The Rab5 effector EEA1 is a core component of endosome docking

Abstract

Intracellular membrane docking and fusion requires the interplay between soluble factors and SNAREs. The SNARE hypothesis postulates that pairing between a vesicular v-SNARE and a target membrane z-SNARE is the primary molecular interaction underlying the specificity of vesicle targeting as well as lipid bilayer fusion. This proposal is supported by recent studies using a minimal artificial system. However, several observations demonstrate that SNAREs function at multiple transport steps and can pair promiscuously, questioning the role of SNAREs in conveying vesicle targeting. Moreover, other proteins have been shown to be important in membrane docking or tethering. Therefore, if the minimal machinery is defined as the set of proteins sufficient to reproduce in vitro the fidelity of vesicle targeting, docking and fusion as in vivo, then SNAREs are not sufficient to specify vesicle targeting. Endosome fusion also requires cytosolic factors and is regulated by the small GTPase Rab5. Here we show that Rab5-interacting soluble proteins can completely substitute for cytosol in an in vivo endosome-fusion assay, and that the Rab5 effector EEA1 is the only factor necessary to confer minimal fusion activity. Rab5 and other associated proteins seem to act upstream of EEA1, implying that Rab5 effectors comprise both regulatory molecules and mechanical components of the membrane transport machinery. We further show that EEA1 mediates endosome docking and, together with SNAREs, leads to membrane fusion.

Distinct molecular mechanisms and divergent endocytotic pathways of AMPA receptor internalization

“…The simplest explanation for this difference in kinetics is that AMPA receptors that are internalized in response to insulin return more slowly to the surface. This hypothesis is consistent with our finding that AMPA receptors internalized in response to AMPA, but not in response to insulin, remain largely within a recycling endosome system, as defined by colocalization with EEA1, syntaxin 13 and TfR [32][33][34][35][36][37][38][39][40] . Unlike AMPA, insulin caused a divergence of internalized AMPA receptors out of the pathway delineated by EEA1, syntaxin 13 and TfR into a separate intracellular compartment.…”

Birbeck Granules Are Subdomains of Endosomal Recycling Compartment in Human Epidermal Langerhans Cells, Which Form Where Langerin Accumulates

“…In another study carried out in the same cell type, we demonstrated that intracellular CD1a also colocalizes with Rab11 (Salamero et al, 2001), a small GTPase that specifically associates with the membrane of the endosomal recycling compartment (ERC) (Ullrich et al, 1996;Ren et al, 1998). Steady-state internal Langerin colocalized strongly with Rab11 in the ERC ( Figure 1E) but poorly with EEA1, a marker of the early/sorting endosomes ( Figure 1F) (Simonsen et al, 1998;Christoforidis et al, 1999;McBride et al, 1999 …”

“…In another study carried out in the same cell type, we demonstrated that intracellular CD1a also colocalizes with Rab11 (Salamero et al, 2001), a small GTPase that specifically associates with the membrane of the endosomal recycling compartment (ERC) (Ullrich et al, 1996;Ren et al, 1998). Steady-state internal Langerin colocalized strongly with Rab11 in the ERC ( Figure 1E) but poorly with EEA1, a marker of the early/sorting endosomes ( Figure 1F) (Simonsen et al, 1998;Christoforidis et al, 1999;McBride et al, 1999). The steady-state localization of Lange- rin was clearly different from that of CD1a, the latter being more equally distributed between the cell surface and internal membranes; however, this could be related to the dual role of the ERC as a sorting compartment for internalized proteins (Ghosh et al, 1998;Mallet and Maxfield, 1999;Wilcke et al, 2000) and as a storage reservoir for the regulated delivery of proteins to the plasma membrane (Johnson et al, 2001).…”

Arf6-independent GPI-anchored Protein-enriched Early Endosomal Compartments Fuse with Sorting Endosomes via a Rab5/Phosphatidylinositol-3′-Kinase–dependent Machinery

“…The Rab5 GTPase forms a domain by recruiting hVps34, causing localized production of PI3P (Christoforidis et al, 1999a). EEA1 stabilization on the GEECs also requires active Rab5 (Murray et al, 2002).…”