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Polarized transport is essential for constructing multiple plasma membrane domains in the cell. Drosophila photoreceptors are an excellent model system to study the mechanisms of polarized transport. Rab11 is the key factor regulating the post-Golgi transport of rhodopsin 1 (Rh1; also known as NinaE), a photoreceptive protein, to the rhabdomere, a photoreceptive plasma membrane. Here, we found that neuronal Synaptobrevin (nSyb) colocalizes with Rab11 on the trans-side of Golgi stacks and post-Golgi vesicles at the rhabdomere base, and nSyb deficiency impairs rhabdomeric transport and induces accumulation of Rh1 and vesicles in the cytoplasm; this is similar to the effects of Rab11 loss. These results indicate that nSyb acts as a post-Golgi SNARE toward rhabdomeres. Surprisingly, in Rab11-, Rip11- and nSyb-deficient photoreceptors, illumination enhances cytoplasmic accumulation of Rh1, which colocalizes with Rab11, Rabenosyn5, nSyb and Arrestin 1 (Arr1). Arr1 loss, but not Rab5 dominant negative (Rab5DN) protein expression, inhibits the light-enhanced cytoplasmic Rh1 accumulation. Rab5DN inhibits the generation of Rh1-containing multivesicular bodies rather than Rh1 internalization. Overall, these results indicate that exocytic Rh1 mingles with endocytosed Rh1 and is then transported together to rhabdomeres.

Golgi stacks are the basic structural units of the Golgi. Golgi stacks are separated from each other and scattered in the cytoplasm of

Golgi stacks are the basic structural units of the Golgi apparatus. Golgi stacks are separated from each other and scattered in the cytoplasm of Drosophila cells. Here, we report that the ARF-GEF inhibitor Brefeldin A (BFA) induces the formation of BFA bodies, which are aggregates of Golgi stacks, trans-Golgi networks, and recycling endosomes. Recycling endosomes are located in the centers of BFA bodies, while Golgi stacks surround them on their trans sides. Live imaging of S2 cells revealed that Golgi stacks repeatedly merged and separated on their trans sides, and BFA caused successive merger by inhibiting separation, forming BFA bodies. S2 cells carrying genome-edited BFA-resistant mutant Sec71M717L did not form BFA bodies at high concentrations of BFA; S2 cells carrying genome-edited BFA-hypersensitive mutant Sec71F713Y produced BFA bodies at low concentrations of BFA. These results indicate that Sec71 is the sole BFA target for BFA body formation and controls Golgi stack separation. Finally, we showed that impairment of Sec71 in fly photoreceptors induces BFA body formation with accumulation of both apical and basolateral cargos, resulting in inhibition of polarized transport.

Historically, the trans-Golgi network (TGN) has been recognized as a sorting center of newly synthesized proteins, whereas recycling endosome (RE) is a compartment where endocytosed materials transit before being recycled to the plasma membrane. However, recent findings revealed that both the TGN and RE connect endocytosis and exocytosis, and thus are functionally overlapping. Here we report, in both Drosophila and microtubule-disrupted HeLa cells, that REs are interconvertible between two distinct states, namely Golgi-associated REs and free REs. Detachment and reattachment of REs and Golgi stacks were often observed. These two types of REs were in the route of Glycosylphosphatidylinositol-anchored cargo protein released from the endoplasmic reticulum, but not in that of Vesicular stomatitis virus G protein. In plants, it has been established that there are two types of TGNs: the Golgi-associated TGN and Golgi-independent TGN. Dynamics of REs in both Drosophila and mammalian cells revealed strong similarity to plant TGNs. Together with the molecular-level similarity, these results indicate that fly/mammalian REs are equivalent organelles to TGNs in plants, and evoke reconsideration of identities and functional relationships between REs and TGNs.

Historically, the trans-Golgi network (TGN) has been recognized as a sorting center of newly synthesized proteins, whereas the recycling endosome (RE) is a compartment where endocytosed materials transit before being recycled to the plasma membrane. However, recent findings revealed that both the TGN and RE connect endocytosis and exocytosis and, thus, are functionally overlapping. Here we report, in both