Poster number 6 submitted by Fiona Fitzgerald
The DEAD-box helicase Drs1 plays a role in compaction of 25S rRNA domain III during nucleolar stages of large subunit assembly
Fiona Fitzgerald (Department of Biological Sciences, Carnegie Mellon University), Collin Bachert (Department of Biological Sciences, Carnegie Mellon University), Stephanie Biedka (Department of Biological Sciences, Carnegie Mellon University), Yungyang Zhang (State Key Laboratory of Membrane Biology, Peking University), Ning Gao (State Key Laboratory of Membrane Biology, Peking University), John L. Woolford (Department of Biological Sciences, Carnegie Mellon University)
The nucleolar stages of large subunit (LSU) assembly are characterized by compaction of individual domains of 25S ribosomal RNA (rRNA) around the root helices, beginning with compaction of domains I, II, and VI, followed by domain III, and ultimately domains IV and V. The compaction of these rRNA domains is important for formation of ribosome functional centers, e.g., the nascent polypeptide exit tunnel (NPET) and the peptidyl-transferase center (PTC). We also believe that before these rRNA domains are compacted, they function to form a network of interactions between pre-ribosomes to create the nucleolar condensate.
Some DEAD-box helicases (DBHs) involved in ribosome biogenesis use their ATPase activity to coordinate the folding of rRNA through removal of structural snoRNAs or assembly factors (AFs), and rearrangement of rRNA structure. However, the precise functions of most DBHs involved in ribosome assembly remain unclear.
The DBH Drs1 functions in early stages of LSU assembly. Depletion of Drs1 leads to accumulation of 27SA2 and 27SA3 pre-rRNAs. In contrast, the drs1-1 mutation near a conserved ATPase motif results in cold-sensitive growth and accumulation of 27SB pre-rRNA. We seek to understand the role of the ATPase activity of Drs1 in late-nucleolar stages of LSU assembly.
Previous studies using protein-protein crosslinking and co-transcriptional rRNA association indicate that Drs1 might bind in or near domain III of assembling pre-ribosomes. Further examination of the drs1-1 missense mutant, using a combination of proteomics, cryo-EM, and genetics, indicates that Drs1 may play an important role in compaction of rRNA domain III.
Keywords: rRNA, ribosome, helicase