The polypeptide exit tunnel of the ribosomal large subunit requires assembly factors for proper construction and function

Abstract

The final product of ribosome assembly consists of two ribosomal subunits capable of functioning with high fidelity. In eukaryotes, this process is facilitated by protein assembly factors that can promote proper formation of functional centers through structural probing, functional test‐driving, acting as a scaffold, or protecting the site from premature activity. The polypeptide exit tunnel (PET) of the ribosomal large subunit (LSU) is the conduit through which nascent polypeptide chains are passed as they are synthesized. Protein and RNA components of the PET lumen actively participate in translation by interacting with the nascent chain in order to regulate ribosome stalling events, thus regulating translation speed and gene expression. Recent cryo‐EM structures of immature LSU particles from different stages of assembly spanning the nucleoplasm to the cytoplasm have shown that the PET is sequentially probed by the C‐termini of three assembly factors: first, Nog1, then Rei1, and finally Reh1. The C‐termini of these three assembly factors mimic nascent chains, plugging the PET for almost the entirety of LSU assembly. No biological function, however, has been assigned to this redundant occupation of the PET. Here, we show that truncating PET‐inclusive residues from the C‐terminus of Nog1 causes LSU assembly defects that are exacerbated by analogous truncations to one or both of the Rei1 and Reh1 C‐termini. These defects manifest in processing of late immature ribosomal RNA intermediates. Interestingly, these assembly defects are further exacerbated at lower temperatures. Despite these mutants displaying slow growth at permissive temperatures, however, mature ribosomes are produced that may be defective in translation. Our data supports the model that Nog1 is necessary for efficient construction of the PET during early stages of assembly while Rei1 and Reh1 carry out checkpoints during later stages.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.