Communication network within the essential AAA-ATPase Rix7 drives ribosome assembly

Area of Science:

• Molecular Biology
• Structural Biology
• Biochemistry

Background:

• Rix7 is a vital AAA+ ATPase involved in early ribosome biogenesis.
• It functions as a protein translocase, removing substrates from preribosomes via its central pore.
• The coordination mechanism between Rix7's domains was previously unknown.

Purpose of the Study:

• To elucidate the structural basis of Rix7's function in ribosome biogenesis.
• To understand how Rix7's N-terminal domain (NTD) and AAA+ domains (D1, D2) coordinate within the hexamer.
• To identify key structural motifs essential for substrate translocation and interdomain communication.

Main Methods:

• Cryo-electron microscopy (cryo-EM) was used to capture structures of Rix7 variants.
• Cross-linking mass spectrometry (XL-MS) investigated the NTD's association with the hexamer.
• Cell-based assays were employed to assess the functional importance of specific Rix7 domains and motifs.

Main Results:

• High-resolution cryo-EM structures of Rix7 hexamers, including a 2.9 Å structure of the NTD-deleted variant, were obtained.
• The N-terminal domain (NTD) was found to associate with the central channel in vitro, though disordered in cryo-EM.
• The linker between D1 and D2 domains and pore-lining loops are crucial for large ribosomal subunit formation.

Conclusions:

• Rix7 employs a sophisticated communication network involving its domains to drive ribosome biogenesis.
• Structural insights reveal critical elements for substrate translocation and interdomain communication within the Rix7 hexamer.
• The linker and pore loops are essential for Rix7's role in large ribosomal subunit assembly.