Nanoscale imaging reveals the mechanisms of ER-to-Golgi transport via a dynamic tubular-vesicular network

Wong-Dilworth, L.; Bregu, G.; Restel, S.; Rodilla-Ramirez, C.; Ebeling, S.; Harel, S.; Leupold, P.; Grimm, J.; Lavis, L.; Bottanelli, F.; Angulo-Capel, J.; Campelo, F.

doi:10.1101/2023.10.27.563951

bioRxiv · 10.1101/2023.10.27.563951

Nanoscale imaging reveals the mechanisms of ER-to-Golgi transport via a dynamic tubular-vesicular network

Wong-Dilworth, L.·Bregu, G.·Restel, S.·Rodilla-Ramirez, C.·Ebeling, S.·Harel, S.·Leupold, P.·Grimm, J.·Lavis, L.·Bottanelli, F.·Angulo-Capel, J.·Campelo, F.

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Abstract

The endoplasmic reticulum (ER) and the Golgi apparatus are the first sorting stations along the secretory pathway of mammalian cells and have a crucial role in protein quality control and cellular homeostasis. While machinery components mediating ER-to-Golgi transport have been mapped, it is unclear how exchange between the two closely juxtaposed organelles is coordinated in living cells. Here, using gene editing to tag machinery components, live-cell confocal and stimulated emission depletion (STED) super-resolution microscopy, we show that ER-to-Golgi transport occurs via a dynamic network of tubules positive for the small GTPase ARF4. swCOPI machinery is tightly associated to this network and moves with tubular-vesicular structures. Strikingly, the ARF4 network appears to be continuous with the ER and ARF4 tubules remodel around static ER exit sites (ERES) defined by COPII machinery. We were further able to dissect the steps of ER-to-Golgi transport with functional trafficking assays. A wave of cargo released from the ER percolates through peripheral and Golgi-tethered ARF4 structures before filling the cis-Golgi. Perturbation via acute degradation of ARF4 shows an active regulatory role for the GTPase and COPI in anterograde transport. Our data supports a model in which anterograde ER-to-Golgi transport occurs via an ARF4 tubular-vesicular network directly connecting the ER and Golgi-associated pre-cisternae.

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Wong-Dilworth, L., Bregu, G., Restel, S., Rodilla-Ramirez, C., Ebeling, S., Harel, S., Leupold, P., Grimm, J., Lavis, L., Bottanelli, F., Angulo-Capel, J., Campelo, F.. 2023-10-27. Nanoscale imaging reveals the mechanisms of ER-to-Golgi transport via a dynamic tubular-vesicular network. https://doi.org/10.1101/2023.10.27.563951

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