Integrin trafficking in cells and tissues

Abstract

Cell adhesion to the extracellular matrix is fundamental to metazoan multicellularity and is accomplished primarily through the integrin family of cell-surface receptors. Integrins are internalized and enter the endocytic-exocytic pathway before being recycled back to the plasma membrane. The trafficking of this extensive protein family is regulated in multiple context-dependent ways to modulate integrin function in the cell. Here, we discuss recent advances in understanding the mechanisms and cellular roles of integrin endocytic trafficking.

Figure 1. Composition and function of the integrin family.

a, The reported pairing between integrin α and β subunits and the ECM ligand/s for each heterodimer are illustrated for mammals, Drosophila and C. elegans. In mammals, α5β1-integrin binds to the fibronectin RGD motif, whereas α4β1 binds to the fibronectin LDV motif. COL: collagen; E-cad: E-cadherin; FN: fibronectin; ICAM: intercellular cell adhesion molecule; VCAM-1: vascular cell adhesion molecule 1. For a more comprehensive/exhaustive list of integrin ligands, please refer to. b, Integrins are bidirectional signalling molecules. Inside-out signals regulate talin binding to integrin β-tails and thus tightly control integrin affinity for ECM ligands. Subsequent ECM binding triggers recruitment of protein complexes (scaffolding and adaptor proteins, kinases and phosphatases, etc.) to the integrin cytoplasmic tails to promote integrin downstream signalling (outside-in signalling). Integrins can also signal from within endosomes (inside-in signalling) to support FAK activity and suppress anoikis or to promote signalling downstream of co-trafficking MET to support anoikis resistance, tumour growth and cancer cell dissemination to lungs. The superscripted P in red indicates phosphorylation. ERK, extracellular signal-regulated kinase; p52SHC, p52 isoform of SHC-transforming protein 1. c, Integrin-ECM adhesions, in vitro, are defined based on localisation, components and maturation stage. Nascent adhesions (focal complexes) represent initial integrin receptor clustering in response to ECM engagement and recruitment of adaptor and signalling proteins to the integrin tails. These small protein assemblies mature into focal adhesions (FA), which serve to anchor actin stress fibres and are vital for generation of contractile force. Fibrillar adhesions are mature α5β1-integrin adhesions, and prominent sites of fibronectin fibrillogenesis, that result from the centripetal translocation of this specific integrin heterodimer towards the cell body.

Figure 2. Fine-tuning integrin endocytosis and recycling.

a, Summary of integrin trafficking pathways. Integrins are internalised into Rab5-positive early endosomes (EE). EE mature into late endosomes (LE), which fuse with lysosomes (Lys) for integrin degradation. Under certain conditions (Rab25 and chloride intracellular channel protein 3 (CLIC3) expression), integrins exit LE/Lys compartments and are recycled back to the plasma membrane. Integrins also traffic to multivesicular bodies (highlighted by *) and to the perinuclear recycling compartment (PNRC). This process occurs within 20 minutes, while degradation takes several hours resulting in the majority of integrins being recycled back to the cell surface. Each step requires a spatiotemporal hierarchy of interactions between integrins, endocytic adaptors, Ras and Arf GTPase family members and other molecules. The Rab GTPases involved are indicated. b, Integrin receptor internalisation mechanisms, broadly classified as clathrin-mediated endocytosis (CME) or clathrin-independent endocytosis (CIE, including caveolin-dependent pathways, micropinocytosis and clathrin-independent carriers (CLICs)). CDR, circular dorsal ruffles. c, Integrin endocytosis can be fine-tuned by extracellular-initiated signals. Left-hand panel: Syndecan-4(Syn-4)─fibronectin interaction activates PKCα and RhoG and promotes caveolin-dependent α5β1-integrin endocytosis, attenuated adhesion and increased cell migration. PKCα also phosphorylate FMNL2 (recruited by RhoC). Phospho-FMNL2 (superscripted P in red indicates phosphorylation) interaction with the α-integrin GFFKR motif drives αβ1-integrin internalisation. The role of actin nucleation and the extracellular signal regulating this pathway remain undefined. *PKCα activation occurs at the plasma membrane. Right-hand panel: MET receptor─β1-integrin interaction induces integrin endocytosis, collective mesenchymal cell migration, in a HIP1- (clathrin adaptor) and RhoA-dependent manner, and “inside-in” signalling (see Fig. 1b). d, Recruitment of specific endosomal adaptors imposes selectivity to integrin recycling pathways. Left-hand panel: GGA2, an Arf effector has been implicated in Rab13-mediated recycling of β1-integrin (a non-peer-reviewed study). Right-hand panel: Different populations of Rab5-positive EE, defined by the presence of Rab5 effectors EEA1 (not shown), APPL1 and the CORVET complex promote different recycling routes^,,. CORVET mediates fission and fusion of EE that mature into LE. Here, Rab5 is replaced by Rab7 and integrins enter LE. Integrins escape degradation by interaction with SNX17 (components in inset)^–. CORVET components Vps3/Vps8 localise to Rab4-positive endosomes (fusion of APPL1-positive EE). Vps3/Vps8 vesicles deliver integrins to the plasma membrane through Rab11-positive recycling endosomes. SE, sorting endosomes.

Figure 3. Integrin trafficking in development.

a, Retrograde transport of inactive β1-integrin (bottom) is required for polarized cell behaviours during development. Epiblast cell rosette structure fails to form in Rab6 mutant embryos (top). Integrin retrograde traffic is also required for persistent migration of a distal tip cell (DTC) in C. elegans gonad (middle). Knockdown of Rab6 or the retromer component Vps35 causes a DTC pathfinding error and morphogenetic defects. b, Integrin CME during optic cup formation is facilitated by binding of Numb and Numb-like to the β1-integrin NPxY motif and is inhibited by competitive binding of these adaptors to the NPxF motif of the membrane protein Ojoplano (Opo). Loss of Opo or overexpression of adaptors result in excess integrin endocytosis, decreased cortical actin in basal endfeet, failure in basal constriction and a flat retina^,. c, In MTJ, the muscle cell (αPS2βPS-integrins) is attached to the tendon cell (αPS1βPS- and αPS2βPS-integrins) indirectly through ECM. Integrin turnover in MTJ is increased by reduced availability of ECM and decreased by raised muscle tension, elevated integrin outside-in activation, or expression of Rab5 mutants^,. Cleavage of PI(3)P by myotubularin (MTM1) prevents receptor accumulation in endosomal-related inclusions. d, Cellular layers forming Drosophila wing are held together by adhesion of αPS1βPS-integrin (dorsal, D) and αPS2βPS-integrin (ventral layer, V) to the ECM secreted in between. In wing imaginal disc, βPS-integrin trafficking is mediated by Rab11, which when disrupted leads to increased apical cell area, intracellular βPS-integrin accumulation and disorganised actin cytoskeleton. The ensuing change in cell shape from columnar to cuboidal (not shown) leads to separation of cell layers and blisters. DN, dominant negative; mtm mutant, myotubularin mutant. e, Recycling of α5β1- and α6β1-integrins is required for migration of cranial NCC. Integrins are recycled through Rab4 and Rab11 pathways in a laminin substrate-dependent manner. f, α9β1-integrin recycling within the growth cone and long-range axonal integrin traffic are required for efficient axon growth. Rab5-regulated endocytosis is followed by recycling through Rab4, Rab11 or Arf6 pathways. Activation of Arf6 promotes retrograde transport of integrin-containing vesicles towards the neuron body. Chemorepellent cues trigger β1-integrin endocytosis on one side of the growth cone, which changes direction of growth cone migration.