During endocytosis, cargo or membrane receptors are internalised into early endosomes (EEs) before being delivered to late endosomes (LEs) on their path to lysosomal degradation. The predominant model of endocytic progression posits that transient EEs containing the GTPase Rab5 mature into LEs in a process that involves the exchange of Rab5 to Rab7a. Now, Oddmund Bakke and colleagues (Skjeldal et al., 2021) suggest that endosomal maturation also induces the de novo formation of Rab5-positive EEs. The authors take advantage of MDCK-Ii cells with enlarged endosomes, in which fluorescent Rab5 remains on the endosomal membrane for a longer time without affecting the transition from EEs to LEs, as previously reported in the journal (Margiotta et al., 2020). Live imaging of these and HeLa cells revealed that Rab5 detachment occurs in two phases. The initial phase is a fast diffusion-like exchange between the endosomal membrane and cytosol, and is independent of Rab7a. Surprisingly, during the latter phase, Rab5 microdomains converge into larger domains, and this is dependent on the recruitment of Rab7a to the maturing endosome. These converged domains then detach and give rise to newly formed and functional EEs. Together, this work thus suggests that Rab5 is not entirely exchanged via the cytosol during endosome maturation, but a fraction of Rab5 stays associated with the endosomal membrane and primes the formation of new EEs, thereby maintaining homeostasis of EE compartments.
