Structural biology of macromolecular complexes involved in RNA metabolism
Translation is the central process in biology during which genetic information encoded on mRNAs is read by the ribosome and polypeptides are synthesized. Extensive biochemical studies on prokaryotic ribosomes have given insights into the assembly of this machine, while structural studies have illuminated its workings during translation.
Eukaryotic ribosome biogenesis is vastly more complex than in prokaryotes, requiring more than 200 additional factors and proceeding through multiple cellular compartments. The process centers around the transcription of a long ribosomal RNA transcript in the nucleolus. Chemical modification of bases and the association of small subunit ribosomal proteins occur at this early stage. A series of RNA cleavage events then separates the pre-40S and pre-60S particles, which exit the nucleus as almost fully assembled pre-ribosomal particles. These particles associate with late-acting biogenesis factors that also act as transport adaptors, mediating interactions with the nucleocytoplasmic transport machinery. Final maturation is completed in the cytoplasm where late-acting biogenesis factors are removed and the mature ribosomal particles can associate on mRNAs. Proteomic studies have generated parts lists of proteins involved in ribosome biogenesis in yeast and many of these proteins are conserved in mammals. Less clear is the order of individual events, particularly in late cytoplasmic maturation of ribosomal particles. I am interested in using structural and biochemical approaches to gain mechanistic insights into the function of ribosome biogenesis factors in yeast and mammalian cells.
Recently, we solved the structure of yTsr1, a conserved ribosome biogenesis factor that associates with pre-40S particles in the nucleus and is removed from particles in the cytoplasm during maturation. Previously, little was known about Tsr1 except that it is essential for generating mature 40S subunits and that it has sequence similarity to structurally related to translational GTPases such as EF-Tu and SelB but has lost catalytic activity and is locked in a conformation resembling the GTP-bound form of small GTPases (Figure. 1A). By docking the structure of Tsr1 into previously published cryo-EM maps of pre-40S particles, we find that Tsr1 occupies a critical part of the surface of the particle and that it likely blocks access to other biogenesis factors such as eIF5B and Rio1 (Figure. 1B). This suggests that Tsr1 exit is a critical step that occurs prior to the action of Rio1 and eIF5B, giving an insight into the temporal order of events that occur on these particles (Figure. 1C).
McCaughan, U.M., Jayachandran, U., Shchepachev, V., Chen, Z.A., Rappsilber, J., Tollervey, D., and Cook, A.G. (2016). Pre-40S ribosome biogenesis factor Tsr1 is an inactive structural mimic of translational GTPases. Nat Commun 7, 11789.
Jayachandran, U., Grey, H., and Cook, A.G. (2016). Nuclear factor 90 uses an ADAR2-like binding mode to recognize specific bases in dsRNA. Nucleic Acids Res 44, 1924-1936.
Wandrey, F., Montellese, C., Koos, K., Badertscher, L., Bammert, L., Cook, A.G., Zemp, I., Horvath, P., and Kutay, U. (2015). The NF45/NF90 Heterodimer Contributes to the Biogenesis of 60S Ribosomal Subunits and Influences Nucleolar Morphology. Mol Cell Biol 35, 3491-3503.