Spatial organization of the plasma membrane is an essential feature of the cellular response to external stimuli. Receptor organization at the cell surface mediates transmission of extracellular stimuli to intracellular signalling molecules and effectors that impact various cellular processes including cell differentiation, metabolism, growth, migration and apoptosis. Membrane domains include morphologically distinct plasma membrane invaginations such as clathrin-coated pits and caveolae, but also less well-defined domains such as lipid rafts and the galectin lattice. In the present chapter, we will discuss interaction between caveolae, lipid rafts and the galectin lattice in the control of cancer cell signalling.

Introduction

The plasma membrane is organized through compartmentalization by the underlying actin membrane skeleton and also by the presence of distinct plasma membrane domains that selectively recruit different classes of lipids and proteins [1]. Domain organization of the plasma membrane is a critical determinant of the spatial distribution and activity of cytokine receptors, adhesion molecules and proximal effectors of signalling that mediate the cellular response to extracellular cues.

Lipid rafts are highly dynamic plasma membrane domains enriched in cholesterol and sphingolipids. Although the existence of lipid rafts has proven to be highly controversial, the fundamental basis of the raft hypothesis, that cholesterol functionally organizes the plasma membrane (and other cellular membranes), is well accepted. Current debate questions the actual nature of a lipid raft and, particularly, the DRM (detergent-resistant membrane) approach used to isolate these dynamic, transient structures. Current definitions of lipid rafts suggest that they are small, transient cholesterol-dependent membrane domains that can form more stable platforms promoting the assembly and interaction of receptors and signalling molecules as well as clathrin-independent endocytosis. For the purposes of this chapter, the terms lipid raft and raft-dependent will refer to cholesterol-dependent membrane domains and we will specifically use DRM when referring to biochemical analysis of these domains.

Caveolae are a subdomain of sphingolipid- and cholesterol-rich lipid rafts that form plasma membrane invaginations 60–80 nm in diameter. The caveolins are key structural components of caveolae. More recently, the cytoplasmic cavins were also shown to be critical to maintain the structure and function of caveolae (Figures 1 and 2).

Cholesterol-dependent membrane domains: caveolae, Cav1 scaffolds and lipid rafts

Figure 1.
Cholesterol-dependent membrane domains: caveolae, Cav1 scaffolds and lipid rafts

Cav1 is associated with caveolae along with cavin-1 and, in some cells, cavin-2. Non-caveolar Cav1 exists at the plasma membrane as oligomerized Cav1 scaffolds. Lipid rafts are cholesterol- and sphingolipid-rich membrane domains, also enriched in glycosylphosphatidylinositol (GPI)-anchored proteins, that vary from highly transient nanoscale domains to larger, more stable structures.

Figure 1.
Cholesterol-dependent membrane domains: caveolae, Cav1 scaffolds and lipid rafts

Cav1 is associated with caveolae along with cavin-1 and, in some cells, cavin-2. Non-caveolar Cav1 exists at the plasma membrane as oligomerized Cav1 scaffolds. Lipid rafts are cholesterol- and sphingolipid-rich membrane domains, also enriched in glycosylphosphatidylinositol (GPI)-anchored proteins, that vary from highly transient nanoscale domains to larger, more stable structures.

Cavins and caveolae

Figure 2.
Cavins and caveolae

Cav1 and cavins are associated with invaginated smooth plasmalemmal vesicles or caveolae. Cavin-1 and cavin-2 (in some tissues) are required for caveolae invagination and cavin-3 for caveolae budding to form caveolar vesicles. Cavin-2 overexpression induces enlarged caveolae and caveolar tubules, defining critical roles for cavin family members in caveolae formation, morphology and dynamics. Modified from [5].

Figure 2.
Cavins and caveolae

Cav1 and cavins are associated with invaginated smooth plasmalemmal vesicles or caveolae. Cavin-1 and cavin-2 (in some tissues) are required for caveolae invagination and cavin-3 for caveolae budding to form caveolar vesicles. Cavin-2 overexpression induces enlarged caveolae and caveolar tubules, defining critical roles for cavin family members in caveolae formation, morphology and dynamics. Modified from [5].

Galectin–glycoprotein lattices form another class of plasma membrane domains that regulates diverse cellular process including receptor signalling. Galectins are galactose-specific lectins that bind and cross-link N-glycans of cell surface glycoproteins, including growth factor receptors such as EGFR (epidermal growth factor receptor) and TGFβR (transforming growth factor β receptor), integrins and cadherins, and glycolipids to form heterogeneous lattices that regulate receptor dynamics and availability to ligand (Figure 3).

Mgat5 and the Gal3 lattice

Figure 3.
Mgat5 and the Gal3 lattice

Modification of N-glycans of newly synthesized glycoproteins takes place in the medial Golgi by N-acetylglucosaminyltransferases (only Mgat5, the N-acetylglucosaminyltransferase responsible for β1–6 branching, is shown in this schematic). Further modification of terminal branches with galactose and sialic acid takes place in medial and trans Golgi cisternae. At the cell surface, glycoproteins with branched glycans display increased affinities for galectins. Interaction of glycoproteins and glycolipids with chimeric Gal3, containing a C-terminal CRD and a non-lectin N-terminal domain responsible for oligomerization and pentamer formation, leads to formation of the cell surface galectin lattice.

Figure 3.
Mgat5 and the Gal3 lattice

Modification of N-glycans of newly synthesized glycoproteins takes place in the medial Golgi by N-acetylglucosaminyltransferases (only Mgat5, the N-acetylglucosaminyltransferase responsible for β1–6 branching, is shown in this schematic). Further modification of terminal branches with galactose and sialic acid takes place in medial and trans Golgi cisternae. At the cell surface, glycoproteins with branched glycans display increased affinities for galectins. Interaction of glycoproteins and glycolipids with chimeric Gal3, containing a C-terminal CRD and a non-lectin N-terminal domain responsible for oligomerization and pentamer formation, leads to formation of the cell surface galectin lattice.

Lipid-, protein- and glycan-dependent interactions are therefore key determinants of the activity of a variety of cellular receptors, such as receptor tyrosine kinases, G-protein-coupled receptors, integrins and cadherins, that mediate the transmission of extracellular signals, cell adhesion and cell migration. Expression of both Cav1 (caveolin-1) and Gal3 (galectin-3) are closely associated with cell migration and cancer metastasis. These plasma membrane domain effectors promote protein and lipid recruitment to domains, including lipid rafts, where homotypic and heterotypic clustering regulates the dynamics and functionality of plasma membrane molecules. How these interactions impact cancer cell signalling, adhesion and migration is the subject of this chapter.

Caveolin-1: caveolae and other roles

Cav1 is the caveola coat protein and is required for caveola formation. Cav2 and Cav3 are the other members of the caveolin family. Cav2 is co-expressed with Cav1 and is required for Cav1 stabilization and plasma membrane localization. Work on Cav2 is limited relative to Cav1, particularly with respect to cancer cell signalling, and the present chapter will not address Cav2, although further study of Cav2 is certainly warranted. Cav3 is muscle specific but is also expressed in glial cells, and it plays an essential role in caveolae biogenesis in tissues where it is expressed [2]. A single caveola contains approximately 150 caveolin molecules. However, newly synthesized Cav1 forms low-molecular-mass oligomers in the endoplasmic reticulum and travels to the Golgi apparatus, where it undergoes oligomerization and palmitoylation [2]. Non-caveolar roles for Cav1 and evidence for the existence of smaller non-invaginated Cav1 domains (or scaffolds) have been reported [3,4] (Figure 1).

Recently, another family of cytoplasmic proteins, the cavins, was shown to be important for the formation of caveolae [5]. Amongst the cavins, cavin-1, also known as PTRF (polymerase I and transcript release factor), is critical for the formation of functional caveolae and stabilization of Cav1 expression. Other members of the cavins include cavin-2 [also known as SDPR (serum deprivation response protein)], cavin-3 [also known as SRBC (serum deprivation response-related gene product that binds to c-kinase)] and cavin-4 [also known as MURC (muscle-restricted coiled-coil protein)]. Cavin-2 promotes caveolar membrane curvature and cavin-3 affects the formation of caveolar endocytic carriers. Cavin-4 is predominantly expressed in cardiac and skeletal muscle and has been linked to myogenesis and muscle hypertrophy [6]. Loss of cavin-1 results in destabilization and degradation of Cav1, and cavin-1−/− mice show both an absence of caveolae and reduced levels of caveolins [7]. Cavin-1 interacts directly with caveolin-1 in a 4:1 stoichiometry independently of cavin-2 and cavin-3. Cavin-2 has been shown to be required for caveolae expression, but only in endothelial cells of select tissues (Figure 2) [8,9]

Cav1 is a 178-amino acid long protein (22 kDa). The cytoplasmic N- and C-termini of Cav1 are separated by a 33-amino-acid-long hydrophobic region (residues 102–134) imparting Cav1 a hairpin structure [10] (Figure 4). Cav1 has several distinct domains, including an oligomerization domain (residues 61–101), and a CSD (caveolin scaffolding domain) (residues 82–101). Cav1 phosphorylation at Ser80 converts Cav1 into a soluble secreted protein. Cav1 directly interacts with and regulates the activity of signalling molecules via its CSD. An aromatic-rich CBM (caveolin-binding motif) (φXφXXXXφ, φXXXXφXXφ or φXφXXXXφXXφ, where φ is an aromatic and X an unspecified amino acid), has been reported in Cav1-interacting partners [11,12]. However, the role of the CBM has been recently questioned; indeed, presence of a CBM is not an absolute criterion for Cav1 binding, and in fact, many proteins with putative CBMs do not interact with Cav1 and many Cav1-binding molecules lack CBMs [13]. Various receptors and signalling molecules such as GPCRs (G-protein-coupled receptors), receptor tyrosine kinases, integrins, steroid hormone receptors, and downstream molecules such as heterotrimeric G-proteins, ion channels and NO synthase interact with Cav1. Receptor interaction with Cav1 leads predominantly, but not exclusively, to inhibition of receptor signalling and consequent reduction of cell proliferation and migration, and in some cases, induction of cell death [14,15] (Figure 5). More recently, caveolae have been shown to control membrane organization and Ras signalling via regulation of membrane lipid composition [16]. The role of caveolins, cavins and caveolae in cellular signalling is therefore complex and still not fully understood.

Cav1 protein structure

Figure 4.
Cav1 protein structure

Both N- and C-terminal regions of Cav1 face the cytoplasm. A mutation at position 132 (P132 L) located close to C-terminal palmitoylation sites disrupts CSD function and Cav1 tumour suppressor function in breast cancer. The Cav1 CSD is a highly conserved region situated between D82 and R101 that mediates interaction with various signalling molecules including tyrosine kinase receptors, G-proteins, PKC, eNOS (endothelial nitric oxide synthase), ERK (extracellular-signal-regulated kinase), Ras family GTPases and phospholipases. Cav1 is phosphorylated at Ser80 and Tyr14. Src, Fyn or Abl-dependent Tyr14 phosphorylation recruits Src, Grb7, Csk, Fyn and c-Abl and has been linked to various cellular phenomena including mechanotransduction, signal transduction, endocytosis, cell migration and focal adhesion dynamics. Modified from [15].

Figure 4.
Cav1 protein structure

Both N- and C-terminal regions of Cav1 face the cytoplasm. A mutation at position 132 (P132 L) located close to C-terminal palmitoylation sites disrupts CSD function and Cav1 tumour suppressor function in breast cancer. The Cav1 CSD is a highly conserved region situated between D82 and R101 that mediates interaction with various signalling molecules including tyrosine kinase receptors, G-proteins, PKC, eNOS (endothelial nitric oxide synthase), ERK (extracellular-signal-regulated kinase), Ras family GTPases and phospholipases. Cav1 is phosphorylated at Ser80 and Tyr14. Src, Fyn or Abl-dependent Tyr14 phosphorylation recruits Src, Grb7, Csk, Fyn and c-Abl and has been linked to various cellular phenomena including mechanotransduction, signal transduction, endocytosis, cell migration and focal adhesion dynamics. Modified from [15].

Roles of Cav1 and Gal3 in signalling at the plasma membrane

Figure 5.
Roles of Cav1 and Gal3 in signalling at the plasma membrane

Interaction of Cav1 with receptor tyrosine kinases such as EGFR inhibits signalling and reduces cell proliferation, supporting a tumour suppressor function for Cav1. EGFR recruitment by Gal3 to the galectin enhances receptor signalling by both promoting cell surface residency and limiting interaction with negative regulatory Cav1 domains stimulating tumour cell proliferation and tumour growth. Synergistic activity of Gal3 and pCav1 at focal adhesions leads to integrin activation and pCav1/Src-dependent activation of RhoA GTPase and downstream ROCK signalling associated with enhanced tumour cell migration and metastasis.

Figure 5.
Roles of Cav1 and Gal3 in signalling at the plasma membrane

Interaction of Cav1 with receptor tyrosine kinases such as EGFR inhibits signalling and reduces cell proliferation, supporting a tumour suppressor function for Cav1. EGFR recruitment by Gal3 to the galectin enhances receptor signalling by both promoting cell surface residency and limiting interaction with negative regulatory Cav1 domains stimulating tumour cell proliferation and tumour growth. Synergistic activity of Gal3 and pCav1 at focal adhesions leads to integrin activation and pCav1/Src-dependent activation of RhoA GTPase and downstream ROCK signalling associated with enhanced tumour cell migration and metastasis.

Tyrosine phosphorylated caveolin-1

Cav1 is phosphorylated at Tyr14 by Src and other tyrosine kinases such as Fyn and c-Abl in response to various stimuli including EGF (epidermal growth factor), PDGF (platelet-derived growth factor), IGF (insulin-like growth factor), PEDF (pigment epithelium-derived factor) and stress [14]. Early studies localized pCav1 to focal adhesions (large macromolecular assemblies that mediate cell–substrate adhesion and integrin signalling); however, Hill et al. [17] demonstrated that the anti-pCav1 monoclonal antibody cross-reacts with phospho-paxillin, questioning the localization of pCav1 to focal adhesions. However, studies using the fluorescent probe laurdan revealed that membrane order in focal adhesions is affected by pCav1 expression [18] and loss of adhesion triggers pCav1-dependent internalization of raft components [19]. pCav1 might then regulate focal adhesion signalling via raft organization and composition within focal adhesions. Cav1 Tyr14 phosphorylation is not required for caveolae formation and caveolae are localized predominantly to the rear of a migrating cell [20,21]. As such, the membrane domain that mediates pCav1-induced raft-dependent internalization and pCav1-dependent focal adhesion dynamics at the leading edge of migrating cells remains to be determined.

In metastatic cancer cells, Src-dependent Cav1 phosphorylation activates RhoA and FAK (focal adhesion kinase) leading to Rho–ROCK (Rho-associated protein kinase) and Src-dependent focal adhesion dynamics, actin cytoskeleton remodelling, and tumour cell migration and invasion [2224]. pCav1 recruits several SH2 domain-containing proteins such as Csk or Grb7 [25,26]. Csk activation induces downstream activation of RhoA and Rho-kinase, leading to myosin light chain phosphorylation and cytoskeletal rearrangements and extracellular matrix remodelling [27]. pCav1 (and Gal3; see below) is a critical mediator of the EGF-induced migratory response [23]. Cav1 phosphorylation induced by EGF is associated with increased caveolae [28]. More recently, pCav1 was shown to act as a mechanotransducer, converting a mechanical stimulus into biochemical activity, that phosphorylates and inactivates the Egr1 (early growth response-1) transcription factor, relieving inhibition of Cav1 and cavin-1 promoter activity via PKC (protein kinase C) signalling [20]. Biosynthesis of Cav1 and induction of caveolae may contribute to maintenance of membrane integrity in response to mechanical stress (i.e. an external force applied to cells by the surrounding environment) [29] (Figure 6). pCav1 therefore promotes focal adhesion dynamics, integrin signalling, growth factor-induced cell migration and mechanotransduction.

Cav1 and mechanotransduction

Figure 6.
Cav1 and mechanotransduction

Caveolae, containing Cav1 and cavin-1, are present at the plasma membrane of resting cells. Upon mechanical stress, caveolae flatten out to provide additional membrane and thereby buffer membrane tension. In parallel, cell stretching and associated actin stress fibre formation and focal adhesion maturation induces Cav1 phosphorylation. pCav1 signals through PKC to phosphorylate and inactivate the Egr1 transcription factor, relieving inhibition of Cav1 and cavin-1 promoter activity and enabling formation of additional caveolae that contribute to maintenance of membrane integrity in response to mechanical stress.

Figure 6.
Cav1 and mechanotransduction

Caveolae, containing Cav1 and cavin-1, are present at the plasma membrane of resting cells. Upon mechanical stress, caveolae flatten out to provide additional membrane and thereby buffer membrane tension. In parallel, cell stretching and associated actin stress fibre formation and focal adhesion maturation induces Cav1 phosphorylation. pCav1 signals through PKC to phosphorylate and inactivate the Egr1 transcription factor, relieving inhibition of Cav1 and cavin-1 promoter activity and enabling formation of additional caveolae that contribute to maintenance of membrane integrity in response to mechanical stress.

Galectin-3, the galectin lattice and interaction with lipid rafts

Most cell surface receptors are glycoproteins whose binding to lectins forms multivalent lattices or nanodomains that regulate glycoprotein mobility. The galectins are N-acetyllactosamine (Galβ1–3GlcNAc or Galβ1–4GlcNAc) binding proteins that bind preferentially to Golgi-remodelled N-glycans of cell surface glycoproteins [30]. Golgi N-acetylglucosaminyltransferase V (encoded by Mgat5) increases β1–6 branching of N-glycans and affinity for galectins [31] (Figure 3). Mgat5 overexpression and β1–6 N-glycan branching are elevated in malignant cancers, increasing receptor association with cell surface Gal3 and enhancing signalling. Dennis and colleagues showed that both the number of N-glycans carried by a glycoprotein and also metabolic flux through the hexosamine pathway to UDP-GlcNAc, the donor substrate common to the Golgi Mgat enzymes, affect β1–6 branching and glycoprotein recruitment to the galectin lattice. Intriguingly, a differential number of N-glycans on receptors that drive growth versus differentiation provides a mechanism for metabolic control of cellular growth and arrest [32]. Lattice affinity of glycoproteins is therefore regulated by N-glycan number, Mgat5 activity, metabolic flux and galectin expression.

The chimaera-type Gal3 has the C-terminal CRD (carbohydrate recognition domain) common to the 15 galectins but also a long N-terminal non-lectin domain responsible for interaction between subunits facilitating oligomerization. Gal3 exists as monomers but forms multimeric structures (up to pentamers) driven by the density of multivalent glycoprotein ligands, resulting in the formation of a lattice having irregular geometry [31]. The Mgat5-dependent Gal3 lattice promotes tyrosine kinase receptor signalling by decreasing receptor internalization after stimulation and counteracting Cav1 negative regulation of receptor tyrosine kinase by sequestering the receptor outside of negative regulatory Cav1 scaffolds [33,34] (Figure 5). Gal3 promotes integrin-dependent epithelial cell migration, and interaction of Gal3 with Mgat5-modified N-glycans stimulates cancer cell motility by promoting integrin activation, focal adhesion turnover and matrix remodelling in addition to destabilizing cadherin-based cell–cell junctions [30]. Mgat5 and Gal3 gene expression are upregulated in cancer cells and, together with metabolic input through UDP-GlcNAc, recruit receptors to the galectin lattice promoting the cell signalling, adhesion and migration that underlie cancer progression and metastasis [31].

Lectin–glycan interactions play important roles in eliciting a proper immune response. For example, the galectin lattice plays a role in B cell maturation and also in developing B cell tolerance [35]. Gal3 lattice also activates neutrophils to phagocytosis and mast cell degranulation by clustering ligands at the cell surface. To maintain adequate cell-surface cytokine receptor expression for cell motility and phagocytosis, macrophages require Mgat5-dependent galectin–glycoprotein lattice formation [34]. The Mgat5-dependent Gal3 lattice is also a key regulator of T-cell function. The Gal3 immunomodulator activity can both trigger T-cell death and control T-cell activation thresholds. Lipid rafts are critical platforms for TCR (T-cell receptor) signalling. TCR cross-linking induces lipid raft aggregation and LCK and TCR enrichment in these domains, enabling LCK-dependent TCR phosphorylation stimulating downstream signalling events (see Chapters 13 [35a] and 14 [35b]). In this context, the actin cytoskeleton and Gal3 lattice dynamically regulate TCR signalling by controlling raft association of T-cell receptors; whereas TCR binding to actin promotes accumulation in rafts, Gal3 sequesters TCR out of rafts, favouring a quiescent state [36]. The idea that Gal3 sequesters glycoproteins out of lipid raft domains is supported by the Gal3-dependent sequestration of EGFR from Cav1 scaffolds, proteomics data showing that competitive disruption of the galectin lattice with lactose increases protein association with DRMs and Gal3-dependent sequestration of glycoproteins away from lipid rafts for incorporation into apically targeted vesicles in the secretory pathway of polarized epithelial MDCK cells [33,37,38].

Galectins can also bind glycosphingolipids with lower affinity in lipid rafts [30] (Figure 3). Mobility of GM1 glycosphingolipids (using cholera toxin B subunit as a reporter) and N-cadherin at cell–cell junctions is enhanced by the Gal3 lattice and promotes increased junctional turnover [37]. Mgat5 expression promotes N-cadherin junctional dynamics via N-glycan branching [39]. However, in cells treated with mannosidase II inhibitor, preventing N-glycan branching, Gal3 enhanced glycosphingolipid mobility, suggesting that Gal3 interacts directly with the glycosphingolipid [37]. Increased mobility of N-cadherin and glycosphingolipids in junctions upon Gal3 binding may be due to the ability of Gal3 to act as a spacer that limits interaction and enhances lateral mobility of glycoproteins and/or glycolipids, thereby accelerating turnover of stable N-cadherin cell adhesions. Indeed, Gal3 is internalized via a non-clathrin raft pathway to apical endosomes, enriched in tubular endosomal intermediates of the CLIC (clathrin-independent carrier)/GEEC (glycosylphosphatidylinositol-enriched early endosomal compartment) raft endocytic pathway and induces the raft-dependent endocytosis of integrin [4042]. This suggests that multivalent galectin interaction with both glycoproteins and glycolipids can impact raft organization, dynamics and function [30].

Co-ordinate Cav1–Gal3 signalling in cancer

The Gal3 lattice also regulates signalling events through Cav1 expression and function. Indeed Mgat5 expression is associated with Cav1 overexpression and phosphorylation [33,43]. Cav1 was originally considered to be a tumour suppressor gene because of its role as an inhibitor of cytokine receptor signalling and cell growth and identification of the sporadic P132 L mutation associated with breast cancer [15]. However, in contrast with its apparent tumour suppressor function, Cav1 expression is associated with a poor prognosis in several different cancer types, including breast and prostate cancer. To explain the diverse roles of Cav1 in various cancers, we showed that Gal3 lattice expression is a determinant of Cav1 tumour promoter or suppressor activity [4,15]. Cav1 oligomers in the plasma membrane prevent EGFR activation [33,44]. This is counteracted by Gal3 lattice clustering, which prevents EGFR internalization, retains receptors outside of Cav1 scaffolds and amplifies the EGFR signal [33,34] (Figure 5). Receptor recruitment by the Mgat5–galectin-dependent lattice competes with negative regulatory Cav1-containing domains and might play a role in enabling Cav1 tumour promoter activity. Cav1 can therefore be regarded as a conditional tumour suppressor whose suppressor function is dependent on expression of Mgat5 and the galectin lattice [4] (Figure 5) and potentially other molecular modulators of Cav1 function.

By promoting cell migration through regulation of raft organization at the level of focal adhesions, Cav1 phosphorylation might contribute to the dual role of Cav1 in cancer progression. pCav1 and the galectin lattice promote raft organization within focal adhesions and raft-dependent internalization of integrins and signalling molecules [18,19]. Furthermore, clustering of activated integrins is dependent on intact lipid rafts, suggesting that activation and subsequent clustering of integrins are favoured in an ordered membrane domain [45]. We observed that the Mgat5–Gal3 lattice induces α5β1 integrin activation [46] and downstream integrin- and Src-dependent RhoA–ROCK activation via pCav1 leading to focal adhesion turnover and tumour cell migration [23,43]. pCav1 and Gal3 are both required for EGF-induced cell migration and formation of circular dorsal ruffles, macropinocytotic actin cytoskeletal structures involved in integrin trafficking in response to cytokine-induced migration [23]. This suggests that Gal3 clustering and activation of integrins might recruit pCav1 [47,48], leading to pCav1-dependent Src–RhoA–ROCK signalling and cell migration [23,24,43] (Figure 5). However the extent to which Gal3 is involved in pCav1-dependent internalization processes remains to be determined.

In tumour cells, Mgat5 and Gal3 expression can therefore override tumour suppressor function of Cav1, but also work in concert with pTyr14 Cav1 to promote tumour cell migration [4] (Figure 5). Consistently, co-expression of Cav1 and Gal3 promote tumour cell migration, RhoA activation and FAK stabilization in focal adhesions of DTC (differentiated thyroid cancer) cell lines. Co-ordinate expression of Cav1 and Gal3 was found to represent a highly precise marker for DTC diagnosis using clinically relevant patient samples representing in vivo validation of a synergistic role of these two domain effectors in cancer progression [49].

Concluding remarks

Complex interactions between membrane domains and their effectors, such as Cav1 and Gal3, influence cell surface receptor activity and downstream signalling and affect various aspects of cell fate decisions, including but certainly not limited to, cancer cell migration and metastasis.

Summary

  • The temporal and spatial organization of the plasma membrane regulates interaction between receptors and downstream signalling molecules to control the cellular response to extracellular signals.

  • Lipid rafts are cholesterol- and sphingolipid-rich membrane domains that vary from highly transient domains to larger, more stable structures and serve as a platform for receptor signalling.

  • Caveolins and cavins are critical components of caveolae, small plasma membrane invaginations involved in diverse cellular processes ranging from cholesterol homoeostasis, endocytosis, cell migration, and mechanotransduction. Cav1 regulates multiple cancer-associated cellular processes and acts as both tumour promoter and suppressor.

  • Galectins are galactose-specific lectins that bind and cross-link N-glycans of cell surface glycoproteins as well as glycolipids to form heterogeneous lattices that regulate receptor dynamics and availability to ligand.

  • In cancer cells, Gal3 and tyrosine phosphorylated Cav1 act synergistically to promote tumour cell migration by promoting focal adhesion turnover, integrin activation and actin cytoskeleton reorganization.

J.S. is the recipient of a fellowship from the Canadian Breast Cancer Foundation, BC/Yukon region (CBCF). Work on caveolin-1 and galectin-3 in the Nabi lab is supported by grants from the Canadian Institutes for Health Research [grant number CIHR MOP-126029] and the Cancer Research Society. We apologize for our inability to cite all of the work that contributed to our current understanding of the field due to limitations on the number of references. We thank Jim Dennis and Radu Stan for critical reading of the text.

References

References
1.
Kusumi
 
A.
Fujiwara
 
T.K.
Chadda
 
R.
Xie
 
M.
Tsunoyama
 
T.A.
Kalay
 
Z.
Kasai
 
R.S.
Suzuki
 
K.G.
 
Dynamic organizing principles of the plasma membrane that regulate signal transduction: commemorating the fortieth anniversary of Singer and Nicolson's fluid-mosaic model
Annu. Rev. Cell Dev. Biol.
2012
, vol. 
28
 (pg. 
215
-
250
)
2.
Parton
 
R.G.
Simons
 
K.
 
The multiple faces of caveolae
Nat. Rev. Mol. Cell Biol.
2007
, vol. 
8
 (pg. 
185
-
194
)
3.
Head
 
B.P.
Insel
 
P.A.
 
Do caveolins regulate cells by actions outside of caveolae?
Trends Cell Biol.
2007
, vol. 
17
 (pg. 
51
-
57
)
4.
Lajoie
 
P.
Goetz
 
J.G.
Dennis
 
J.W.
Nabi
 
I.R.
 
Lattices, rafts, and scaffolds: domain regulation of receptor signaling at the plasma membrane
J. Cell Biol.
2009
, vol. 
185
 (pg. 
381
-
385
)
5.
Nabi
 
I.R.
 
Cavin fever: regulating caveolae
Nat. Cell Biol.
2009
, vol. 
11
 (pg. 
789
-
791
)
6.
Parton
 
R.G.
del Pozo
 
M.A.
 
Caveolae as plasma membrane sensors, protectors and organizers
Nat. Rev. Mol. Cell Biol.
2013
, vol. 
14
 (pg. 
98
-
112
)
7.
Liu
 
L.
Brown
 
D.
McKee
 
M.
Lebrasseur
 
N.K.
Yang
 
D.
Albrecht
 
K.H.
Ravid
 
K.
Pilch
 
P.F.
 
Deletion of Cavin/PTRF causes global loss of caveolae, dyslipidemia, and glucose intolerance
Cell Metab.
2008
, vol. 
8
 (pg. 
310
-
317
)
8.
Hansen
 
C.G.
Shvets
 
E.
Howard
 
G.
Riento
 
K.
Nichols
 
B.J.
 
Deletion of cavin genes reveals tissue-specific mechanisms for morphogenesis of endothelial caveolae
Nat. Commun.
2013
, vol. 
4
 pg. 
1831
 
9.
Ludwig
 
A.
Howard
 
G.
Mendoza-Topaz
 
C.
Deerinck
 
T.
Mackey
 
M.
Sandin
 
S.
Ellisman
 
M.H.
Nichols
 
B.J.
 
Molecular composition and ultrastructure of the caveolar coat complex
PLoS Biol.
2013
, vol. 
11
 pg. 
e1001640
 
10.
Parton
 
R.G.
 
Caveolae and caveolins
Curr. Opin. Cell Biol.
1996
, vol. 
8
 (pg. 
542
-
548
)
11.
Couet
 
J.
Li
 
S.
Okamoto
 
T.
Ikezu
 
T.
Lisanti
 
M.P.
 
Identification of peptide and protein ligands for the caveolin-scaffolding domain. Implications for the interaction of caveolin with caveolae-associated proteins
J. Biol. Chem.
1997
, vol. 
272
 (pg. 
6525
-
6533
)
12.
Okamoto
 
T.
Schlegel
 
A.
Scherer
 
P.E.
Lisanti
 
M.P.
 
Caveolins, a family of scaffolding proteins for organizing ‘preassembled signaling complexes’ at the plasma membrane
J. Biol. Chem.
1998
, vol. 
273
 (pg. 
5419
-
5422
)
13.
Collins
 
B.M.
Davis
 
M.J.
Hancock
 
J.F.
Parton
 
R.G.
 
Structure-based reassessment of the caveolin signaling model: do caveolae regulate signaling through caveolin-protein interactions?
Dev. Cell
2012
, vol. 
23
 (pg. 
11
-
20
)
14.
Boscher
 
C.
Nabi
 
I.R.
 
Caveolin-1: role in cell signaling
Adv. Exp. Med. Biol.
2012
, vol. 
729
 (pg. 
29
-
50
)
15.
Goetz
 
J.G.
Lajoie
 
P.
Wiseman
 
S.M.
Nabi
 
I.R.
 
Caveolin-1 in tumor progression: the good, the bad and the ugly
Cancer Metastasis Rev.
2008
, vol. 
27
 (pg. 
715
-
735
)
16.
Ariotti
 
N.
Fernández-Rojo
 
M.A.
Zhou
 
Y.
Hill
 
M.M.
Rodkey
 
T.L.
Inder
 
K.L.
Tanner
 
L.B.
Wenk
 
M.R.
Hancock
 
J.F.
Parton
 
R.G.
 
Caveolae regulate the nanoscale organization of the plasma membrane to remotely control Ras signaling
J. Cell Biol.
2014
, vol. 
204
 (pg. 
777
-
792
)
17.
Hill
 
M.M.
Scherbakov
 
N.
Schiefermeier
 
N.
Baran
 
J.
Hancock
 
J.F.
Huber
 
L.A.
Parton
 
R.G.
Parat
 
M.O.
 
Reassessing the role of phosphocaveolin-1 in cell adhesion and migration
Traffic
2007
, vol. 
8
 (pg. 
1695
-
1705
)
18.
Gaus
 
K.
Le Lay
 
S.
Balasubramanian
 
N.
Schwartz
 
M.A.
 
Integrin-mediated adhesion regulates membrane order
J. Cell Biol.
2006
, vol. 
174
 (pg. 
725
-
734
)
19.
del Pozo
 
M.A.
Balasubramanian
 
N.
Alderson
 
N.B.
Kiosses
 
W.B.
Grande-Garcia
 
A.
Anderson
 
R.G.
Schwartz
 
M.A.
 
Phospho-caveolin-1 mediates integrin-regulated membrane domain internalization
Nat. Cell Biol.
2005
, vol. 
7
 (pg. 
901
-
908
)
20.
Joshi
 
B.
Bastiani
 
M.
Strugnell
 
S.S.
Boscher
 
C.
Parton
 
R.G.
Nabi
 
I.R.
 
Phosphocaveolin-1 is a mechanotransducer that induces caveola biogenesis via Egr1 transcriptional regulation
J. Cell Biol.
2012
, vol. 
199
 (pg. 
425
-
435
)
21.
Parat
 
M.-O.
Anand-Apte
 
B.
Fox
 
P.L.
 
Differential caveolin-1 polarization in endothelial cells during migration in two and three dimensions
Mol. Biol. Cell
2003
, vol. 
14
 (pg. 
3156
-
3168
)
22.
Joshi
 
B.
Strugnell
 
S.S.
Goetz
 
J.G.
Kojic
 
L.D.
Cox
 
M.E.
Griffith
 
O.L.
Chan
 
S.K.
Jones
 
S.J.
Leung
 
S.P.
Masoudi
 
H.
, et al 
Phosphorylated caveolin-1 regulates Rho/ROCK-dependent focal adhesion dynamics and tumor cell migration and invasion
Cancer Res.
2008
, vol. 
68
 (pg. 
8210
-
8220
)
23.
Boscher
 
C.
Nabi
 
I.R.
 
Galectin-3- and phospho-caveolin-1-dependent outside-in integrin signaling mediates the EGF motogenic response in mammary cancer cells
Mol. Biol. Cell
2013
, vol. 
24
 (pg. 
2134
-
2145
)
24.
Grande-Garcia
 
A.
Echarri
 
A.
de Rooij
 
J.
Alderson
 
N.B.
Waterman-Storer
 
C.M.
Valdivielso
 
J.M.
del Pozo
 
M.A.
 
Caveolin-1 regulates cell polarization and directional migration through Src kinase and Rho GTPases
The J. Cell Biol.
2007
, vol. 
177
 (pg. 
683
-
694
)
25.
Cao
 
H.
Courchesne
 
W.E.
Mastick
 
C.C.
 
A phosphotyrosine-dependent protein interaction screen reveals a role for phosphorylation of caveolin-1 on tyrosine 14: recruitment of C-terminal Src kinase
J. Biol. Chem.
2002
, vol. 
277
 (pg. 
8771
-
8774
)
26.
Lee
 
H.
Volonte
 
D.
Galbiati
 
F.
Iyengar
 
P.
Lublin
 
D.
Bregman
 
D.B.
Wilson
 
M.T.
Campos-Gonzalez
 
R.
Bouzahzah
 
B.
Pestell
 
R.G.
, et al 
Constitutive and growth factor-regulated phosphorylation of caveolin-1 occurs at the same site (Tyr-14) in vivo: identification of a c-Src/Cav-1/Grb7 signaling cassette
Mol. Endocrinol.
2000
, vol. 
14
 (pg. 
1750
-
1775
)
27.
Radel
 
C.
Rizzo
 
V.
 
Integrin mechanotransduction stimulates caveolin-1 phosphorylation and recruitment of Csk to mediate actin reorganization
Am. J. Physiol. Heart Circ. Physiol.
2005
, vol. 
288
 (pg. 
H936
-
945
)
28.
Orlichenko
 
L.
Huang
 
B.
Krueger
 
E.
McNiven
 
M.A.
 
Epithelial growth factor-induced phosphorylation of caveolin 1 at tyrosine 14 stimulates caveolae formation in epithelial cells
J. Biol. Chem.
2006
, vol. 
281
 (pg. 
4570
-
4579
)
29.
Sinha
 
B.
Koster
 
D.
Ruez
 
R.
Gonnord
 
P.
Bastiani
 
M.
Abankwa
 
D.
Stan
 
R.
Butler-Browne
 
G.
Vedie
 
B.
Johannes
 
L.
, et al 
Cells respond to mechanical stress by rapid disassembly of caveolae
Cell
2010
, vol. 
144
 (pg. 
402
-
413
)
30.
Boscher
 
C.
Dennis
 
J.W.
Nabi
 
I.R.
 
Glycosylation, galectins and cellular signaling
Curr. Opin. Cell Biol.
2011
, vol. 
23
 (pg. 
383
-
392
)
31.
Dennis
 
J.W.
Nabi
 
I.R.
Demetriou
 
M.
 
Metabolism, cell surface organization, and disease
Cell
2009
, vol. 
139
 (pg. 
1229
-
1241
)
32.
Lau
 
K.S.
Partridge
 
E.A.
Grigorian
 
A.
Silvescu
 
C.I.
Reinhold
 
V.N.
Demetriou
 
M.
Dennis
 
J.W.
 
Complex N-glycan number and degree of branching cooperate to regulate cell proliferation and differentiation
Cell
2007
, vol. 
129
 (pg. 
123
-
134
)
33.
Lajoie
 
P.
Partridge
 
E.A.
Guay
 
G.
Goetz
 
J.G.
Pawling
 
J.
Lagana
 
A.
Joshi
 
B.
Dennis
 
J.W.
Nabi
 
I.R.
 
Plasma membrane domain organization regulates EGFR signaling in tumor cells
J. Cell Biol.
2007
, vol. 
179
 (pg. 
341
-
356
)
34.
Partridge
 
E.A.
Le Roy
 
C.
Di Guglielmo
 
G.M.
Pawling
 
J.
Cheung
 
P.
Granovsky
 
M.
Nabi
 
I.R.
Wrana
 
J.L.
Dennis
 
J.W.
 
Regulation of cytokine receptors by Golgi N-glycan processing and endocytosis
Science
2004
, vol. 
306
 (pg. 
120
-
124
)
35.
Gauthier
 
L.
Rossi
 
B.
Roux
 
F.
Termine
 
E.
Schiff
 
C.
 
Galectin-1 is a stromal cell ligand of the pre-B cell receptor (BCR) implicated in synapse formation between pre-B and stromal cells and in pre-BCR triggering
Proc. Natl. Acad. Sci. U.S.A.
2002
, vol. 
99
 (pg. 
13014
-
13019
)
35a.
Nika
 
K.
Acuto
 
O.
 
Membrane nanodomains in T-cell antigen receptor signalling
Essays Biochem.
2015
, vol. 
57
 (pg. 
165
-
175
)
35b.
Byrum
 
J.N.
Rodgers
 
W.
 
Membrane–cytoskeleton interactions in cholesterol-dependent domain formation
Essays Biochem.
, vol. 
57
 (pg. 
177
-
187
)
36.
Rabinovich
 
G.A.
Toscano
 
M.A.
Jackson
 
S.S.
Vasta
 
G.R.
 
Functions of cell surface galectin-glycoprotein lattices
Curr. Opin. Struct. Biol.
2007
, vol. 
17
 (pg. 
513
-
520
)
37.
Boscher
 
C.
Zheng
 
Y.Z.
Lakshminarayan
 
R.
Johannes
 
L.
Dennis
 
J.W.
Foster
 
L.J.
Nabi
 
I.R.
 
Galectin-3 protein regulates mobility of N-cadherin and GM1 ganglioside at cell-cell junctions of mammary carcinoma cells
J. Biol. Chem.
2012
, vol. 
287
 (pg. 
32940
-
32952
)
38.
Delacour
 
D.
Greb
 
C.
Koch
 
A.
Salomonsson
 
E.
Leffler
 
H.
Le Bivic
 
A.
Jacob
 
R.
 
Apical sorting by galectin-3-dependent glycoprotein clustering
Traffic
2007
, vol. 
8
 (pg. 
379
-
388
)
39.
Guo
 
H.B.
Johnson
 
H.
Randolph
 
M.
Pierce
 
M.
 
Regulation of homotypic cell-cell adhesion by branched N-glycosylation of N-cadherin extracellular EC2 and EC3 domains
J. Biol. Chem.
2009
, vol. 
284
 (pg. 
34986
-
34997
)
40.
Howes
 
M.
Kirkham
 
M.
Riches
 
J.
Cortese
 
K.
Walser
 
P.
Simpson
 
F.
Hill
 
M.
Jones
 
A.
Lundmark
 
R.
Lindsay
 
M.R.
, et al 
Clathrin-independent carriers form a high capacity endocytic sorting system at the leading edge of migrating cells
J. Cell Biol.
2010
, vol. 
190
 (pg. 
675
-
691
)
41.
Furtak
 
V.
Hatcher
 
F.
Ochieng
 
J.
 
Galectin-3 mediates the endocytosis of β-1 integrins by breast carcinoma cells
Biochem. Biophys. Res. Comm.
2001
, vol. 
289
 (pg. 
845
-
850
)
42.
Straube
 
T.
von Mach
 
T.
Honig
 
E.
Greb
 
C.
Schneider
 
D.
Jacob
 
R.
 
pH-dependent recycling of galectin-3 at the apical membrane of epithelial cells
Traffic
2013
, vol. 
14
 (pg. 
1014
-
1027
)
43.
Goetz
 
J.G.
Joshi
 
B.
Lajoie
 
P.
Strugnell
 
S.S.
Scudamore
 
T.
Kojic
 
L.D.
Nabi
 
I.R.
 
Concerted regulation of focal adhesion dynamics by galectin-3 and tyrosine-phosphorylated caveolin-1
J. Cell Biol.
2008
, vol. 
180
 (pg. 
1261
-
1275
)
44.
Couet
 
J.
Sargiacomo
 
M.
Lisanti
 
M.P.
 
Interaction of a receptor tyrosine kinase, EGF-R, with caveolins. Caveolin binding negatively regulates tyrosine and serine/threonine kinase activities
J. Biol. Chem.
1997
, vol. 
272
 (pg. 
30429
-
30438
)
45.
Leitinger
 
B.
Hogg
 
N.
 
The involvement of lipid rafts in the regulation of integrin function
J. Cell Sci.
2002
, vol. 
115
 (pg. 
963
-
972
)
46.
Lagana
 
A.
Goetz
 
J.G.
Cheung
 
P.
Raz
 
A.
Dennis
 
J.
Nabi
 
I.R.
 
Galectin binding to Mgat5-modified N-glycans regulates fibronectin matrix remodeling in tumor cells
Mol. Cell. Biol.
2006
, vol. 
26
 (pg. 
3181
-
3193
)
47.
Wary
 
K.K.
Mariotti
 
A.
Zurzolo
 
C.
Giancotti
 
F.G.
 
A requirement for caveolin-1 and associated kinase Fyn in integrin signaling and anchorage-dependent cell growth
Cell
1998
, vol. 
94
 (pg. 
625
-
634
)
48.
Wei
 
Y.
Yang
 
X.
Liu
 
Q.
Wilkins
 
J.A.
Chapman
 
H.A.
 
A role for caveolin and the urokinase receptor in integrin-mediated adhesion and signaling
J. Cell Biol.
1999
, vol. 
144
 (pg. 
1285
-
1294
)
49.
Shankar
 
J.
Wiseman
 
S.M.
Meng
 
F.
Kasaian
 
K.
Strugnell
 
S.
Mofid
 
A.
Gown
 
A.
Jones
 
S.J.
Nabi
 
I.R.
 
Coordinated expression of galectin-3 and caveolin-1 in thyroid cancer
J. Pathol.
2012
, vol. 
228
 (pg. 
56
-
66
)