Architecture of the yeast Elongator complex.

Maria I Dauden,Karin D Breunig,Sebastian Glatt,Ambroise Desfosses,Martin Beck,Bertrand Séraphin,Alessandro Ori,Christoph W Müller,Osita F Onuma,Olga Kolaj‐Robin,Jan Kosinski,Niklas A Hoffmann,Carsten Sachse,Celine Faux

doi:10.15252/embr.201643353

Abstract

The highly conserved eukaryotic Elongator complex performs specific chemical modifications on wobble base uridines of tRNAs, which are essential for proteome stability and homeostasis. The complex is formed by six individual subunits (Elp1‐6) that are all equally important for its tRNA modification activity. However, its overall architecture and the detailed reaction mechanism remain elusive. Here, we report the structures of the fully assembled yeast Elongator and the Elp123 sub‐complex solved by an integrative structure determination approach showing that two copies of the Elp1, Elp2, and Elp3 subunits form a two‐lobed scaffold, which binds Elp456 asymmetrically. Our topological models are consistent with previous studies on individual subunits and further validated by complementary biochemical analyses. Our study provides a structural framework on how the tRNA modification activity is carried out by Elongator.

Highlights

During the elongation phase of the ribosome-mediated translation process, transient pausing events support proper domain folding of the nascent polypeptide chains, which gain their threedimensional conformations immediately after they have left the exit tunnel of the ribosomes, a process sometimes facilitated by chaperones [1,2,3]
In an initial attempt of assembling the Elongator complex from individually purified proteins, expressed heterologously in Escherichia coli, we were able to observe a direct interaction between Elp1 and Elp3, and Elp1 and Elp456
Consistent with substoichiometric cellular amounts of Elp456 in vivo [28,36,37], purifications of Elp1-tandem-affinity purification (TAP) resulted in an excess of Elp123 sub-complex [20], whereas Elp6-TAP purifications resulted in reduced quantities but yielded highly pure, complete, and stoichiometric Elongator complex

Summary

Introduction

During the elongation phase of the ribosome-mediated translation process, transient pausing events support proper domain folding of the nascent polypeptide chains, which gain their threedimensional conformations immediately after they have left the exit tunnel of the ribosomes, a process sometimes facilitated by chaperones [1,2,3]. Previous studies indicated that specific base modifications in the wobble base position of tRNAs are crucial to maintain these highly dynamic and complex mechanisms. Because they influence the recognition rate and affinity between incoming tRNAs and codons in the A site of the translating ribosome [4,5,6]. The detailed chemistry of the Elongator modification reaction is insufficiently described, and the role of the resulting modifications is not fully understood [11,12,13] It is currently unclear how tRNA is delivered to the active center and how the high modification turnover can be achieved in the context of the full complex. The cellular role of Elongator is of fundamental clinical relevance, as mutations affecting the integrity and activity of this macromolecular complex are related to the onset of neurodegenerative diseases [14,15,16], cancer [17,18], and intellectual disabilities [19]

Methods

Results

Conclusion