Regulation of cargo-selective endocytosis by dynamin 2 GTPase-activating protein girdin (original) (raw)
Related papers
Cargo-specific recruitment in clathrin and dynamin-independent endocytosis
bioRxiv, 2020
Spatially controlled, cargo-specific endocytosis is essential for development, tissue homeostasis, and cancer invasion and is often hijacked by viral infections 1. Unlike clathrin-mediated endocytosis, which exploits cargo-specific adaptors for selective protein internalization, the clathrin and dynamin-independent endocytic pathway (CLIC-GEEC, CG-pathway) has until now been considered a bulk internalization route for the fluid phase, glycosylated membrane proteins and lipids 2,3. Although the core molecular players of CG endocytosis have been recently defined, no cargo-specific adaptors are known and evidence of selective protein uptake into the pathway is lacking 3. Here, we identify the first cargo-specific adaptor for CG-endocytosis and demonstrate its clinical relevance in breast cancer progression. By combining unbiased molecular characterization and super-resolution imaging, we identified the actin-binding protein swiprosin-1 (EFHD2) as a cargo-specific adaptor regulating int...
2017
Numerous endocytic pathways operate simultaneously at the cell surface. Here we focus on the molecular machinery involved in the generation of endocytic vesicles of the clathrin and dynamin-independent CLIC/GEEC (CG) pathway. This pathway internalises many GPI-anchored proteins and a large fraction of the fluid-phase in different cell types. We developed a real-time TIRF assay using pH-sensitive GFP-GPI to identify nascent CG endocytic sites. The temporal profile of known CG pathway modulators showed that ARF1/GBF1 (GTPase/GEF pair) and CDC42 (RhoGTPase) are recruited sequentially to CG endocytic sites, ∼60s and ∼9s prior to scission. Using a limited RNAi screen, we found several BAR domain proteins affecting CG endocytosis and focused on IRSp53 and PICK1 that have interactions with CDC42 and ARF1 respectively. IRSp53, an I-BAR domain containing protein, was recruited to the plasma membrane at the site of forming CG endocytic vesicles and in its absence, nascent endocytic CLICs, did...
Clathrin-independent pathways of endocytosis
Cold Spring Harbor perspectives in biology, 2014
There are many pathways of endocytosis at the cell surface that apparently operate at the same time. With the advent of new molecular genetic and imaging tools, an understanding of the different ways by which a cell may endocytose cargo is increasing by leaps and bounds. In this review we explore pathways of endocytosis that occur in the absence of clathrin. These are referred to as clathrin-independent endocytosis (CIE). Here we primarily focus on those pathways that function at the small scale in which some have distinct coats (caveolae) and others function in the absence of specific coated intermediates. We follow the trafficking itineraries of the material endocytosed by these pathways and finally discuss the functional roles that these pathways play in cell and tissue physiology. It is likely that these pathways will play key roles in the regulation of plasma membrane area and tension and also control the availability of membrane during cell migration.
Actin and dynamin2 dynamics and interplay during clathrin-mediated endocytosis
The Journal of cell biology, 2014
Clathrin-mediated endocytosis (CME) involves the recruitment of numerous proteins to sites on the plasma membrane with prescribed timing to mediate specific stages of the process. However, how choreographed recruitment and function of specific proteins during CME is achieved remains unclear. Using genome editing to express fluorescent fusion proteins at native levels and live-cell imaging with single-molecule sensitivity, we explored dynamin2 stoichiometry, dynamics, and functional interdependency with actin. Our quantitative analyses revealed heterogeneity in the timing of the early phase of CME, with transient recruitment of 2-4 molecules of dynamin2. In contrast, considerable regularity characterized the final 20 s of CME, during which ∼26 molecules of dynamin2, sufficient to make one ring around the vesicle neck, were typically recruited. Actin assembly generally preceded dynamin2 recruitment during the late phases of CME, and promoted dynamin recruitment. Collectively, our resu...
Discovery of New Cargo Proteins that Enter Cells through Clathrin-Independent Endocytosis
Traffic, 2009
Clathrin-independent endocytosis (CIE) allows internalization of plasma membrane proteins lacking clathrin-targeting sequences, such as the major histocompatibility complex Class I protein (MHCI), into cells. After internalization, vesicles containing MHCI fuse with transferrincontaining endosomes generated from clathrin-dependent endocytosis. In HeLa cells, MHCI is subsequently routed to late endosomes or recycled back out to the plasma membrane (PM) in distinctive tubular carriers. Arf6 is associated with endosomal membranes carrying CIE cargo and expression of an active form of Arf6 leads to the generation of vacuolar structures that trap CIE cargo immediately after endocytosis, blocking the convergence with transferrin-containing endosomes. We isolated these trapped vacuolar structures and analyzed their protein composition by mass spectrometry. Here we identify and validate six new endogenous cargo proteins (CD44, CD55, CD98, CD147, Glut1 and ICAM1) that use CIE to enter cells. CD55 and Glut1 appear to closely parallel the trafficking of MHCI, merging with transferrin endosomes before entering the recycling tubules. In contrast, CD44, CD98 and CD147 appear to directly enter the recycling tubules and bypass the merge with EEA1-positive, transferrin-containing endosomes. This divergent itinerary suggests that sorting may occur along this CIE pathway. Furthermore, the identification of new cargo proteins will assist others studying CIE in different cell types and tissues.
ArhGEF37 assists Dynamin2 during Clathrin-mediated endocytosis
Journal of Cell Science, 2019
Clathrin-mediated endocytosis (CME) engages over 30 proteins to secure efficient cargo and membrane uptake. While the function of most core CME components is well established, auxiliary mechanisms critical for fine-tuning and adaptation remain largely elusive. In this study, we identify ArhGEF37, a currently uncharacterized protein, as novel constituent of CME. Structure prediction together with quantitative cellular and biochemical studies present a unique BAR domain and PI(4,5)P2-dependent protein/membrane interactions. Functional characterization yields accumulation of ArhGEF37 at Dynamin2-rich late endocytic sites and increased endocytosis rates in the presence of ArhGEF37. Together, these results introduce ArhGEF37 as novel regulatory protein involved in endocytosis.