Matriz Extracelular

REVIEW

Extracellular matrix and cell signalling: the dynamic cooperation of integrin, proteoglycan and growth factor receptor

Soo-Hyun Kim, Jeremy Turnbull1 and Scott Guimond1

Division of Biomedical Sciences, St George’s Medical School, University of London, Cranmer Terrace, London SW17 0RE, UK 1Institute of Integrative Biology, Centre for Glycobiology, University of Liverpool, Crown Street, Liverpool L69 7ZB, UK (Correspondence should be addressed to S-H Kim; Email: skim@sgul.ac.uk)

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Abstract

Extracellular matrices (ECM) are secreted molecules that constitute the cell microenvironment, composed of a dynamic and complex array of glycoproteins, collagens, glycosamino- glycans and proteoglycans. ECM provides the bulk, shape and strength of many tissues in vivo, such as basement membrane, bone and cartilage. In vitro, most animal cells can only grow when they are attached to surfaces through ECM. ECM is also the substrate for cell migration. However, ECM provides much more than just mechanical and structural support, with implications in developmental patterning, stem cell niches and cancer. ECM imparts spatial context for signalling events by various cell surface growth factor receptors and adhesion molecules such as integrins. The external physical properties of ECM may also have a role in the signalling process. ECM molecules can be flexible and extendable, and mechanical tension can expose cryptic sites, which could further interact with growth factors or their receptors. ECM proteins and

structures can determine the cell behaviour, polarity, migration, differentiation, proliferation and survival by communicating with the intracellular cytoskeleton and transmission of growth factor signals. Integrins and proteo- glycans are the major ECM adhesion receptors which cooperate in signalling events, determining the signalling outcomes, and thus the cell fate. This review focuses on the emerging concept of spatial cell biology of ECM, especially the current understanding of integrins and heparan sulphate proteoglycans as the essential cellular machineries that sense, integrate and respond to the physical and chemical environ- mental information either by directly connecting with the local adhesion sites or by regulating global cellular processes through growth factor receptor signalling pathways, leading to the integration of both external and internal signals in space and time.

Journal of Endocrinology (2011) 209, 139–151

Extracellular matrix as structural support and binding platform

In order for cells to function, they must be properly supported, having contacts with neighbouring cells and/or the extracellular matrix (ECM). The ECM provides much of the structural support available to parenchymal cells in tissues. In the skin, it provides the dermis and the basement membrane, on which sit the basal cells that give rise to the stratified skin layers. All this provides the tensile strength and flexibility inherent to skin. In other tissues, basement membranes provide anchoring support to epithelial and endothelial cells. The ECM is produced by epithelial cells and stromal cells found within the matrix itself, including fibroblasts, osteoblasts and basal epithelial cells.

The primary proteins present in the ECM and indeed the entire body are the collagens. Collagens are a family of

proteins with at least 29 members; though not all are found in the ECM, they share a common structural motif of helical fibrils formed by three protein subunits. There are many types of collagens present in the ECM and basement membrane, including, but not limited to type I, III, IV, V and the glycosaminoglycan-containing type XI (Hulmes 2002). The primary function of the collagens is to act as the structural support and binding partners for other ECM proteins. Along with collagen, elastin is the major structural protein in the ECM (Kielty et al. 2002). Individual tropoelastin protein subunits are crosslinked together to give the mature elastin fibre. Elastin, along with fibrillin, is responsible for the flexibility inherent in many tissues.

The diverse array of ECM proteins not only support the physical structure of the cell but also various biological functions, largely through their ability to bind multiple interacting partners such as other ECM proteins, growth

Journal of Endocrinology (2011) 209, 139–151 DOI: 10.1530/JOE-10-0377

0022–0795/11/0209–139 q 2011 Society for Endocrinology Printed in Great Britain Online version via http://www.endocrinology-journals.org

factors, signal receptors and the adhesion molecules, which is mediated by the multiple, specific domains present within each protein. The best example is fibronectin (FN), which, like other ECM proteins, is produced by fibroblasts among other cell types. FN exists as a dimer and can bind to collagens and heparan sulphate proteoglycans (HSPGs) (Ruoslahti 1988), also see later section), thus contributing to the structural framework for many cell surface receptor systems. One of the major functional domains contained in FN is the FN type III (FNIII) domain. FnIII domains contain about 100 amino acids in two, anti-parallel b-sheets, which are also present in collagens, neural cell adhesion molecules (NCAMs) and some cytokine receptors. The FNIII domains contain the amino acid sequence responsible for integrin- binding (RGD motif) and heparin-binding domains, which are further discussed later. Two other domains of FN, the FNI and FNII domains, are smaller than the FNIII domains, with 45–60 amino acids, but they share the basic anti-parallel b-sheet structure stabilised by disulphide bonds. The FN domains are primarily responsible for the binding of FN to collagen, fibrin and other FN molecules and are often present in multiple copies within each protein (for example, there are 15 FNIII domains in FN), allowing for multiple interactions, thus providing a protein-binding platform (Ruoslahti 1988). Another well-known example would be laminins, which are mainly present in the basement membranes and partly responsible for providing the tensile strength of the tissue. Laminin consists of three subunits – a, b and g – which come together to form a characteristic cross pattern that can bind to other laminins as well as proteoglycans and other ECM proteins (Colognato & Yurchenco 2000). Moreover, vitronectin can bind to and regulate components of the plasminogen activator signal complex, in addition to its cell adhesion duties (Preissner & Seiffert 1998).

HSPGs are proteoglycans found in ECM with multiple heparan sulphate (HS) side chains covalently coupled to the

core protein. HSPGs present in the matrix include perlecan, agrin, collagen type XI, syndecans and glypicans. The perlecan, agrin and collagens are actively secreted into

the ECM, while the syndecans and glypicans are cleaved from the cell surface by proteases and phospholipases respectively (Brunner et al. 1994, Manon-Jensen et al.

2010). Secreted HSPGs bind to almost all of the structural proteins in the ECM via both protein–protein interactions and HS–protein interactions. The cleaved HSPGs interactions are primarily, but not exclusively, via HS chains.

Features of the physical adhesion surface and cell signalling

Studies using fabricated inert matrix substrates such as polyethylene glycol have suggested that the binding interaction of cell surface receptors to specific adhesion ligands can be purely dependent on the mechanical attributes of the surface (Marastoni et al. 2008), and the matrix stiffness has profound effects on cell fate and behaviour (Discher et al.

2009). For example, mesenchymal stem cells differentiate into specific cell types only when grown on the appropriate physiological stiffness; thus, matrix elasticity can direct stem cell lineage specification (Engler et al. 2006). Matrix stiffness also has effects on cell migration, proliferation and survival (Wells 2008), and focal adhesions can form and grow only if they experience pulling forces through their cytoskeleton. It is known that growth on soft substrates leads to smaller focal adhesions, containing less phosphotyrosine, and reduced cytoskeletal organisation (Pelham & Wang 1997); how changes in focal adhesion size and composition as well as other tension-dependent mechanisms drive the genetic programs responsible for the differential responses is under investigation. Through the recent progress in nanotechnol- ogy, it is now possible to engineer specific nanopatterned surfaces, providing new insights into the mechanical proper- ties of ECM (Geiger et al. 2001).

Integrins, the inside-out and outside-in signalling

Cells respond to the mechanical and biochemical changes in ECM through the crosstalk between integrins and the actin cytoskeleton. Integrins are heterodimeric transmembrane receptors composed of eighteen a subunits and eight b subunits that can be non-covalently assembled into 24 combinations. The integrin dimers bind to an array of different ECM molecules with overlapping binding affinities, as summarised in a review by Alam et al. (2007). Therefore, the specific integrin expression patterns by a cell dictate which ECM substrate the cell can bind (Hemler & Lobb 1995) and the composition of integrin adhesomes determines the downstream signalling events, thus the eventual cell behaviour and fate. Integrins have unique ability to respond

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