Golgi Tubule Traffic and the Effects of Brefeldin A Visualized in Living Cells

TLDR

The Golgi complex is a dynamic organelle engaged in both secretory and retrograde membrane traffic. Here, we use green fluorescent protein–Golgi protein chimeras to study Golgi morphology in vivo. Analysis of the kinetics of tubule formation suggests a wetting‑like, tension‑driven membrane flow that supplements diffusive transfer of Golgi membrane into the ER. In living cells, Golgi membrane tubules are ubiquitous and interconnect Golgi elements, and upon Brefeldin A treatment they form an extensive network that persists for minutes before rapidly transferring both lipid and protein contents into the ER, demonstrating a nonselective, flow‑assisted transport that implies Golgi membranes are at higher free energy than ER and that retrograde tubule dynamics are essential for organelle maintenance.

Abstract

The Golgi complex is a dynamic organelle engaged in both secretory and retrograde membrane traffic. Here, we use green fluorescent protein–Golgi protein chimeras to study Golgi morphology in vivo. In untreated cells, membrane tubules were a ubiquitous, prominent feature of the Golgi complex, serving both to interconnect adjacent Golgi elements and to carry membrane outward along microtubules after detaching from stable Golgi structures. Brefeldin A treatment, which reversibly disassembles the Golgi complex, accentuated tubule formation without tubule detachment. A tubule network extending throughout the cytoplasm was quickly generated and persisted for 5–10 min until rapidly emptying Golgi contents into the ER within 15–30 s. Both lipid and protein emptied from the Golgi at similar rapid rates, leaving no Golgi structure behind, indicating that Golgi membranes do not simply mix but are absorbed into the ER in BFA-treated cells. The directionality of redistribution implied Golgi membranes are at a higher free energy state than ER membranes. Analysis of its kinetics suggested a mechanism that is analogous to wetting or adsorptive phenomena in which a tension-driven membrane flow supplements diffusive transfer of Golgi membrane into the ER. Such nonselective, flow-assisted transport of Golgi membranes into ER suggests that mechanisms that regulate retrograde tubule formation and detachment from the Golgi complex are integral to the existence and maintenance of this organelle.