The Ribosome Maturation Factor Rea1 utilizes nucleotide independent and ATP-hydrolysis driven Linker remodelling for the removal of ribosome assembly factors

Abstract

The essential ribosome maturation factor Rea1 (also known as Midasin) catalyses the removal of assembly factors from precursors of the large ribosomal subunit and subsequently promotes their export from the nucleus to the cytosol. Rea1 is a large protein of nearly 5000 residues and a member of the AAA+ (ATPases associated with various cellular activities) protein family. It consists of a concatenated ring of six AAA+ domains from which the ≈ 1700 residue linker emerges that is subdivided into stem, middle and top domains. A flexible and unstructured D/E rich region connects the linker top to a MIDAS (metal ion dependent adhesion site) domain, which is able to bind the assembly factor substrates. Despite its key importance for ribosome maturation, the Rea1 mechanism driving assembly factor removal is still poorly understood. Although early studies on Rea1 demonstrated that assembly factor removal requires ATP hydrolysis, the Rea1 conformations associated with ATP hydrolysis have not been identified yet. Here we demonstrate that the Rea1 linker is essential for assembly factor removal. It rotates and swings towards the AAA+ ring following a complex remodelling scheme involving nucleotide independent as well as nucleotide dependent steps. ATP hydrolysis is required to engage the linker top with the AAA+ ring and ultimately with the AAA+ docked MIDAS domain. The interaction between the linker top and the MIDAS domain allows force transmission for assembly factor removal.