TFG facilitates outer coat disassembly on COPII transport carriers to promote tethering and fusion with ER–Golgi intermediate compartments

Abstract

The conserved coat protein complex II (COPII) mediates the initial steps of secretory protein trafficking by assembling onto subdomains of the endoplasmic reticulum (ER) in two layers to generate cargo-laden transport carriers that ultimately fuse with an adjacent ER–Golgi intermediate compartment (ERGIC). Here, we demonstrate that Trk-fused gene (TFG) binds directly to the inner layer of the COPII coat. Specifically, the TFG C terminus interacts with Sec23 through a shared interface with the outer COPII coat and the cargo receptor Tango1/cTAGE5. Our findings indicate that TFG binding to Sec23 outcompetes these other associations in a concentration-dependent manner and ultimately promotes outer coat dissociation. Additionally, we demonstrate that TFG tethers vesicles harboring the inner COPII coat, which contributes to their clustering between the ER and ERGIC in cells. Together, our studies define a mechanism by which COPII transport carriers are retained locally at the ER/ERGIC interface after outer coat disassembly, which is a prerequisite for fusion with ERGIC membranes.

Membrane Transport at an Organelle Interface in the Early Secretory Pathway: Take Your Coat Off and Stay a While

“…Strikingly, depletion of TFG not only results in the stabilization of COPII‐coated transport carriers, but also their aberrant appearance away from the ER/ERGIC interface, suggesting an additional role for TFG in tethering COPII carriers prior to full uncoating and fusion with ERGIC membranes . Consistent with this idea, recombinant TFG is sufficient to cluster inner COPII‐coated liposomes in vitro . In particular, the carboxyl‐terminal intrinsically disordered domain of TFG is critical for this activity.…”

“…In particular, the carboxyl‐terminal intrinsically disordered domain of TFG is critical for this activity. Previous work suggested that this region of TFG is capable of self‐assembly into larger structures (≈200–300 nm in size), raising the possibility that elevated concentrations of TFG can lead to phase separation within the cytoplasm, potentially forming a membrane‐less liquid droplet . By generating a phase separated liquid droplet at the interface between ER and ERGIC membranes, COPII transport carriers may become transiently trapped, providing sufficient time for the inner coat to disassociate (Figure ).…”

“…However, the cost of augmenting this system has been far from negligible, requiring a unique step of transport that allows for plasticity in carrier size, facilitates rapid carrier uncoating, and restricts diffusion away from ERGIC (ER‐Golgi intermediate compartment) membranes, all without assistance from canonical motor proteins and cytoskeletal filaments. To achieve this, metazoans appear to have separately evolved two distinct families of proteins, Trk‐fused gene (TFG) and Tango1/cTage5, which are both used to enable efficient transport of small and large cargoes between the ER and ERGIC in mammals …”

“…Fairly elaborate mechanisms have been proposed to describe how Tango1/cTage5 complexes may facilitate this action, based on the premise that the inner COPII coat exists in two distinct complexes after assembly on ER membranes, bound either to Tango1/cTage5 or Sec13‐Sec31 . However, direct evidence for this is lacking, and localization studies consistently show the presence of both Tango1/cTage5 and Sec13‐Sec31 at all ER subdomains harboring Sec23‐Sec24 . Instead, a simpler explanation may reconcile the role of Tango1/cTage5 complexes in early secretory pathway function.…”

“…However, recent studies suggest a key role for TFG in this process (Figure ). Specifically, TFG was shown to bind directly to Sec23 through a shared interface with Sec31, promoting disassembly of the outer coat during the trafficking of COPII carriers within the ER/ERGIC interface . Additionally, post‐translation modifications on COPII coat proteins may further facilitate their disassembly, although neither biochemical or structural data are currently available to support this idea …”