Role of the extracellular matrix in morphogenesis

The extracellular matrix is a complex, dynamic and critical component of all tissues. It functions as a scaffold for tissue morphogenesis, provides cues for cell proliferation and differentiation, promotes the maintenance of differentiated tissues and enhances the repair response after injury. Various amounts and types of collagens, adhesion molecules, proteoglycans, growth factors and cytokines or chemokines are present in the tissue- and temporal-specific extracellular matrices. Tissue morphogenesis is mediated by multiple extracellular matrix components and by multiple active sites on some of these components. Biologically active extracellular matrix components may have use in tissue repair, regeneration and engineering, and in programming stem cells for tissue replacement.     

Structure And Evolution Of Calcium-Modulated Protein

Abstract

The question posed by the science of analytical histology is how the properties and interactions of the components of the tissues determine their organization in the organs. The relevant components of the tissues are the cells and the extracellular matrix. The ability of cohering populations of cells to self-assemble structured tissues by cell sorting out offers an important opportunity for the experimental study of the mechanisms by which the cells and extracellular matrix interact to determine structure. The investigator can manipulate the initial organization and the cellular composition of the system and, in favorable situations, the composition of the extracellular matrix and the activities of candidate adhesive molecules.     

Syndecan, a developmentally regulated cell surface proteoglycan that binds extracellular matrix and growth factors

Cellular behaviour during development is dictated, in part, by the insoluble extracellular matrix and the soluble growth factor peptides, the major molecules responsible for integrating cells into morphologically and functionally defined groups. These extracellular molecules influence cellular behaviour by binding at the cell surface to specific receptors that transduce intracellular signals in various ways not yet fully clear. Syndecan, a cell surface proteoglycan found predominantly on epithelia in mature tissues binds both extracellular matrix components (fibronectin, collagens I, III, V, and thrombospondin) and basic fibroblast growth factor (bFGF). Syndecan consists of chondroitin sulfate and heparan sulphate chains linked to a 31 kilodalton (kDa) integral membrane protein. Syndecan represents a family of integral membrane proteoglycans that differ in extracellular domains, but share cytoplasmic domains. Syndecan behaves as a matrix receptor: it binds selectively to components of the extracellular matrix, associates intracellularly with the actin cytoskeleton when cross-linked at the cell surface, its extracellular domain is shed upon cell rounding and it localizes solely to basolateral surfaces of simple epithelia. Mammary epithelial cells made syndecan-deficient become fibroblastic in morphology and cell behaviour, showing that syndecan maintains epithelial cell morphology. Syndecan changes in quantity, location and structure during development: it appears initially on four-cell embryos (prior to its known matrix ligands), becomes restricted in the pre-implementation embryo to the cells that will form the embryo proper, changes its expression due to epithelial-mesenchymal interactions (for example, induced in kidney mesenchyme by the ureteric bud), and with association of cells with extracellular matrix (for example, during B-cell differentiation), and ultimately, in mature tissues becomes restricted to epithelial tissues. The number and size of its glycosaminoglycan chains vary with changes in cell shape and organization yielding tissue type-specific polymorphic forms of syndecan. Its interactions with the major extracellular effector molecules that influence cell behaviour, its role in maintaining cell shape and its spatial and temporal changes in expression during development indicate that syndecan is involved in morphogenesis.     

Matrix biology meets toxinology

Venoms are cocktails containing pharmacologically active compounds, which drastically affect essential functions of the neuromuscular and cardiovascular system, as well as of blood, kidney and other organs. As the extracellular matrix and its contacts with cells are responsible for maintaining the integrity and functionality of these organs and tissues, it is not surprising that several venom components target matrix molecules and their respective cellular receptors. Many venom components, such as matrix-degrading enzymes, disintegrins, and C-type lectin-like proteins, have been identified and have laid the foundation for the frontier research field of matrix toxinology. Interestingly, many toxins consist of domains which are structurally homologous to modules and domains of matrix proteins, their proteinases and cellular receptors. In addition to finding new agents and tools, which specifically interact with matrix molecules and their receptors, the characterization of known matrix-targeting toxins will provide insights into their molecular modes of action and thus may lead to potential new therapeutic strategies for treating matrix-related diseases, such as blood clotting and thrombocyte-mediated disorders, but also tumor malignancies.     

Herpes simplex virus type 1 preferentially targets human colon carcinoma: role of extracellular matrix

Viral therapy of cancer (viral oncolysis) is dependent on selective destruction of the tumor tissue compared with healthy tissues. Several factors, including receptor expression, extracellular components, and intracellular mechanisms, may influence viral oncolysis. In the present work, we studied the potential oncolytic activity of herpes simplex virus type 1 (HSV-1), using an organ culture system derived from colon carcinoma and healthy colon tissues of mouse and human origin. HSV-1 infected normal colons ex vivo at a very low efficiency, in contrast to high-efficiency infection of colon carcinoma tissue. In contrast, adenoviral and lentiviral vectors infected both tissues equally well. To investigate the mechanisms underlying the preferential affinity of HSV-1 for the carcinoma tissue, intracellular and extracellular factors were investigated. Two extracellular components, collagen and mucin molecules, were found to restrict HSV-1 infectivity in the healthy colon. The mucin layer of the healthy colon binds to HSV-1 and thereby blocks viral interaction with the epithelial cells of the tissue. In contrast, colon carcinomas express small amounts of collagen and mucin molecules and are thus permissive to HSV-1 infection. In agreement with the ex vivo system, HSV-1 injected into a mouse colon carcinoma in vivo significantly reduced the volume of the tumor. In conclusion, we describe a novel mechanism of viral selectivity for malignant tissues that is based on variance of the extracellular matrix between tumor and healthy tissues. These insights may facilitate new approaches to the application of HSV-1 as an oncolytic virus.     

Epithelial-mesenchymal interactions in tooth morphogenesis: the roles of extracellular matrix, growth factors, and cell surface receptors.

Morphogenesis and cell differentiation in the developing tooth are controlled by a series of reciprocal interactions between the epithelial and mesenchymal tissues. The exact molecular mechanisms operating in these interactions are unknown at present, but both structural components of the extracellular matrix (ECM) and diffusible growth factors have been suggested to be involved. In this review article we summarize our findings on the distribution patterns of three ECM molecules and two cell surface receptors during tooth morphogenesis through bud, cap, and bell stages of development. The examined molecules include fibronectin, type III collagen, and tenascin, which all represent components of the mesenchymal ECM, the cell surface proteoglycan, syndecan, which functions as a receptor for interstitial matrix, and the cell surface receptor for epidermal growth factor. Based on the observed changes in distribution patterns and on experimental evidence, roles are suggested for these molecules in epithelial-mesenchymal interactions during tooth development. Fibronectin is suggested to be involved in the cell-matrix interaction that controls odontoblast differentiation. Epidermal growth factor and its receptors are suggested to be involved in a paracrine fashion in the epithelial-mesenchymal interactions regulating morphogenesis of bud- and cap-stage teeth. Tenascin and syndecan are accumulated in the dental mesenchyme during the bud stage of development, and it is suggested that they represent a couple of a cell surface receptor and its matrix ligand and that they are involved in mesenchymal cell condensation during the earliest stages of tooth morphogenesis.     

Extracellular Matrix in Neural Plasticity and Regeneration

The extracellular matrix (ECM) is a fundamental component of biological tissues. The ECM in the central nervous system (CNS) is unique in both composition and function. Functions such as learning, memory, synaptogenesis, and plasticity are regulated by numerous ECM molecules. The neural ECM acts as a non-specific physical barrier that modulates neuronal plasticity and axon regeneration. There are two specialized types of ECM in the CNS, diffuse perisynaptic ECM and condensed ECM, which selectively surround the perikaryon and initial part of dendritic trees in subtypes of neurons, forming perineuronal nets. This review presents the current knowledge about the role of important neuronal ECM molecules in maintaining the basic functions of a neuron, including electrogenesis and the ability to form neural circuits. The review mainly focuses on the role of ECM components that participate in the control of key events such as cell survival, axonal growth, and synaptic remodeling. Particular attention is drawn to the numerous molecular partners of the main ECM components. These regulatory molecules are integrated into the cell membrane or disposed into the matrix itself in solid or soluble form. The interaction of the main matrix components with molecular partners seems essential in molecular mechanisms controlling neuronal functions. Special attention is paid to the chondroitin sulfate proteoglycan 4, type 1 transmembrane protein, neural-glial antigen 2 (NG2/CSPG4), whose cleaved extracellular domain is such a molecular partner that it not only acts directly on neural and vascular cells, but also exerts its influence indirectly by binding to resident ECM molecules.

     

Bacterial adhesion to animal tissues: protein determinants for recognition of extracellular matrix components

The extracellular matrix (ECM) is present within all animal tissues and organs. Actually, it surrounds the eukaryotic cells composing the four basic tissue types, i.e. epithelial, muscle, nerve and connective. ECM does not solely refer to connective tissue but composes all tissues where its composition, structure and organization vary from one tissue to another. Constituted of the four main fibrous proteins, i.e. collagen, fibronectin, laminin and elastin, ECM components form a highly structured and functional network via specific interactions. From the basement membrane to interstitial matrix, further heterogeneity exists in the organization of the ECM in various tissues and organs also depending on their physiological state. Back to a molecular level, bacterial proteins represent the most significant part of the microbial surface components recognizing adhesive matrix molecules (MSCRAMM). These cell surface proteins are secreted and localized differently in monoderm and diderm–LPS bacteria. While one collagen‐binding domain (CBD) and different fibronectin‐binding domains (FBD1 to 8) have been registered in databases, much remains to be learned on specific binding to other ECM proteins via single or supramolecular protein structures. Besides theinteraction of bacterial proteins with individual ECM components, this review aims at stressing the importance of fully considering the ECM at supramolecular, cellular, tissue and organ levels. This conceptual view should not be overlooked to rigorously comprehend the physiology of bacterial interaction from commensal to pathogenic species.     

Current viewpoint on structure and on evolution of collagens. II. Fibril-associated collagens

Fibril-associated collagens (FACITs) form one of subfamilies included in family of collagens. Being minor components of connective tissue of multicellular animals, FACITs play an important role in structurization of extracellular matrix whose peculiarities determine essential intertissue differences. FACITs participate in regulation of sizes of banded collagen fibrils as well as are connecting links between various components extracellular matrix and cells in different tissues. Functional characteristics of FACIT molecules are determined by peculiarities of structural organization of their α-chains (breakdowns in collagenous domains and module structure of N-terminal noncollagenous sites), trimeric molecules (domains of trimerization) and supramolecular assemblies (mainly association with banded collagen fibrils and the inability to form homopolymeric supramolecular aggregates). The problem of evolution of this group of collagen molecules is also discussed. A hypothetical model of structural changes leading to formation of the FACIT subfamily is proposed.     

Interactions between the extracellular matrix and the cell surface determine tooth morphogenesis and the cellular differentiation of the dental mesenchyme

A series of reciprocal interactions between epithelial and mesenchymal tissues control the morphogenesis and cell differentiation in the developing tooth. The molecular mechanisms operating in these interactions are, however, unknown at present. Structural components of the extracellular matrix (ECM) affect cellular behavior in the embryo and appear to be involved also in these regulatory processes. The ECM molecules exert their effects on cells through binding to specific matrix receptors on the cell surface. This review article summarizes our findings on the distribution patterns during tooth development of the ECM glycoproteins, fibronectin and tenascin, and of the cell surface proteoglycan, syndecan, which functions as a receptor for interstitial matrix. Based on the observed changes in these distribution patterns and on experimental evidence, roles for these molecules in epithelial-mesenchymal interactions during tooth development are suggested. Fibronectin and tenascin are enriched in the dental basement membrane at the time of odontoblast differentiation. These matrix glycoproteins may be involved in the cell-matrix interaction which controls differentiation of the dental mesenchymal cells into odontoblasts. Tenascin and syndecan are accumulated in the dental mesenchyme during bud stage of development. We have shown in tissue recombination experiments that the presumptive dental epithelium induces the expression of tenascin and syndecan in mesenchyme. We suggest that these molecules are involved in cell-matrix interactions, which regulate mesenchymal cell condensation during the earliest stages of tooth morphogenesis.     

Evidence of low molecular weight components in the organic matrix of the reef building coral, Stylophora pistillata

Biominerals contain both inorganic and organic components. Organic components are collectively termed the organic matrix, and this matrix has been reported to play a crucial role in mineralization. Several matrix proteins have been characterized in vertebrates, but only a few in invertebrates, primarily in Molluscs and Echinoderms. Methods classically used to extract organic matrix proteins eliminate potential low molecular weight matrix components, since cut-offs ranging from 3.5 to 10 kDa are used to desalt matrix extracts. Consequently, the presence of such components remains unknown and these are never subjected to further analyses. In the present study, we have used microcolonies from the Scleractinian coral Stylophora pistillata to study newly synthesized matrix components by labelling them with 14C-labelled amino acids. Radioactive matrix components were investigated by a method in which both total organic matrix and fractions of matrix below and above 5 kDa were analyzed. Using this method and SDS-PAGE analyses, we were able to detect the presence of low molecular mass matrix components (<3.5 kDa), but no free amino acids in the skeletal organic matrix. Since more than 98% of the 14C-labelled amino acids were incorporated into low molecular weight molecules, these probably form the bulk of newly synthesized organic matrix components. Our results suggest that these low molecular weight components may be peptides, which can be involved in the regulation of coral skeleton mineralization.     

Extracellular vesicles are integral and functional components of the extracellular matrix

Extracellular vesicles (EV) are small plasma membrane-derived particles released into the extracellular space by virtually all cell types. Recently, EV have received increased interest because of their capability to carry nucleic acids, proteins, lipids and signaling molecules and to transfer their cargo into the target cells. Less attention has been paid to their role in modifying the composition of the extracellular matrix (ECM), either directly or indirectly via regulating the ability of target cells to synthesize or degrade matrix molecules. Based on recent results, EV can be considered one of the structural and functional components of the ECM that participate in matrix organization, regulation of cells within it, and in determining the physical properties of soft connective tissues, bone, cartilage and dentin. This review addresses the relevance of EV as specific modulators of the ECM, such as during the assembly and disassembly of the molecular network, signaling through the ECM and formation of niches suitable for tissue regeneration, inflammation and tumor progression. Finally, we assess the potential of these aspects of EV biology to translational medicine.     

Epitope tagging of legume root nodule extensin modifies protein structure and crosslinking in cell walls of transformed tobacco leaves

Root nodule extensins (RNEs) are highly glycosylated plant glycoproteins localized in the extracellular matrix of legume tissues and in the lumen of Rhizobium-induced infection threads. In pea and other legumes, a family of genes encode glycoproteins of different overall length but with the same basic composition. The predicted polypeptide sequence reveals repeating and alternating motifs characteristic of extensins and arabinogalactan proteins. In order to monitor the behavior of individual RNE gene products in the plant extracellular matrix, the coding sequence of PsRNE1 from Pisum sativum was expressed in insect cells and in tobacco leaves. RNE products extracted from tobacco tissues were of high molecular weight (in excess of 80 kDa), indicating extensive glycosylation similar to that in pea tissues. Epitope-tagged derivatives of PsRNE1 could be localized in cell walls. However, the introduction of epitope tags at the C-terminus of RNE altered the behavior of RNE in the extracellular matrix, apparently preventing intermolecular crosslinking of RNE molecules and their covalent association with other cell wall components. These observations are discussed in the light of a computational model for the RNE glycoprotein that is consistent with an extended rod-like structure. It is proposed that RNE can undergo three classes of tyrosine-based crosslinking. Intramolecular crosslinking of vicinal Tyr residues is rod stiffening, end-to-end linkage is rod lengthening, and side-to-side intermolecular crosslinking is rod bundling. The control of these interconversions could have important implications for the biomechanics of infection thread growth.     

Ontogeny of the basal lamina in the sea urchin embryo

The patterns of expression for several extracellular matrix components during development of the sea urchin embryo are described. An immunofluorescence assay was employed on paraffin-sectioned material using (i) polyclonal antibodies against known vertebrate extracellular matrix components: laminin, fibronectin, heparan sulfate proteoglycan, collagen types I, III, and IV; and (ii) monoclonal antibodies generated against sea urchin embryonic components. Most extracellular matrix components studied were found localized within the unfertilized egg in granules (0.5-2.0 micron) distinct from the cortical granules. Fertilization initiated trafficking of the extracellular matrix (ECM) components from within the egg granules to the basal lamina of the developing embryo. The various ECM components arrived within the developing basal lamina at different times, and not all components were unique to the basal lamina. Two ECM components were not found within the egg. These molecules appeared de novo at the mesenchyme blastula stage, and remained specific to the mesoderm through development. The reactivity of antibodies to vertebrate ECM antigens with components of the sea urchin embryo suggests the presence of immunologically similar ECM molecules between the phyla.     

Revisiting the mystery of fibronectin multimers: The fibronectin matrix is composed of fibronectin dimers cross-linked by non-covalent bonds

Fibronectin (FN) matrix fibrils have long been thought to be formed by disulfide-bonded FN multimers, although there is no direct evidence that they are covalently linked with each other. To understand the biochemical properties of these fibrils, we extracted a crude FN matrix from FN-YPet transfected 3T3 cell culture using 0.2% deoxycholate and DNase. The insoluble extracted matrix preserved fibrillar structures and a major portion of the extracted proteins migrated as FN monomers on an SDS gel under reducing conditions. Under non-reducing conditions, some FN molecules appeared to be trapped at the top of the stacking gel. We tested this by mixing fluorescently labeled FN dimers with the extracted matrix just before loading on an SDS gel, and found that most of them were trapped with the extracted proteins at the top of the stacking gel. These results suggested that some components of the extracted matrix plugged the stacking gel and FN dimers were trapped with them. Rotary shadowing electron microscopy showed that the extracted matrix had some fibers that resembled fibrillin microfibrils. Peptide mass fingerprinting confirmed the presence of fibrillin in the extracted matrix. Fibrillin is known to form disulfide-bonded multimers and it is likely to be one of the components that plug the stacking gel and trap FN molecules in this system. The phenomenon by which FN molecules appear to migrate as multimers on SDS gels is thus an artifact rising from the presence of other large components in the extract. We conclude that FN matrix fibrils are made of FN dimers that are further cross-linked by non-covalent protein–protein bonds.     

Spatially resolved analysis of small molecules by matrix-assisted laser desorption/ionization mass spectrometric imaging (MALDI-MSI)

? Matrix-assisted laser desorption/ionization mass spectrometric imaging (MALDI-MSI) of tissues provides the means to analyse the spatial distributions of small molecules and proteins within tissues. This imaging technique is commonplace in medicinal and pharmaceutical research, but its application in plant science is very recent. Broader introduction requires specific adaptations for plant tissues. Sample preparation is of paramount importance in order to obtain high-quality spectra providing sufficient spatial resolution for compounds. Optimization is required for sectioning, choice of matrix and means of matrix deposition. ? Here, we present our current protocols for the detection of small molecules in cryodissected immature barley (Hordeum vulgare) grains and tobacco (Nicotiana tabacum) roots. ? Examples of MALDI-MSI measurements are provided, and the level of reproducibility across biological replicates is addressed. Furthermore, our approaches for the validation of distribution patterns and for the identification of molecules are described. ? Finally, we discuss how MALDI-MSI can contribute to applied plant research.     

The biochemical, biological, and pathological kaleidoscope of cell surface substrates processed by matrix metalloproteinases

Matrix metalloproteinases (MMPs) constitute a family of more than 20 endopeptidases. Identification of specific matrix and non-matrix components as MMP substrates showed that, aside from their initial role as extracellular matrix modifiers, MMPs play significant roles in highly complex processes such as the regulation of cell behavior, cell-cell communication, and tumor progression. Thanks to the comprehensive examination of the expanded MMP action radius, the initial view of proteases acting in the soluble phase has evolved into a kaleidoscope of proteolytic reactions connected to the cell surface. Important classes of cell surface molecules include adhesion molecules, mediators of apoptosis, receptors, chemokines, cytokines, growth factors, proteases, intercellular junction proteins, and structural molecules. Proteolysis of cell surface proteins by MMPs may have extremely diverse biological implications, ranging from maturation and activation, to inactivation or degradation of substrates. In this way, modification of membrane-associated proteins by MMPs is crucial for communication between cells and the extracellular milieu, and determines cell fate and the integrity of tissues. Hence, insights into the processing of cell surface proteins by MMPs and the concomitant effects on physiological processes as well as on disease onset and evolution, leads the way to innovative therapeutic approaches for cancer, as well as degenerative and inflammatory diseases.     

The Biochemical,Biological, and Pathological Kaleidoscope of Cell Surface Substrates Processed by Matrix Metalloproteinases

ABSTRACT

Matrix metalloproteinases (MMPs) constitute a family of more than 20 endopeptidases. Identification of specific matrix and non-matrix components as MMP substrates showed that, aside from their initial role as extracellular matrix modifiers, MMPs play significant roles in highly complex processes such as the regulation of cell behavior, cell-cell communication, and tumor progression. Thanks to the comprehensive examination of the expanded MMP action radius, the initial view of proteases acting in the soluble phase has evolved into a kaleidoscope of proteolytic reactions connected to the cell surface. Important classes of cell surface molecules include adhesion molecules, mediators of apoptosis, receptors, chemokines, cytokines, growth factors, proteases, intercellular junction proteins, and structural molecules. Proteolysis of cell surface proteins by MMPs may have extremely diverse biological implications, ranging from maturation and activation, to inactivation or degradation of substrates. In this way, modification of membrane-associated proteins by MMPs is crucial for communication between cells and the extracellular milieu, and determines cell fate and the integrity of tissues. Hence, insights into the processing of cell surface proteins by MMPs and the concomitant effects on physiological processes as well as on disease onset and evolution, leads the way to innovative therapeutic approaches for cancer, as well as degenerative and inflammatory diseases.     

Adhesion determinants of the Streptococcus species

Streptococci are clinically important Gram-positive bacteria that are capable to cause a wide variety of diseases in humans and animals. Phylogenetic analyses based on 16S rRNA sequences of the streptococcal species reveal a clustering pattern, reflecting, with a few exceptions, their pathogenic potential and ecological preferences. Microbial adhesion to host tissues is the initial critical event in the pathogenesis of most infections. Streptococci use multiple adhesins to attach to the epithelium, and their expression is regulated in response to environmental and growth conditions. Bacterial adhesins recognize and bind cell surface molecules and extracellular matrix components through specific domains that for certain adhesin families have been well defined and found conserved across the streptococcal species. In this review, we present the different streptococcal adhesin families categorized on the basis of their adhesive properties and structural characteristics, and, when available, we focus the attention on conserved functional domains.