechanisms that allow the Kti11/Kti13 heterodimer to coregulate two consecutive steps in ribosomal protein synthesis

Structural basis for tRNA modification by Elp3 from Dehalococcoides mccartyi

International audienceDuring translation elongation, decoding is based on the recognition of codons by corresponding tRNA anticodon triplets. Molecular mechanisms that regulate global protein synthesis via specific base modifications in tRNA anticodons are receiving increasing attention. The conserved eukaryotic Elongator complex specifically modifies uridines located in the wobble base position of tRNAs. Mutations in Elongator subunits are associated with certain neurodegenerative diseases and cancer. Here we present the crystal structure of D. mccartyi Elp3 (DmcElp3) at 2.15-Å resolution. Our results reveal an unexpected arrangement of Elp3 lysine acetyltransferase (KAT) and radical S-adenosyl methionine (SAM) domains, which share a large interface and form a composite active site and tRNA-binding pocket, with an iron–sulfur cluster located in the dimerization interface of two DmcElp3 molecules. Structure-guided mutagenesis studies of yeast Elp3 confirmed the relevance of our findings for eukaryotic Elp3s and should aid in understanding the cellular functions and pathophysiological roles of Elongator