Structural view on 60S ribosome biogenesis

Barrio Garcia, Clara (2016): Structural view on 60S ribosome biogenesis: remodeling and quality control mechanisms. Dissertation, LMU München: Fakultät für Chemie und Pharmazie

Abstract

Eukaryotic ribosomes undergo a complex maturation process through which the ribosomal RNA (rRNA) must bind to ribosomal proteins (r-proteins) and fold into its native state. This requires numerous auxiliary factors responsible of rRNA processing, remodeling, intracellular transport and quality control checkpoints. Impairments in the process of ribosome production can lead to different diseases known as ribosomopathies. However, little is known about which defects in the ribosome biogenesis pathway can escape all quality checkpoints and what targets these faulty ribosomes and their putative products for degradation. The first part of this thesis constitutes a first attempt to address these questions, focusing on a specific impairment of the 60S biogenesis pathway that leads to the production of structurally deficient 80S ribosomes. Here, using cryo-EM and biochemical analysis, it is shown that large structural defects in the ribosome may bypass all quality control mechanisms in the nucleus, but are recognized in the cytoplasm after faulty ribosomes engage in translation. Then, the resulting proteins are targeted for degradation by the ribosome quality-control complex (RQC) after subunit splitting is carried out. The second part of this thesis focuses on a nucleoplasmic step of 60S ribosome biogenesis in which both the Rix1 complex and the dynein related AAA+ ATPase Rea1 bind to maturing pre-60S particles. Rea1 is required for the ATP-dependent dissociation of the assembly factor Rsa4 and may be involved in rearranging the Central Protuberance (CP). Several cryo-electron microscopy (cryo-EM) structures of native pre-60S particles bound to the Rix1-Rea1 machinery are presented in this thesis. Overall, the Rix1-Rea1 particle is similar to the earlier intermediate, the socalled Arx1 particle. However, a 180° rotation of the CP required to reach the final conformation of the 60S, has already happened in the Rix1-Rea1 state, constituting the largest remodeling step at the end of the maturation pathway. By performing cryo-EM analysis on two similar particles that were mutated on either Rix1 or Rea1 and by comparing the structures of the Arx1 and Rix1-Rea1 particles, a mechanistic model for the transition between the two intermediates is provided.

• Dokumententyp: Dissertationen (Dissertation, LMU München)
• Keywords: ribosome, biogenesis, 60S, cryo-EM, Rix1, Rea1
• Themengebiete: 500 Naturwissenschaften und Mathematik ; 500 Naturwissenschaften und Mathematik > 540 Chemie
• Fakultäten: Fakultät für Chemie und Pharmazie
• Sprache der Hochschulschrift: Englisch
• Datum der mündlichen Prüfung: 19. September 2016
• 1. Bericht­erstatter:in: Beckmann, Roland