In agreement with this conclusion, budding yeast actin patches (which are known to be endocytic sites) are polarized to regions of cell surface growth. Mechanisms must, therefore, exist to target components involved in endocytic site initiation to specific regions of the plasma membrane. The plasma membrane is highly organized and the composition of distinct regions is controlled in part through endocytosis and exocytosis. The initiation of endocytic sites is closely linked to their placement on the plasma membrane. Further analysis of early-arriving proteins will be key in determining how assembly of endocytic sites is initiated. Intriguingly, Ede1p exhibits distinct dynamics to clathrin, suggesting that there are two different activities in endocytic site formation ( Kaksonen et al., 2005 Toshima et al., 2006). Ede1p is a homologue of the mammalian adapter Eps15, which localizes to endocytic sites but has as yet undefined dynamics ( Tebar et al., 1996 van Delft et al., 1997). An additional yeast protein, Ede1p, arrives early and plays a role in endocytic site formation, although its dynamics and function have yet to be fully investigated ( Kaksonen et al., 2005 Toshima et al., 2006). In yeast, the role of AP-2 is unclear, but clathrin, a coat module component, marks the first stages of endocytosis, and its deletion causes severe defects in the number of sites formed ( Huang et al., 1999 Kaksonen et al., 2005 Newpher et al., 2005 Newpher and Lemmon, 2006). Additionally, the adapter AP-2, is critical for site formation and has similar dynamic behavior to clathrin and so is thought to arrive early ( Hinrichsen et al., 2003 Motley et al., 2003 Ehrlich et al., 2004 Keyel et al., 2004). In mammalian cells the classical coat protein clathrin marks the earliest known stage of endocytic site formation ( Merrifield et al., 2002).
The best candidates for proteins that initiate endocytic site formation are those that arrive earliest. In Saccharomyces cerevisiae four endocytic modules have been defined that each contain proteins with similar dynamics: the coat, WASP/myo, amphiphysin, and actin modules ( Kaksonen et al., 2005).ĭespite detailed knowledge of events at endocytic sites, little is understood about the early stages of their formation. It is now evident that the dynamic recruitment and disappearance of endocytic proteins are precisely coordinated for productive internalization and that each protein has defined dynamics at endocytic sites. These studies have identified numerous proteins that sequentially assemble at endocytic sites and have shown that actin polymerization can power internalization. The dynamics of protein recruitment to sites of clathrin-mediated endocytosis have been revealed by live-cell microscopy in budding yeast and mammalian cells ( Merrifield et al., 2002 Kaksonen et al., 2003 Kaksonen et al., 2005). Thus Ede1p and Syp1p are conserved, early-arriving endocytic proteins with roles in the formation and placement of endocytic sites, respectively.
Additionally, Ede1p is important for endocytic site formation, whereas Syp1p acts as a polarized factor that recruits both Ede1p and endocytic sites to the necks of emerging buds. In yeast, both Syp1p and Ede1p play important roles in the rate of endocytic site turnover. Like Syp1p, SGIP1-α arrives early at sites of clathrin-mediated endocytosis, suggesting that Syp1p/Ede1p and SGIP1-α/Eps15 may have a conserved function. We find that Syp1p is related in amino acid sequence to several mammalian proteins one of which, SGIP1-α, is an endocytic component that binds the Ede1p homolog Eps15. Syp1p arrives at endocytic sites early in their formation and shares unique dynamics with the EH-domain protein Ede1p. Here we identify the septin-associated protein Syp1p as a component of the machinery that drives clathrin-mediated endocytosis in budding yeast. However, little is understood about the early stages of their formation.
Recent studies have revealed the detailed timing of protein recruitment to endocytic sites in budding yeast.
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