Remodelling the extracellular matrix in development and disease

Abstract

The extracellular matrix (ECM) is a highly dynamic structure that is present in all tissues and continuously undergoes controlled remodelling. This process involves quantitative and qualitative changes in the ECM, mediated by specific enzymes that are responsible for ECM degradation, such as metalloproteinases. The ECM interacts with cells to regulate diverse functions, including proliferation, migration and differentiation. ECM remodelling is crucial for regulating the morphogenesis of the intestine and lungs, as well as of the mammary and submandibular glands. Dysregulation of ECM composition, structure, stiffness and abundance contributes to several pathological conditions, such as fibrosis and invasive cancer. A better understanding of how the ECM regulates organ structure and function and of how ECM remodelling affects disease progression will contribute to the development of new therapeutics.

Figure 1. Structure and targets substrates of metalloproteinases

Metalloproteinases belong to the metzincin enzyme family, which includes matrix metalloproteinases(MMPs), adamalysins (which includes ADAMs (a disintegrin and metalloproteinases) and ADAMTS (ADAMs with a thrombospondin motif)) and astacins (including meprins). They are multidomain enzymes that contain the highly conserved motif HEXXHXXGXXH (where X is any amino acid), in which three His residues chelate a zinc ion in the catalytic site. Metalloproteinases are produced either as soluble or membrane-anchored enzymes that cleave components of the extracellular matrix (ECM). MMPs are composed of several shared functional domains: signal peptide domain, propeptide domain, catalytic domain and haemopexin-like domain (except MMP7, MMP23 and MMP26). The amino-terminal signal peptide domain is required for the secretion of MMPs. The propeptide domain contains the Cys-switch motif PRCGXPD. The catalytic domain (which has proteolytic activity) contains the zinc-binding motif; the Cys residue in this motif interacts with the zinc ion that keeps pro-MMPs inactive until the propeptide domain is removed. The carboxy-terminal haemopexin-like domain, which is present in almost all MMPs, is involved in substrate specificity and in the non-proteolytic functions of MMPs. Membrane-type MMPs (MTMMPs) such as MMP14 are anchored to the cell surface by either a transmembrane domain followed by a short cytoplasmic tail or a glycosylphosphatidylinositol (GPI) sequence. Some MMPs, including MTMMPs, MMP11, MMP17, MMP21, MMP23, MMP25 and MMP28 can be activated by the furin convertase, which cleaves the propeptide of inactive precursors in the Golgi apparatus, to release functional proteins. ADAMs are transmembrane proteins that are structurally similar to MTMMPs, except that they lack the haemopexin domain and instead have three other domains: the Cys-rich domain, the epidermal growth factor (EGF)-like repeat domain (except ADAM10 and ADAM17) and the disintegrin domain. Only ADAM9, ADAM10, ADAM12 and ADAM15 are shown, as the other ADAMs do not have known ECM protein substrates. ADAMTSs are secreted proteinases and have thrombospondin type I-like repeats in their C-terminal sequence. In addition to the metalloproteinase domains, the meprins also have an astacin-like catalytic domain (Ast-like Cat), a MAM (meprin A5 protein Tyr phosphatase) domain, a TRAF (TNFR-associated factor) domain and a C-terminal cytosolic tail. Meprin-α also contains a furin cleavage domain, cleavage of which results in the loss of the EGF-like transmembrane domain and the cytosolic domain and release of the enzyme into the extracellular space. MMP23 contains an immunoglobulin (Ig) domain that is unique among the MMPs. This Ig domain facilitates protein–protein or lipid–protein interactions similar to the haemopexin domain of other MMPs.

Figure 2. Extracellular matrix remodelling during intestinal development

a | Tadpole-to-adult intestinal epithelium remodelling during Xenopus laevis morphogenesis. In the pre-metamorphosis tadpole, the small intestine consists of a single layer of larval epithelium (also known as typhlosole), connective tissue and a thin muscle layer. During metamorphosis, thyroid hormone (TH) is produced in high levels, inducing the release of matrix metalloproteinase 11 (MMP11) by stromal cells to trigger apoptosis of larval epithelial cells. At the same time, proliferating stem and progenitor cells give rise to new adult epithelial cells that replace the larval epithelium. During metamorphosis, the basement membrane and the muscle layer are thicker. The levels of other MMPs, such as MMP2 and/or MMP9 and MMP14, increase during tadpole metamorphosis after epithelial cell death, suggesting that they may have a role post-apoptosis. At the end of metamorphosis, the differentiated adult intestine becomes capable of self-renewal and forms a multiply folded epithelium, similar to the mammalian adult intestine. b–d | Intestinal epithelium remodelling in other vertebrates. Laminin distribution during mammalian intestine development determines small intestine and colon architecture. The basement membrane of villi in the intestine is composed mainly of laminin 511 α5 chain. In mice, a lack of laminin 511 in the intestinal basement membrane leads to a compensatory deposition of colonic laminins (laminin 111 and laminin 411), which results in the transformation of the small intestinal to a tissue with a colon-like mucosal structure that shows high levels of cell proliferation, low levels of the cell cycle inhibitor cyclin-dependent kinase inhibitor B (CDKN1B), and higher numbers of goblet cells (part b). RGD-dependent substrates such as fibronectin can bind to α8β1 integrin. This anchorage prevents anoikis in undifferentiated human intestinal epithelial crypt (HIEC) cells through the recruitment of vinculin and the activation of the PI3K–AKT signalling pathway (part c). Collagen VI is produced by HIEC cells and regulates fibronectin assembly by restraining cell–fibronectin interactions, which influences cell functions such as migration. A lack of collagen VI leads to recruitment of tensin at the fibrillar adhesion points via the activation of myosin light chain kinases (MLCKs), which mediate actomyosin contractility, extensive fibrillogenesis and cell migration (part d). FAK, focal adhesion kinase. Figure part a modified from Establishment of intestinal stem cell niche during amphibian metamorphosis. Curr. Top. Dev. Biol Volume 103. Chapter 11. Pages 305–327. 2013. With permission from Elsevier.

Figure 3. Extracellular matrix remodelling during branching morphogenesis

A| Ductal elongation and branching of the mammary and submandibular glands. Aa | Collagen is locally synthesized and aligned to increase extracellular matrix (ECM) stiffness and create a mechanical anisotropy that will drive branching. Collagen synthesis is mediated by the activation of the RHOA–RHO-associated protein kinase (ROCK) signalling pathway. The ECM at the end bud tip is much thinner than in cleft region and around the duct. Matrix metalloproteinase 2 (MMP2) and MMP14 are expressed and active at the end bud tip, whereas MMP3 is involved inside branching. Ab | Cleft formation and deepening in the submandibular glands. Fibronectin is locally assembled in the basement membrane and induces BTBD7 at the base of forming clefts, which in turn upregulates the transcription factor SNAIL2 and downregulates the adhesion molecule E-cadherin. These molecular events promote alterations in cell shape, decreasing cell–cell adhesion and promoting a motile phenotype to promote cleft progression. Fibronectin assembly requires focal adhesion kinase (FAK) activation and RHOA–ROCK-mediated actomyosin contraction. B | Role of elastin and collagen deposition in alveolar branching in the lung. Elastin and collagen deposition promotes ECM stiffness in the neonatal lung and facilitates signalling through the endothelial lipoprotein receptor-related protein 5 (LRP5)–TIE2 (also known as angiopoietin 1 receptor) pathway, which is required for normal lung development. Consistent with this, disrupting lung collagen I, III and VI and elastin expression and localization through treatment with β-aminopropionitrile, an inhibitor of the collagen crosslinking enzyme lysyl oxidase (LOX), softens neonatal mouse lung tissue and downregulates the expression of LRP5 and TIE2, which leads to an inhibition of vascular and alveolar morphogenesis in neonatal mice. Ca | Heparan sulphate proteoglycans (HSPGs) bind fibroblast growth factors (FGFs) with different affinity and help to create a concentration gradient that can control cell fate in submandibular glands. In contrast to FGF10, FGF7 binds HSPG with low affinity and diffuses broadly, promoting branching in the submandibular gland. Cb| HSPGs such as perlecan bind FGF10 with high affinity. Following cleavage by heparanase, perlecan releases FGF10, which can then diffuse locally and promote duct elongation. FGFR2, FGF receptor 2; MLC, myocin light chain; MYPT, myosin phosphatase. Figure part Ca modified from Differential interactions of FGFs with heparin sulfate control gradient formation and branching morphogenesis. Sci. Signal. Volume 2. Issue 88. Page ra55. 2009. Reprinted with permission from AAAS.

Figure 4. Aberrant extracellular matrix remodelling leads to numerous human diseases

A | Over-degradation of the extracellular matrix (ECM), mediated primarily by matrix metalloproteinases (MMPs) and ADAMTS (ADAMs with a thrombospondin motif), results in osteoarthritis and increased breakdown of the connective tissue. B | As a result of chronic inflammation or tissue injury, transforming growth factor-β (TGFβ), connective tissue growth factor (CTGF), interleukin-13 (IL-13) and other factors stimulate fibroblasts and myofibroblasts (the main ECM producers) to produce more ECM, resulting in pathological fibrosis. The excess ECM further stimulates fibroblasts to continue making ECM, forming a positive feedback loop. Fibrosis is a major risk factor for developing cancer, including hepatocellular carcinoma and breast cancer. C | The ECM contributes to cancer pathogenesis by several mechanisms: functioning as a barrier to chemotherapy, to monoclonal antibodies such as cetuximab and to immune therapy mediated, for example, by cytotoxic T cells (CTLs) (part Ca); forming migration track ‘highways’ that regulate the interaction of immune cells with cancer cells (part Cb); stimulating integrin signalling through increased ECM stiffness, which promotes ECM synthesis, invasion and proliferation (lysyl oxidase (LOX) and LOX-like 2 (LOXL2) are enzymes that crosslink collagen and are the main enzymes responsible for increasing ECM stiffness) (part Cc); forming a niche for new metastatic cells and providing survival and proliferative signals (part Cd); generating novel bioactive ECM fragments from native ECM chains, and stimulating cell migration or immune cell recruitment (part Ce); activating cell–ECM receptors such as discoidin domain-containing receptor 1 (DDR1) and DDR2, which bind directly to collagen and regulate transcriptional pathways to increase MMP and epithelial–mesenchymal transition (EMT) marker expression (part Cf); and sequestering growth factors that can be released by proteolytic cleavage, which then diffuse to bind receptors to stimulate cell growth, EMT or angiogenesis (part Cg).