The ghost in the machine: small GTPases as spatial regulators of exocytosis

Abstract

Temporal and spatial regulation of membrane-trafficking events is crucial to both membrane identity and overall cell polarity. Small GTPases of the Rab, Ral and Rho protein families have been implicated as important regulators of vesicle docking and fusion events. This review focuses on how these GTPases interact with the exocyst complex, which is a multisubunit tethering complex involved in the regulation of cell-surface transport and cell polarity. The Rab and Ral GTPases are thought to function in exocyst assembly and vesicle-tethering processes, whereas the Rho family GTPases seem to function in the local activation of the exocyst complex to facilitate downstream vesicle-fusion events. The localized activation of the exocyst by Rho GTPases is likely to have an important role in spatial regulation of exocytosis.

Figure 1

Alignment of mammalian and yeast exocyst subunits that interact with Rho, Ral and Rab small GTPases. Regions of each subunit that are conserved between yeast and mammals are shown in yellow. Regions lacking obvious sequence similarity are blue in mammalian cells and red in yeast.

Figure 2

A three-step model for vesicle docking, exocyst activation and vesicle fusion regulated by small GTPases. (1) The initial vesicle-docking or tethering event is regulated by Rab and Ral GTPases, perhaps by promoting exocyst assembly. The association of particular exocyst subunits with the vesicle or plasma membrane in this diagram is speculative. There is evidence that exocyst assembly is regulated by Ral and this function, like that of Rab GTPases, is first required for vesicle tethering rather than fusion [67]. Fluorescence recovery after photobleaching (FRAP) studies in yeast have indicated that all of the exocyst subunits except Sec3 are likely to be delivered to sites of polarized growth through vesicle-mediated events [68]. (2) This is followed by local activation of the exocyst complex by Rho3, Cdc42 or TC10 family GTPases in their active GTP-bound state. Exocyst activation results in a stimulation of downstream fusion activity, probably by promoting assembly of active t-SNARE heterodimers. (3) The presence of active t-SNARE dimers results in SNARE-mediated fusion of the secretory vesicles at the site of exocyst activation.

Figure 3

A model for activation of the exocyst complex by Rho family GTPases. (a) Domain organization and molecular regulation of formins. In the absence of Rho GTP, formins are maintained in an inactive state by an autoinhibitory interaction between the DAD and DID domains, which is relieved by association of an active, GTP-bound Rho GTPases with the GBD domain. This interaction enables DID to adopt a structural conformation that induces release of the DAD domain and leads to the activation of the formin protein. (b) Domain organization and model for molecular regulation of Exo70 and the exocyst complex by Rho GTPase. The D domain of Exo70 interacts with phospholipids containing phosphatidylinositol (4,5)-bisphosphate (PIP₂) and the C domain is necessary for the interaction with Rho family small GTPases. In the absence of Rho GTP, Exo70, along with other components of the exocyst complex, remain in the inactive or basal activity state. Upon interaction of Exo70 with Rho GTP, Exo70 adopts an alternative conformation that leads to the activation of the exocyst complex. The activation in this case could be the result of disrupting an inhibitor interaction between Exo70 and another subunit of the exocyst complex or a direct change in the conformation of Exo70 itself, which then leads to a change in the overall structure of the complex.