Regulatory sites in the Mon1-Ccz1 complex control Rab5 to Rab7 transition and endosome maturation

Abstract

Maturation from early to late endosomes depends on the exchange of their marker proteins Rab5 to Rab7. This requires Rab7 activation by its specific guanine nucleotide exchange factor (GEF) Mon1-Ccz1. Efficient GEF activity of this complex on membranes depends on Rab5, thus driving Rab-exchange on endosomes. However, molecular details on the role of Rab5 in Mon1-Ccz1 activation are unclear. Here we identify key features in Mon1 involved in GEF regulation. We show that the intrinsically disordered N-terminal domain of Mon1 autoinhibits Rab5-dependent GEF-activity on membranes. Consequently, Mon1 truncations result in higher GEF activityin vitro, and a shift from Rab5 to more Rab7 positive structures inDrosophilanephrocytes and yeast, suggesting faster endosomal maturation. Using modeling, we further identify a conserved Rab5 binding site in Mon1. Mutations impairing Rab5 interaction result in poor GEF activity on membranes and growth defectsin vivo. Our analysis provides a framework to understand the mechanism of Rab-conversion and organelle maturation along the endomembrane system.SummaryTransport of proteins via the endolysosomal pathway requires Rab5 on early endosomes, which is replaced by Rab7 on late endosomes. Here, we identify distinct regulatory sites in the Rab7 activator, the Mon1-Ccz1 complex, shedding light on the regulation of Rab5 to Rab7 transition.

Structure of the metazoan Rab7 GEF complex Mon1–Ccz1–Bulli

“…Furthermore, MCBulli and CPLANE may contain a conserved binding site for a GTPase on Mon1 and Fuzzy, respectively. Functional studies on the regulation of the yeast MC1 and Drosophila MCBulli complexes provide experimental support for the proposed Rab5–MCBulli binding site ( 33 ). However, in contrast to Rab5, Rsg1 does not contain a lipidation motif and is unlikely a recruiter of CPLANE to membranes.…”

Spatiotemporal recruitment of the ubiquitin-specific protease USP8 directs endosome maturation

“…The Mon1–Ccz1 complex is a heterodimer in yeast, and a heterotrimer in metazoan cells (Dehnen, Janz et al, 2020, Vaites, Paulo et al, 2018, van den Boomen, Sienkiewicz et al, 2020, Wang, Stromhaug et al, 2002). The activation of Mon1-Ccz1 is depended on Rab5 interaction (Borchers, Janz et al, 2023, Langemeyer, Borchers et al, 2020) and Mon1 also coordinates endosome maturation together with the Rab5 GAP TBC1D18 (Hiragi, Matsui et al, 2022). Additionally, the Mon1-Ccz1 complex is able to interact with Rabex5 (Poteryaev, Datta et al, 2010), causing dissociation of Rabex5 from the membrane, which probably terminates the positive feedback loop of Rab5 activation and then promotes the recruitment and activation of Rab7 on endosomes (Nordmann, Cabrera et al, 2010, Poteryaev et al, 2010).…”

“…The activation of Mon1-Ccz1 is depended on Rab5 interaction (Borchers, Janz et al, 2023, Langemeyer, Borchers et al, 2020 and Mon1 also coordinates endosome maturation together with the Rab5 GAP TBC1D18 (Hiragi, Matsui et al, 2022).…”

Coordination between ESCRT function and Rab conversion during endosome maturation

“…Rab conversion is thought to be initiated by coincident detection of the Rab5GEF Rabex5 (RABX-5 in Caenorhabditis elegans ) and increasing levels of PI3P on endosomes by the Rab7GEF Mon1/Ccz1 (Mon1 is called SAND-1 or CMON-1 in C. elegans ) (Cabrera et al , 2014; Langemeyer et al , 2020; Poteryaev et al ., 2010; Poteryaev et al , 2007). In addition, Rab5 can directly bind to Mon1/Ccz1 and modulate its activity (Borchers et al , 2023; Herrmann et al , 2023; Kinchen & Ravichandran, 2010; Langemeyer et al ., 2020). These processes lead to the replacement of Rab5 by Rab7 on maturing endosomes (Borchers et al , 2021; Podinovskaia & Spang, 2018).…”