Membrane trafficking: GTPases stack up

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The GTPases RAB6 and ARL1 form an unusual tetrameric complex that jointly localizes the GCC185 tether protein at the trans-Golgi network (TGN).

The small GTPases RAB and ARL regulate vesicular transport to and from the Golgi by coordinating the packaging of cargo proteins and their subsequent delivery to specific cellular subdomains. Once a vesicle reaches its destination, a tether protein brings the vesicle and target membrane into close proximity to permit fusion, which is catalysed by SNARE proteins. RAB and ARL were thought to have distinct roles in these processes, but Burguete et al. now report in Cell that an unusual tetrameric complex of RAB6 and ARL1 jointly localizes a tether protein at the trans-Golgi network (TGN).

The golgin-family member GCC185 is a tether protein that is required for the receipt of vesicles from late endosomes during retrograde transport. A coiled-coil domain and a GRIP domain in GCC185 are vital for its Golgi localization. The GRIP domain binds to ARL1, which in other golgins directs their localization to the Golgi. Interestingly, although GCC185 was localized to the Golgi and required ARL1 for proper localization, it displayed poor affinity for ARL1.

So how is GCC185 recruited to the Golgi? GCC185 is necessary for the transport of RAB9-bearing vesicles to the TGN, and the authors showed that GCC185 also binds to RAB6. RAB6 and RAB9 interact with distinct, yet overlapping, subdomains of GCC185; RAB6 binds predominantly to the coiled-coil domain and RAB9 requires the coiled-coil domain as well as portions of the GRIP domain. Determining the structure of RAB6 in complex with the isolated RAB-binding domain (RBD) of GCC185 revealed that two molecules of RAB6 interact with a dimer of the GCC185 RBD.

Because RAB6 and ARL1 are both necessary but not sufficient for GCC185 localization in cultured cells, could the two proteins cooperate to regulate GCC185 function? Indeed, Burguete et al. found that RAB6 robustly promotes ARL1 binding to GCC185 in vitro. A model of the hexameric structure indicates that binding of two RAB6 monomers to dimeric GCC185 may stabilize the GRIP domain to facilitate ARL1 binding. The RBD is N-terminal to the GRIP domain, suggesting a symmetrical complex in which RAB6 is stacked above ARL1; four GTPase molecules sandwich a GCC185 dimer. The different mechanisms by which ARL1 and RAB6 are anchored to cellular membranes may underpin this heterohexamer formation: the N terminus of ARL1 is myristoylated and embedded into cellular membranes, which keeps ARL1 close to the membrane surface. Conversely, RAB6 is linked to membranes by a long, flexible hypervariable domain, which allows it to stretch over ARL1 to contact GCC185, and reach as far as $sim$ 100 Å from the membrane surface.

Together, the data suggest a model in which GCC185 may be transferred from RAB9-bearing vesicles to the TGN, where RAB6 — on the TGN membrane — displaces RAB9 to bind GCC185. RAB6 binding in some way enables ARL1 binding and thus transfers GCC185 in a unidirectional manner from late endosomes to the TGN. GCC185 also mediates microtubule nucleation, an event that probably follows the interaction of GCC185 with RAB6 and ARL1.

Assistant Editor, UCSD–Nature Signaling Gateway

Emily J. Chenette

References

Burguete, A. S. et al. Rab and Arl GTPase family members cooperate in the localization of the Golgin GCC185. Cell 132, 286–298 (2008) | Article | PubMed |
Bonifacino, J. S. & Rojas, R. Retrograde transport from endosomes to the trans-Golgi network. Nature Rev. Mol. Cell Biol. 7, 568–579 (2006) | Article | PubMed |