Rab11 Regulates the Compartmentalization of Early Endosomes Required for Efficient Transport from Early Endosomes to the Trans-Golgi Network

TLDR

Rab GTPases regulate membrane traffic between organelles, and Rab11 has been localized to the pericentriolar recycling compartment, post‑Golgi vesicles, and the trans‑Golgi network. The study compared the effects of overexpressing wild‑type and mutant Rab11 on intracellular transport steps in the endocytic/degradative and biosynthetic/exocytic pathways in HeLa cells. Transport from endosomes to the Golgi was examined using Shiga toxin B subunit and TGN38 as reporter molecules. Overexpressing Rab11 (wild‑type, Q70L, or S25N) altered TfR, Tf, STxB, and TGN38 localization, with wt and Q70L directing them to sorting endosomes lacking cellubrevin and S25N redistributing them into a cellubrevin‑rich tubular network that is inaccessible to low‑temperature‑internalized Tf, and all mutants inhibited STxB/TGN38 transport to the TGN, indicating Rab11 regulates endosome‑to‑TGN trafficking by modulating membrane distribution in early endosomes.

Abstract

Several GTPases of the Rab family, known to be regulators of membrane traffic between organelles, have been described and localized to various intracellular compartments. Rab11 has previously been reported to be associated with the pericentriolar recycling compartment, post-Golgi vesicles, and the trans-Golgi network (TGN). We compared the effect of overexpression of wild-type and mutant forms of Rab11 on the different intracellular transport steps in the endocytic/degradative and the biosynthetic/exocytic pathways in HeLa cells. We also studied transport from endosomes to the Golgi apparatus using the Shiga toxin B subunit (STxB) and TGN38 as reporter molecules. Overexpression of both Rab11 wild-type (Rab11wt) and mutants altered the localization of the transferrrin receptor (TfR), internalized Tf, the STxB, and TGN38. In cells overexpressing Rab11wt and in a GTPase-deficient Rab11 mutant (Rab11Q70L), these proteins were found in vesicles showing characteristics of sorting endosomes lacking cellubrevin (Cb). In contrast, they were redistributed into an extended tubular network, together with Cb, in cells overexpressing a dominant negative mutant of Rab11 (Rab11S25N). This tubularized compartment was not accessible to Tf internalized at temperatures <20°C, suggesting that it is of recycling endosomal origin. Overexpression of Rab11wt, Rab11Q70L, and Rab11S25N also inhibited STxB and TGN38 transport from endosomes to the TGN. These results suggest that Rab11 influences endosome to TGN trafficking primarily by regulating membrane distribution inside the early endosomal pathway.