Figure 13–13 The role of dynamin in pinching off clathrin-coated vesicles. (a) Multiple dynamin molecules assemble into a spiral around the neck of the forming bud. The dynamin spiral is thought to recruit other proteins to the bud neck, which, together with dynamin, destabilize the interacting lipid bilayers so that the noncytoplasmic leaflets flow together. The newly formed vesicle then pinches off from the membrane. specific mutations in dynamin can either enhance or block the pinching-off process. (b) dynamin was discovered as the protein defective in the shibire mutant of Drosophila. These mutant flies become paralyzed because clathrin-mediated endocytosis stops, and the synaptic vesicle membrane fails to recycle, blocking neurotransmitter release. deeply invaginated clathrin-coated pits form in the nerve endings of the fly’s nerve cells, with a belt of mutant dynamin assembled around the neck, as shown in this thin-section electron micrograph. The pinching-off process fails because the required membrane fusion does not take place. (c, d) a model of how conformational changes in the gTpase domains of membrane-assembled dynamin may power a conformational change that constricts the neck of the bud. a single dynamin molecule is shown in orange in d. (b, from J.h. koenig and k. Ikeda, J. neurosci. 9:3844–3860, 1989. with permission from the society of neuroscience; c and d, adapted from M.g.J. ford, s. Jenni and J. nunnari, nature 477:561–566, 2011. with permission from Macmillan publishers.)
To balance the vesicle traffic to and from a compartment, coat proteins must assemble only when and where they are needed. While local production of PIPs plays a major part in regulating the assembly of clathrin coats on the plasma membrane and Golgi apparatus, cells superimpose additional ways of regulating coat formation. Coat-recruitment GTPases, for example, control the assembly of clathrin coats on endosomes and the COPI and COPII coats on Golgi and ER membranes.
Many steps in vesicle transport depend on a variety of GTP-binding proteins that control both the spatial and temporal aspects of vesicle formation and fusion. As discussed in Chapter 3, GTP-binding proteins regulate most processes in eukaryotic cells. They act as molecular switches, which flip between an active state with GTP bound and an inactive state with GDP bound. Two classes of proteins regulate the flipping: guanine nucleotide exchange factors (GEFs) activate the proteins by catalyzing the exchange of GDP for GTP, and GTPase-activating proteins (GAPs) inactivate the proteins by triggering the hydrolysis of the bound GTP to GDP (see Figures 3–68 and 15–7). Although both monomeric GTP-binding proteins (monomeric GTPases) and trimeric GTP-binding proteins (G proteins) have important roles in vesicle transport, the roles of the monomeric GTPases are better understood, and we focus on them here.