The Arf6 GTPase-activating Proteins ARAP2 and ACAP1 Define Distinct Endosomal Compartments That Regulate Integrin α5β1 Traffic

Chen, Pei-Wen; Luo, Ruibai; Jian, Xiaoying; Randazzo, Paul A.

doi:10.1074/jbc.m114.596155

Cited by 45 publications

(68 citation statements)

References 48 publications

(81 reference statements)

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“…They were fixed with 4% paraformaldehyde, and DIC images were taken using Nikon Eclipse E600 microscope (Nikon) with a 100X/1. 21 . Clot retraction was initiated with thrombin (0.1 U/mL), and images were taken at the indicated times (i) and quantified (ii).…”

Section: Discussionmentioning

confidence: 99%

“…(B) Internalized FITC-Fg goes through Rab4-and Rab11-positive compartments in WT mouse platelets. Washed WT mouse platelets (5 3 10 8 /mL) in HEPES Tyrode buffer containing 1 mM Ca 21 were incubated with FITC-Fg (0.05 mg/mL) for the indicated times and then fixed with 2% paraformaldehyde at room temperature. After washing with PBS, the fixed platelets (5 3 10 7 /mL) were allowed to adhere to poly-D-lysinecoated coverslips.…”

Section: Discussionmentioning

confidence: 99%

“…(B) Quantification of platelet surface area when spread on Fg-coated surfaces. Washed platelets from WT and KO mice (2.0 3 10 7 /mL) were supplemented with 1 mM Ca 21 and then incubated on Fgcoated surfaces for the indicated times. They were fixed with 4% paraformaldehyde, and DIC images were taken using Nikon Eclipse E600 microscope (Nikon) with a 100X/1.…”

Section: Discussionmentioning

confidence: 99%

“…7,17,18 Although unknown in platelets, Arf6 clearly affects integrin trafficking in other cells: both endocytosis and recycling. [19][20][21] In cancer cells, active and inactive b 1 integrins traffic via different endocytic recycling routes, although their endocytosis equally requires both clathrin and dynamin. 22 Integrins can be recycled back to the plasma membrane, either via a Rab4-mediated short loop, recycling from early endosomes, or via a Rab11-mediated long loop, recycling from a perinuclear The online version of this article contains a data supplement.…”

Section: Introductionmentioning

confidence: 99%

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Arf6 controls platelet spreading and clot retraction via integrin αIIbβ3 trafficking

Huang¹,

Joshi²,

Xiang

et al. 2016

Blood

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• Arf6 selectively regulates endocytic trafficking of platelet a IIb b 3 .• Endocytosis contributes to acute platelet function.Platelet and megakaryocyte endocytosis is important for loading certain granule cargo (ie, fibrinogen [Fg] and vascular endothelial growth factor); however, the mechanisms of platelet endocytosis and its functional acute effects are understudied. Adenosine 5'-diphosphate-ribosylation factor 6 (Arf6) is a small guanosine triphosphate-binding protein that regulates endocytic trafficking, especially of integrins. To study platelet endocytosis, we generated platelet-specific Arf6 knockout (KO) mice. Arf6 KO platelets had less associated Fg suggesting that Arf6 affects a IIb b 3 -mediated Fg uptake and/or storage. Other cargo was unaffected. To measure Fg uptake, mice were injected with biotinylated-or fluorescein isothiocyanate (FITC)-labeled Fg. Platelets from the injected Arf6 KO mice showed lower accumulation of tagged Fg, suggesting an uptake defect. Ex vivo, Arf6 KO platelets were also defective in FITC-Fg uptake and storage. Immunofluorescence analysis showed initial trafficking of FITC-Fg to a Rab4-positive compartment followed by colocalization with Rab11-positive structures, suggesting that platelets contain and use both early and recycling endosomes. Resting and activated a IIb b 3 levels, as measured by flow cytometry, were unchanged; yet, Arf6 KO platelets exhibited enhanced spreading on Fg and faster clot retraction. This was not the result of alterations in a IIb b 3 signaling, because myosin light-chain phosphorylation and Rac1/RhoA activation were unaffected. Consistent with the enhanced clot retraction and spreading, Arf6 KO mice showed no deficits in tail bleeding or FeCl 3 -induced carotid injury assays. Our studies present the first mouse model for defining the functions of platelet endocytosis and suggest that altered integrin trafficking may affect the efficacy of platelet function. (Blood. 2016;127(11):1459-1467 IntroductionIt is increasingly clear that platelets are capable of different types of intracellular membrane trafficking. Platelet exocytosis and autophagy are critical for hemostasis. [1][2][3][4] Platelet endocytosis is important for granule cargo loading. Platelets selectively take up certain cargo (ie, fibrinogen [Fg] and vascular endothelial growth factor) from plasma and package it into a-granules. [5][6][7][8] However, the molecular mechanisms of platelet endocytosis are largely unknown. It is also unclear whether platelet endocytosis contributes to acute platelet functions beyond granule cargo loading. The goal of this study is to begin to address these fundamental questions.The Ras-like, small guanosine triphosphate (GTP)-binding proteins called adenosine 5'-diphosphate-ribosylation factors (Arf's), are key intracellular trafficking regulators. They cycle between inactive guanosine diphosphate (GDP)-bound and active GTP-bound states and are important for multiple functions (eg, actin cytoskeleton remodeling, lipid metabolism, and vesicle traffickin...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Arf6 controls platelet spreading and clot retraction via integrin αIIbβ3 trafficking

Huang¹,

Joshi²,

Xiang

et al. 2016

Blood

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“…The ARAPs control trafficking of a number of transmembrane receptors including a 5 b 1 integrin and EGFR and affect cell adhesion, migration, insulin secretion and lymphatic vascular development. [64][65][66][67][68][69] ARAP3 has been reported to inhibit dissemination of scirrhous gastric cancer. 70 AGAPs are composed of a G-protein like, PH, Arf GAP and Ank repeat domains.…”

Section: Introductionmentioning

confidence: 99%

Allosteric properties of PH domains in Arf regulatory proteins

et al. 2016

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Pleckstrin Homology (PH) domains bind phospholipids and proteins. They are critical regulatory elements of a number enzymes including guanine nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs) for Ras-superfamily guanine nucleotide binding proteins such as ADP-ribosylation factors (Arfs). Recent studies have indicated that many PH domains may bind more than one ligand cooperatively. Here we discuss the molecular basis of PH domain-dependent allosteric behavior of 2 ADPribosylation factor exchange factors, Grp1 and Brag2, cooperative binding of ligands to the PH domains of Grp1 and the Arf GTPase-activating protein, ASAP1, and the consequences for activity of the associated catalytic domains.

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Integrating intracellular nanovesicles into integrin trafficking pathways and beyond

Larocque

Royle

2022

Cell. Mol. Life Sci.

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Membrane traffic controls the movement of proteins and lipids from one cellular compartment to another using a system of transport vesicles. Intracellular nanovesicles (INVs) are a newly described class of transport vesicles. These vesicles are small, carry diverse cargo, and are involved in multiple trafficking steps including anterograde traffic and endosomal recycling. An example of a biological process that they control is cell migration and invasion, due to their role in integrin recycling. In this review, we describe what is known so far about these vesicles. We discuss how INVs may integrate into established membrane trafficking pathways using integrin recycling as an example. We speculate where in the cell INVs have the potential to operate and we identify key questions for future investigation.

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The Arf6 GTPase-activating Proteins ARAP2 and ACAP1 Define Distinct Endosomal Compartments That Regulate Integrin α5β1 Traffic

Cited by 45 publications

References 48 publications

Arf6 controls platelet spreading and clot retraction via integrin αIIbβ3 trafficking

Arf6 controls platelet spreading and clot retraction via integrin αIIbβ3 trafficking

Allosteric properties of PH domains in Arf regulatory proteins

Integrating intracellular nanovesicles into integrin trafficking pathways and beyond

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