The roles of GTPases in biogenesis of the large ribosomal subunit

von Chairini Cássia Thomé
Dissertation
Datum der mündl. Prüfung:2025-02-18
Erschienen:2025-09-22
Betreuer:Dr. Katherine E Bohnsack
Gutachter:Dr. Katherine E Bohnsack
Gutachter:Prof. Dr. Ralf Ficner
Gutachter:Prof. Dr. Jörg Stülke
Gutachter:Dr. Ricarda Richter-Dennerlein
Gutachter:Prof. Dr. Matthias Dobbelstein
Gutachter:Prof. Dr. Henning Urlaub
Zum Verlinken/Zitieren: http://dx.doi.org/10.53846/goediss-11520

 

 

Dateien

Name:Thome_PhD_Thesis_Final_digital.pdf
Size:44.0Mb
Format:PDF

Diese Datei ist bis 17.02.2026 gesperrt.

Zusammenfassung

Englisch

Ribosomes are ribonucleoprotein complexes responsible for protein synthesis, and their biogenesis is a complex, highly orchestrated process requiring a significant amount of energy and the coordinated action of all three RNA polymerases. Disruptions in ribosomal subunit assembly can lead to dysfunctional ribosomes, causing severe congenital diseases (ribosomopathies), and may also contribute to tumorigenesis. Formation of the ribosomal subunits is coordinated by trans-acting ribosome biogenesis factors (RBFs) that mediate ribosomal RNA (rRNA) modification, folding and processing, as well as the incorporation and positioning of ribosomal proteins (RPs). GTPases are among the RBFs that consume energy to drive structural rearrangements of pre-ribosomal particles, and several of these enzymes play important roles in assembly of the large subunit. Much knowledge on ribosome biogenesis comes from the yeast Saccharomyces cerevisiae, and despite the conservation of key aspects of the process across eukaryotes, many human RBFs, including GTPases, remain to be fully characterised. Recent structural studies of human pre-60S particles have identified GTPBP4, GNL3, and GNL2 on the particles, suggest roles for these GTPases in 60S assembly, but their precise functions have remained elusive. Furthermore, another putative GTPase, GNL3L is enriched in nucleoli, a major site of ribosome assembly, but potential contributions of this enzyme to subunit assembly have remained unclear. This study focused on analysing the roles of GTPBP4, GNL2, GNL3 and GNL3L during large ribosomal subunit maturation. Proteomics analyses provide insights into the timing of recruitment of these proteins to pre-ribosomal particles and their subsequent release, as well as the influence of nucleotides on their pre-ribosome association. Analyses of pre-rRNA processing in cells expressing GTP binding and hydrolysis mutants allowed to differentiate between the requirement for the presence of these proteins and the importance of their nucleotide binding/catalytic activities. Protein-RNA crosslinking approaches were used to identify GNL3L binding sites on pre-ribosomes and map RNA contact sites on the GTPase. Expression of truncated versions of GNL3 and GNL3L allowed the contribution of individual domains to pre-ribosome association to be determined. Also, the importance of the GTPase activity of GNL3L for ribosome function was explored. Collectively, these data provide new insights into the functions of GTPBP4, GNL3, and GNL2 during assembly of the large ribosomal subunit in humans and support the involvement of an additional GTPase, GNL3L. These findings contribute to further understand the biogenesis of human cytosolic ribosomes and the roles of GTPases in this process.
Keywords: Ribosome biogenesis; GTPase; GTPBP4; GNL2; GNL3; GNL3L; ribosomal protein; pre-rRNA processing; ribosome biogenesis factors; translation; ribosomal RNA