Monash Biomedicine Discovery Institute, Faculty of Medicine, Nursing and Health Sciences, Monash University, Clayton/Melbourne, VIC, 3800, Australia.
Department of Physics and Astronomy, Purdue University, West Lafayette, IN, 47907, USA.
Nat Commun. 2023 Aug 2;14(1):4652. doi: 10.1038/s41467-023-40428-1.
Endosomal maturation is critical for robust and timely cargo transport to specific cellular compartments. The most prominent model of early endosomal maturation involves a phosphoinositide-driven gain or loss of specific proteins on individual endosomes, emphasising an autonomous and stochastic description. However, limitations in fast, volumetric imaging long hindered direct whole cell-level measurements of absolute numbers of maturation events. Here, we use lattice light-sheet imaging and bespoke automated analysis to track individual very early (APPL1-positive) and early (EEA1-positive) endosomes over the entire population, demonstrating that direct inter-endosomal contact drives maturation between these populations. Using fluorescence lifetime, we show that this endosomal interaction is underpinned by asymmetric binding of EEA1 to very early and early endosomes through its N- and C-termini, respectively. In combination with agent-based simulation which supports a 'trigger-and-convert' model, our findings indicate that APPL1- to EEA1-positive maturation is driven not by autonomous events but by heterotypic EEA1-mediated interactions, providing a mechanism for temporal and population-level control of maturation.
内体成熟对于将货物有效地、及时地运输到特定的细胞隔室至关重要。早期内体成熟的最突出模型涉及单个内体上特定蛋白质的磷酸肌醇驱动的获得或丧失，强调了自主和随机的描述。然而，快速、大容量成像的局限性长期以来一直阻碍了对成熟事件绝对数量的直接全细胞水平测量。在这里，我们使用晶格光片成像和定制的自动分析来跟踪整个群体中的个体早期（APPL1 阳性）和早期（EEA1 阳性）内体，证明了这些群体之间的成熟是由直接的内体间接触驱动的。我们使用荧光寿命成像，表明这种内体相互作用是通过 EEA1 的 N 端和 C 端分别与早期和早期内体的不对称结合来支撑的。结合支持“触发-转化”模型的基于代理的模拟，我们的研究结果表明，APPL1 阳性到 EEA1 阳性的成熟不是由自主事件驱动的，而是由异质 EEA1 介导的相互作用驱动的，为成熟的时间和群体水平控制提供了一种机制。
