Approaches in extracellular matrix engineering for determination of adhesion molecule mediated single cell function

The native extracellular matrix (ECM) and the cells that comprise human tissues are together engaged in a complex relationship; cells alter the composition and structure of the ECM to regulate the material and biologic properties of the surrounding environment while the composition and structure of the ECM modulates cellular processes that maintain healthy tissue and repair diseased tissue. This reciprocal relationship occurs via cell adhesion molecules (CAMs) such as integrins, selectins, cadherins and IgSF adhesion molecules. To study these cell-ECM interactions, researchers use two-dimensional substrates or three-dimensional matrices composed of native proteins or bioactive peptide sequences to study single cell function. While two-dimensional substrates provide valuable information about cell-ECM interactions, three-dimensional matrices more closely mimic the native ECM; cells cultured in three-dimensional matrices have demonstrated greater cell movement and increased integrin expression when compared to cells cultured on two-dimensional substrates. In this article we review a number of cellular processes (adhesion, motility, phagocytosis, differentiation and survival) and examine the cell adhesion molecules and ECM proteins (or bioactive peptide sequences) that mediate cell functionality.     

Modulation of epithelial morphogenesis and cell fate by cell-to-cell signals and regulated cell adhesion

Cell-cell and cell-extracellular matrix (ECM) interactions control many developmental decisions of epithelial cell fate and morphogenesis. Protein tyrosine kinases are one class of regulatory molecules that have been implicated in the modulation of these processes. Several protein tyrosine kinases co-localize with cell-cell (cadherin) and cell-ECM (integrin) adhesion molecules at specific adhesion domains of epithelial cells. Protein tyrosine kinases may regulate epithelial development by modulating cell-cell and cell-ECM interactions and by relaying signals initiated by these interactions to other cellular components that determine cell structure and function.     

Cell-Extracellular Matrix Interactions in Normal and Diseased Skin

Mammalian skin comprises a multi-layered epithelium, the epidermis, and an underlying connective tissue, the dermis. The epidermal extracellular matrix is a basement membrane, whereas the dermal ECM comprises fibrillar collagens and associated proteins. There is considerable heterogeneity in ECM composition within both epidermis and dermis. The functional significance of this extends beyond cell adhesion to a range of cell autonomous and nonautonomous processes, including control of epidermal stem cell fate. In skin, cell-ECM interactions influence normal homeostasis, aging, wound healing, and disease. Disturbed integrin and ECM signaling contributes to both tumor formation and fibrosis. Strategies for manipulating cell-ECM interactions to repair skin defects and intervene in a variety of skin diseases hold promise for the future.The focus of this review is the role of cell-ECM interactions in the physiology of normal and diseased mammalian skin. The skin has epithelial and mesenchymal components and contains ECM comprising both fibrillar collagen and basement membrane. Experimentally, it is a highly tractable tissue, and a range of in vitro and in vivo approaches are available to explore cell-ECM interactions. Such studies are of medical importance because of the wide variety of benign and malignant skin diseases. Research on skin therefore provides an integrated, in vivo, context for understanding the functional significance of specific molecular interactions and signaling pathways involved in cell-ECM adhesion.     

Integrin receptors: the dynamic modulators of endometrial function

Cell-cell and cell-extracellular matrix (ECM) interactions play a critical role in various developmental processes, including differentiation, proliferation and migration of cells. ECM proteins can influence cellular function thus creating a complex feedback mechanism. The adhesion of cells to each other, their ECM proteins and endothelial surfaces is mediated by a variety of membrane proteins collectively known as adhesion molecules. Adhesion molecules have been further divided into five subfamilies, the integrins, the selectins, the cadherins, the mucins and the immunoglobulin superfamily. Members of the integrin family of cell surface adhesion receptors are important mediators of cell-ECM contact. Integrin receptors are alpha beta heterodimers with a transmembrane segment, a short cytoplasmic domain and a large extracellular domain. The role of integrins in reproduction has been established. Several reasons make these molecules very attractive due to their constant involvement from egg to birth. They participate in sperm-egg interaction, fertilization, implantation and placentation in many species including humans. Integrins provide signals to individual cells essential for growth and development of different tissues. In the present review, we describe (1) the regulatory pathways for controlling expression of integrins in the endometrium, (2) various biomarkers and their role in endometrial function, (3) reproductive disorders in women related to aberrant integrin expression in the endometrium and (4) the functional significance of integrins available from gene knockout studies.     

Focal Adhesion: A Focal Point in Current Cell Biology and Molecular Medicine

Cell-extracellular matrix (ECM) is an important property of virtually all cells in multi-cellular organisms. Cell-ECM adhesion research, therefore, has broad impact on biology and medicine. Studies over the past three decades have resulted in tremendous advance in our understanding of the molecular basis and functions of cell-ECM adhesion. Here, I focus on some of the general lessons that we have learned from recent studies on cell-ECM adhesion. In addition, I highlight several topics in this rapidly advancing research area. These topics, which include assembly, disassembly and regulation of cell-ECM adhesion structures, the molecular mechanisms of bi-directional signaling through cell-ECM adhesions, and the tissue and organ pathobiology of cell-ECM adhesion, are pertinent to our understanding of cell-ECM adhesion and signaling.Key Words: Focal adhesion, integrins, extracellular matrix, cytoskeleton, cell migrationCell-ECM adhesion is a fundamental process through which cells interact and communicate with the environment. Cell-ECM adhesion is essential for organogenesis during embryonic development. In adult, it is vital for maintenance of tissue integrity and organ functions. Alterations of cell-ECM adhesion hence are frequently associated with human diseases. Because of the broad significance of cell-ECM adhesion in biology and pathology, understanding how cell-ECM adhesion is mediated and regulated and determining how cell-ECM adhesion influences cell behavior have been the subjects of numerous studies. In particular, studies over the past three decades have led to major breakthroughs in our understanding of cell-ECM adhesion. Many of the key discoveries, including identification of integrins as major transmembrane receptors for ECM proteins, demonstration of integrins as bi-directional (outside-in and inside-out) transmembrane signaling machines, identification of talin, focal adhesion kinase (FAK), integrin-linked kinase (ILK) and other cytoplasmic and membrane-associated proteins as key regulators and effectors of integrins, and delineation of multiple downstream signaling pathways that relay signals from cell surface integrins to diverse cytoplasmic and nuclear effectors, have been reviewed in refs. ^1–12. In this brief article, I will focus on some of the general features of cell-ECM adhesion and discuss from my personal perspective several key questions that remain to be answered in future studies.     

Lateral spacing of integrin ligands influences cell spreading and focal adhesion assembly

Cell-extracellular matrix (cell-ECM) interactions mediated by integrin receptors are essential for providing positional and environmental information necessary for many cell functions, such as proliferation, differentiation and survival. In vitro studies on cell adhesion to randomly adsorbed molecules on substrates have been limited to sub-micrometer patches, thus preventing the detailed study of structural arrangement of integrins and their ligands. In this article, we illustrate the role of the distance between integrin ligands, namely the RGD (arginine-glycine-aspartate) sequence present in ECM proteins, in the control of cell adhesion. By using substrates, which carry cyclic RGD peptides arranged in highly defined nanopatterns, we investigated the dynamics of cell spreading and the molecular composition of adhesion sites in relation to a fixed spacing between the peptides on the surface. Our novel approach for in vitro studies on cell adhesion indicates that not only the composition, but also the spatial organization of the extracellular environment is important in regulating cell-ECM interactions.     

The extracellular matrix in development and morphogenesis: A dynamic view

The extracellular matrix (ECM) is synthesized and secreted by embryonic cells beginning at the earliest stages of development. Our understanding of ECM composition, structure and function has grown considerably in the last several decades and this knowledge has revealed that the extracellular microenvironment is critically important for cell growth, survival, differentiation and morphogenesis. ECM and the cellular receptors that interact with it mediate both physical linkages with the cytoskeleton and the bidirectional flow of information between the extracellular and intracellular compartments. This review considers the range of cell and tissue functions attributed to ECM molecules and summarizes recent findings specific to key developmental processes. The importance of ECM as a dynamic repository for growth factors is highlighted along with more recent studies implicating the 3-dimensional organization and physical properties of the ECM as it relates to cell signaling and the regulation of morphogenetic cell behaviors. Embryonic cell and tissue generated forces and mechanical signals arising from ECM adhesion represent emerging areas of interest in this field.     

Extracellular matrix networks in bone remodeling

Bones are constantly remodeled throughout life to maintain robust structure and function. Dysfunctional remodeling can result in pathological conditions such as osteoporosis (bone loss) or osteosclerosis (bone gain). Bone contains 100s of extracellular matrix (ECM) proteins and the ECM of the various bone tissue compartments plays essential roles directing the remodeling of bone through the coupled activity of osteoclasts (which resorb bone) and osteoblasts (which produce new bone). One important role for the ECM is to serve as a scaffold upon which mineral is deposited. This scaffold is primarily type I collagen, but other ECM components are involved in binding of mineral components. In addition to providing a mineral scaffolding role, the ECM components provide structural flexibility for a tissue that would otherwise be overly rigid. Although primarily secreted by osteoblast-lineage cells, the ECM regulates cells of both the osteoblast-lineage (such as progenitors, mature osteoblasts, and osteocytes) and osteoclast-lineage (including precursors and mature osteoclasts), and it also influences the cross-talk that occurs between these two oppositional cells. ECM influences the differentiation process of mesenchymal stem cells to become osteoblasts by both direct cell-ECM interactions as well as by modulating growth factor activity. Similarly, the ECM can influence the development of osteoclasts from undifferentiated macrophage precursor cells, and influence osteoclast function through direct osteoclast cell binding to matrix components. This comprehensive review will focus on how networks of ECM proteins function to regulate osteoclast- and osteoblast-mediated bone remodeling. The clinical significance of these networks on normal bone and as they relate to pathologies of bone mass and geometry will be considered. A better understanding of the dynamic role of ECM networks in regulating tissue function and cell behavior is essential for the development of new treatment approaches for bone loss.     

Fibroblast-Derived Extracellular Matrices: An Alternative Cell Culture System That Increases Metastatic Cellular Properties

Poor survival rates from lung cancer can largely be attributed to metastatic cells that invade and spread throughout the body. The tumor microenvironment (TME) is composed of multiple cell types, as well as non-cellular components. The TME plays a critical role in the development of metastatic cancers by providing migratory cues and changing the properties of the tumor cells. The Extracellular Matrix (ECM), a main component of the TME, has been shown to change composition during tumor progression, contributing to cancer cell invasion and survival away from the primary cancer site. Although the ECM is well-known to influence the fate of tumor progression, little is known about the molecular mechanisms that are affected by the cancer cell-ECM interactions. It is imperative that these mechanisms are elucidated in order to properly understand and prevent lung cancer dissemination. However, common in vitro studies do not incorporate these interactions into everyday cell culture assays. We have adopted a model that examines decellularized human fibroblast-derived ECM as a 3-dimensional substrate for growth of lung adenocarcinoma cell lines. Here, we have characterized the effect of fibroblast-derived matrices on the properties of various lung-derived epithelial cell lines, including cancerous and non-transformed cells. This work highlights the significance of the cell-ECM interaction and its requirement for incorporation into in vitro experiments. Implementation of a fibroblast-derived ECM as an in vitro technique will provide researchers with an important factor to manipulate to better recreate and study the TME.     

Dynamic imaging of cell, extracellular matrix, and tissue movements during avian vertebral axis patterning

Vertebrate axis patterning depends on cell and extracellular matrix (ECM) repositioning and proper cell-ECM interactions. However, there are few in vivo data addressing how large-scale tissue deformations are coordinated with the motion of local cell ensembles or the displacement of ECM constituents. Combining the methods of dynamic imaging and experimental biology allows both cell and ECM fate-mapping to be correlated with ongoing tissue deformations. These fate-mapping studies suggest that the axial ECM components "move" both as a composite meshwork and as autonomous particles, depending on the length scale being examined. Cells are also part of this composite, and subject to passive displacements resulting from tissue deformations. However, in contrast to the ECM, cells are self-propelled. The net result of cell and ECM displacements, along with proper ECM-cell adhesion, is the assembly of new tissue architecture. Data herein show that disruption of normal cell-ECM interactions during axis formation results in developmental abnormalities and a disorganization of the ECM. Our goal in characterizing the global displacement patterns of axial cells and ECM is to provide critical information regarding existing strain fields in the segmental plate and paraxial mesoderm. Deducing the mechanical influences on cell behavior is critical, if we are to understand vertebral axis patterning. Supplementary material for this article is available online at http://www.mrw.interscience.wiley.com/suppmat/1542-975X/suppmat/72/v72.266.html.     

Role of the extracellular matrix in lymphocyte migration

The extracellular matrix (ECM) exists in various biochemical and structural forms that can act either as a barrier to migrating leukocytes, in the case of basement membranes, or provide a physical scaffold supporting or guiding migration (interstitial matrix). This review focuses on basement membranes and our current knowledge of the way that leukocytes transmigrate this protein barrier, with emphasis on T lymphocytes. Recent data suggest that the classical concept of cell-matrix adhesion requires revision with respect to leukocyte-ECM interactions. Whereas specific receptors may be required for leukocyte recognition of ECM molecules or three-dimensional structural domains, the role of adhesion in migration as perceived from the traditional studies of adherent cell-ECM interactions is less clear. Further, the indirect effects of ECM such as the binding and presentation of cytokines or chemotactic factors may more profoundly influence the directed migration of normally non-adherent leukocytes than the migration of adherent cells such as epithelial cells or fibroblasts. Proteases (in particular matrix metalloproteinases) released at sites of inflammation can selectively process ECM, cell surface molecules or soluble factors, which may result in the release of bioactive fragments that can function as chemoattractants for different leukocyte subsets or may modulate the activity/function of resident mesenchymal and immune cells. Current findings suggest that different leukocyte types employ different mechanisms to migrate across or through the ECM; this might be determined by the composition and organization of the ECM itself.     

Mechanical signaling through the discoidin domain receptor 1 plays a central role in tissue fibrosis

ABSTRACT

The preservation of tissue and organ architecture and function depends on tightly regulated interactions of cells with the extracellular matrix (ECM). These interactions are maintained in a dynamic equilibrium that balances intracellular, myosin-generated tension with extracellular resistance conferred by the mechanical properties of the extracellular matrix. Disturbances of this equilibrium can lead to the development of fibrotic lesions that are associated with a wide repertoire of high prevalence diseases including obstructive cardiovascular diseases, muscular dystrophy and cancer. Mechanotransduction is the process by which mechanical cues are converted into biochemical signals. At the core of mechanotransduction are sensory systems, which are frequently located at sites of cell-ECM and cell-cell contacts. As integrins (cell-ECM junctions) and cadherins (cell-cell contacts) have been extensively studied, we focus here on the properties of the discoidin domain receptor 1 (DDR1), a tyrosine kinase that mediates cell adhesion to collagen. DDR1 expression is positively associated with fibrotic lesions of heart, kidney, liver, lung and perivascular tissues. As the most common end-point of all fibrotic disorders is dysregulated collagen remodeling, we consider here the mechanical signaling functions of DDR1 in processing of fibrillar collagen that lead to tissue fibrosis.     

Genetic Analyses of Integrin Signaling

The development of multicellular organisms, as well as maintenance of organ architecture and function, requires robust regulation of cell fates. This is in part achieved by conserved signaling pathways through which cells process extracellular information and translate this information into changes in proliferation, differentiation, migration, and cell shape. Gene deletion studies in higher eukaryotes have assigned critical roles for components of the extracellular matrix (ECM) and their cellular receptors in a vast number of developmental processes, indicating that a large proportion of this signaling is regulated by cell-ECM interactions. In addition, genetic alterations in components of this signaling axis play causative roles in several human diseases. This review will discuss what genetic analyses in mice and lower organisms have taught us about adhesion signaling in development and disease.Almost all cells in multicellular organisms are surrounded by a three-dimensional organized meshwork of macromolecules that constitute the extracellular matrix (ECM). The ECM is a dynamic structure that is generated and constantly remodeled by cells that secrete and manipulate its components into a precise configuration. It functions as a structural framework that provides cells with positional and environmental information, but also forms specialized structures such as cartilage, tendons, basement membranes (BM), bone, and teeth. In addition to its structural properties, the ECM acts as a signaling platform that regulates a large number of cellular functions. It is capable of binding growth factors, chemokines, and cytokines thereby modulating their bioavailability and activity. On the other hand, the ECM is recognized by multiple cell surface receptors that transmit information from the extracellular environment by propagating intracellular signals (for a review, see Hynes 2009).The major cell surface receptors that recognize and assemble the ECM are integrins. Integrins are heterodimeric transmembrane proteins composed of α and β subunits. Eighteen α subunits and eight β subunits can assemble in 24 different combinations with overlapping substrate specificity and cell-type-specific expression patterns (Hynes 2002; Humphries et al. 2006). This, together with the ability of different heterodimers to assemble specific intracellular signaling complexes, provides multiple layers of signaling specificity to these receptors. Conversely, the integrin expression profile of a given cell type determines which ECM components it can bind. Signals arising from integrins regulate virtually all aspects of cell behavior, including cell migration, survival, cell cycle progression, and differentiation.Genetics has proven to be a powerful tool to dissect the functions of ECM–cell interactions in complex organisms. To date, all of the integrin subunits and their major ligands have been deleted in mice. Given the large variety of cellular processes regulated by adhesion signaling, it is not surprising that a significant subset of these proteins has proven to be essential for embryonic development and/or tissue maintenance. However, in addition to underlining the importance of cell-ECM interactions in development, genetic studies also revealed critical roles for tissue- and cell-type-specific modes of adhesion signaling and provided important insights into human disease.     

Integrin-mediated calcium signaling and regulation of cell adhesion by intracellular calcium

Integrins are ubiquitous trans-membrane adhesion molecules that mediate the interaction of cells with the extracellular matrix (ECM). Integrins link cells to the ECM by interacting with the cell cytoskeleton. In cases such as leukocyte binding, integrins mediate cell-cell interactions and cell-ECM interactions. Recent research indicates that integrins also function as signal transduction receptors, triggering a number of intracellular signaling pathways that regulate cell behavior and development. A number of integrins are known to stimulate changes in intracellular calcium levels, resulting in integrin activation. Although changes in intracellular calcium regulate a vast number of cellular functions, this review will discuss the stimulation of calcium signaling by integrins and the role of intracellular calcium in the regulation of integrin-mediated adhesion.     

Niche interactions in epidermal stem cells

Within the epidermis and dermis of the skin, cells secrete and are surrounded by the extracellular matrix(ECM), which provides structural and biochemical support. The ECM of the epidermis is the basement membrane, and collagen and other dermal components constitute the ECM of the dermis. There is significant variation in the composition of the ECM of the epidermis and dermis, which can affect "cell to cell" and "cell to ECM" interactions. These interactions, in turn, can influence biological responses, aging, and wound healing; abnormal ECM signaling likely contributes toskin diseases. Thus, strategies for manipulating cellECM interactions are critical for treating wounds and a variety of skin diseases. Many of these strategies focus on epidermal stem cells, which reside in a unique niche in which the ECM is the most important component; interactions between the ECM and epidermal stem cells play a major role in regulating stem cell fate. As they constitute a major portion of the ECM, it is likely that integrins and type Ⅳ collagens are important in stem cell regulation and maintenance. In this review, we highlight recent research-including our previous work-exploring the role that the ECM and its associated components play in shaping the epidermal stem cell niche.     

Extracellular matrix (ECM) microstructural composition regulates local cell-ECM biomechanics and fundamental fibroblast behavior: a multidimensional perspective.

The extracellular matrix (ECM) provides the principal means by which mechanical information is communicated between tissue and cellular levels of function. These mechanical signals play a central role in controlling cell fate and establishing tissue structure and function. However, little is known regarding the mechanisms by which specific structural and mechanical properties of the ECM influence its interaction with cells, especially within a tissuelike context. This lack of knowledge precludes formulation of biomimetic microenvironments for effective tissue repair and replacement. The present study determined the role of collagen fibril density in regulating local cell-ECM biomechanics and fundamental fibroblast behavior. The model system consisted of fibroblasts seeded within collagen ECMs with controlled microstructure. Confocal microscopy was used to collect multidimensional images of both ECM microstructure and specific cellular characteristics. From these images temporal changes in three-dimensional cell morphology, time- and space-dependent changes in the three-dimensional local strain state of a cell and its ECM, and spatial distribution of beta1-integrin were quantified. Results showed that fibroblasts grown within high-fibril-density ECMs had decreased length-to-height ratios, increased surface areas, and a greater number of projections. Furthermore, fibroblasts within low-fibril-density ECMs reorganized their ECM to a greater extent, and it appeared that beta1-integrin localization was related to local strain and ECM remodeling events. Finally, fibroblast proliferation was enhanced in low-fibril-density ECMs. Collectively, these results are significant because they provide new insight into how specific physical properties of a cell's ECM microenvironment contribute to tissue remodeling events in vivo and to the design and engineering of functional tissue replacements.     

The interplay between tissue growth and scaffold degradation in engineered tissue constructs

In vitro tissue engineering is emerging as a potential tool to meet the high demand for replacement tissue, caused by the increased incidence of tissue degeneration and damage. A key challenge in this field is ensuring that the mechanical properties of the engineered tissue are appropriate for the in vivo environment. Achieving this goal will require detailed understanding of the interplay between cell proliferation, extracellular matrix (ECM) deposition and scaffold degradation. In this paper, we use a mathematical model (based upon a multiphase continuum framework) to investigate the interplay between tissue growth and scaffold degradation during tissue construct evolution in vitro. Our model accommodates a cell population and culture medium, modelled as viscous fluids, together with a porous scaffold and ECM deposited by the cells, represented as rigid porous materials. We focus on tissue growth within a perfusion bioreactor system, and investigate how the predicted tissue composition is altered under the influence of (1) differential interactions between cells and the supporting scaffold and their associated ECM, (2) scaffold degradation, and (3) mechanotransduction-regulated cell proliferation and ECM deposition. Numerical simulation of the model equations reveals that scaffold heterogeneity typical of that obtained from $\mu $ CT scans of tissue engineering scaffolds can lead to significant variation in the flow-induced mechanical stimuli experienced by cells seeded in the scaffold. This leads to strong heterogeneity in the deposition of ECM. Furthermore, preferential adherence of cells to the ECM in favour of the artificial scaffold appears to have no significant influence on the eventual construct composition; adherence of cells to these supporting structures does, however, lead to cell and ECM distributions which mimic and exaggerate the heterogeneity of the underlying scaffold. Such phenomena have important ramifications for the mechanical integrity of engineered tissue constructs and their suitability for implantation in vivo.     

Cell-interactive 3D-scaffold; advances and applications

Culturing cells ex vivo that differentiate and maintain in vivo characteristics holds great promise not only for the pragmatic revelations of cell function but also in tissue engineering and regenerative medicine. Lack of de-novo extra-cellular matrix (ECM) milieu, which plays a crucial role in generating physical and chemical signals besides providing structural support is attributed to be the major hurdle in normal cell growth in vitro. Hence, to comprehend the outcome of cell biology research in clinical context, it is important that the cell culture based models should incorporate both the three dimensional (3D) organization and multi cellular complexity of an organ while allowing experimental interventions in a desirable manner. This calls for the development of ECM-mimicking 3D scaffold, which can be integrated with relevant ECM cues to offer cell interactive versatility for different medical and non-medical applications. Present review discusses the status of ECM mimicking for 3D cell culture and its diverse implications.     

Myofibroblasts. I. Paracrine cells important in health and disease

Myofibroblasts are aunique group of smooth-muscle-like fibroblasts that have a similarappearance and function regardless of their tissue of residence.Through the secretion of inflammatory and anti-inflammatory cytokines,chemokines, growth factors, both lipid and gaseous inflammatorymediators, as well as extracellular matrix proteins and proteases, theyplay an important role in organogenesis and oncogenesis, inflammation,repair, and fibrosis in most organs and tissues. Platelet-derivedgrowth factor (PDGF) and stem cell factor are two secreted proteinsresponsible for differentiating myofibroblasts from embryological stemcells. These and other growth factors cause proliferation ofmyofibroblasts, and myofibroblast secretion of extracellular matrix(ECM) molecules and various cytokines and growth factors causesmobility, proliferation, and differentiation of epithelial orparenchymal cells. Repeated cycles of injury and repair lead to organor tissue fibrosis through secretion of ECM by the myofibroblasts.Transforming growth factor- and the PDGF family of growth factorsare the key factors in the fibrotic response. Because of theirubiquitous presence in all tissues, myofibroblasts play important rolesin various organ diseases and perhaps in multisystem diseases as well.

     

Fibronectin-dependent collagen I deposition modulates the cell response to fibronectin

Communication between cells and the extracellular matrix (ECM) is critical for regulation of cell growth, survival, migration, and differentiation. Remodeling of the ECM can occur under normal physiological conditions, as a result of tissue injury, and in certain pathological conditions. ECM remodeling leads to alterations in ECM composition and organization that can alter many aspects of cell behavior, including cell migration. The cell migratory response varies depending on the type, amount, and organization of ECM molecules present, as well as the integrin and proteoglycan repertoire of the cells. We and others have shown that the deposition of several ECM molecules, including collagen types I and III, depends on the presence and stability of ECM fibronectin. Hence, the effect of fibronectin and fibronectin matrix on cell function may partially depend on its ability to direct the deposition of collagen in the ECM. In this study, we used collagen-binding fibronectin mutants and recombinant peptides that interfere with fibronectin-collagen binding to show that fibronectin-dependent collagen I deposition regulates the cell migratory response to fibronectin. These data show that the ability of fibronectin to organize other proteins in the ECM is an important aspect of fibronectin function and highlight the importance of understanding how interactions between ECM proteins influence cell behavior.