The GTPase-activating protein GRAF1 regulates the CLIC/GEEC endocytic pathway

Medical Research Council, Laboratory of Molecular Biology, Hills Road, Cambridge, CB2 0QH, UK.
Current Biology (Impact Factor: 9.57). 12/2008; 18(22):1802-8. DOI: 10.1016/j.cub.2008.10.044
Source: PubMed

ABSTRACT Clathrin-independent endocytosis is an umbrella term for a variety of endocytic pathways that internalize numerous cargoes independently of the canonical coat protein Clathrin [1, 2]. Electron-microscopy studies have defined the pleiomorphic CLathrin-Independent Carriers (CLICs) and GPI-Enriched Endocytic Compartments (GEECs) as related major players in such uptake [3, 4]. This CLIC/GEEC pathway relies upon cellular signaling and activation through small G proteins, but mechanistic insight into the biogenesis of its tubular and tubulovesicular carriers is lacking. Here we show that the Rho-GAP-domain-containing protein GRAF1 marks, and is indispensable for, a major Clathrin-independent endocytic pathway. This pathway is characterized by its ability to internalize bacterial exotoxins, GPI-linked proteins, and extracellular fluid. We show that GRAF1 localizes to PtdIns(4,5)P2-enriched, tubular, and punctate lipid structures via N-terminal BAR and PH domains. These membrane carriers are relatively devoid of caveolin1 and flotillin1 but are associated with activity of the small G protein Cdc42. This study provides the first specific noncargo marker for CLIC/GEEC endocytic membranes and demonstrates how GRAF1 can coordinate small G protein signaling and membrane remodeling to facilitate internalization of CLIC/GEEC pathway cargoes.

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    • "CLDN18 protein was observed in the plasma membrane of epithelial cells lining the gastric pit region and at the base of the gastric glands as previously reported in normal human stomach specimens (Sahin et al., 2008; Figure 4A). ARHGAP26 was previously detected on pleiomorphic tubular and punctate membrane structures in HeLa cells (Lundmark et al., 2008). We observed ARHGAP26 in normal stomach on vesicular structures "
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    • "Nevertheless, these arguments all highlight the complexities in studying vesicular uptake. It is also worth mentioning that there are several other clathrin-independent endocytic pathways, such as the recently described CLIC/GLEEC pathway (78, 152). The extent to which these other pathways may be involved remains to be determined. "
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    • "The activity of Arf1 itself appears to be regulated by a GEF called GBF1 at the cell surface [8]. The Bar-domain protein GRAF1 participates in the post-endocytic dynamics of the GEECs [15]. It is clear that a host of other core and peripheral molecules must drive the trafficking of these compartments and their cargo to specific destinations inside the cell. "
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