ABSTRACT
Endosomal trafficking in single cells entails the generation of membrane vesicles; their motor protein-mediated transport; morphological alterations such as tubulation, fusion and fission; as well as dynamic maintenance of various identities, defined by the lipid composition and localisation of specific proteins on their membranes. Endosomal maturation is a major feature of this process. It is classically described as endosomes shedding one specific protein and acquiring another, resulting in an identity change. This step governs the flux and the number of distinct endosomal populations. Many of the molecular players that characterise different compartments have been identified—for example, APPL1 on very early endosomes, and EEA1 on early endosomes. However, how an ensemble population of endosomes matures at a whole cell level has remained unstudied. Here, we describe a novel inter-endosomal interaction-based feed-forward mechanism that drives endosomal conversions and controls ensemble conversion rates at the whole cell level. Furthermore, using live-cell Förster Resonance Energy Transfer, we demonstrate that this process is underpinned by co-ordinated precedence in EEA1 binding via its N- and C-terminal domains, ensuring the biochemical maturation of these vesicles. Using simulations, we provide a quantitative framework to recapture the experimentally observed characteristics in the reaction scheme and the activity of EEA1. We demonstrate that the model of APPL1 to EEA1 endosomal maturation needs to be extended to include heterotypic interactions between endosomes that result in conversions, which form a significant fraction of events, in addition to the previously reported single endosome-centric maturation models. Based on these results, we motivate how temporal specificity emerges in an inherently stochastic system—namely, through a timekeeping mechanism via an N-terminal EEA1 trigger that enables deterministic outcomes in ensemble endosomal conversions.
Competing Interest Statement
The authors have declared no competing interest.
