MALDI-TOF mass spectrometric identification of novel intercellular space peptides

We performed the matrix-assisted laser desorption/ionisation, time-of-flight mass spectrometry (MALDI-TOF) analysis of the peptides entering into the composition of not yet explored bioregulators derived from the extracellular matrix of the tissues of the various organs of the mammals, and also plants and fungi. The study included 15 different mammalian tissues, 13 species of plants, and 2 species of fungi. Exploring the bioregulators derived from eye tissues, we demonstrated that their composition includes peptide components with the same values of the molecular weight. The composition of the bioregulators derived from the tissues of various organs of mammals or different species of plants and fungi includes the peptides with different values of molecular weight. Obtained data indicate the growing evidence of the assumptions about the major function of the bioregulators of this group—their involvement in the regulation of tissue-organ homeostasis in the biological systems.     

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.     

Regulation of cell adhesion and migration by cell-derived matrices

Three-dimensional in vitro extracellular matrix models provide a physiological alternative to regular two-dimensional cell culture, though they lack the full diversity of molecular composition and physical properties of whole-animal systems. Cell-derived matrices are extracellular matrices that are the product of matrix secretion and assembly by cells cultured at high density in vitro. After the removal of the cells that produced the matrix, an assembled matrix scaffold is left that closely mimics native stromal fiber organization and molecular content. Cell-derived matrices have been shown to impart in vivo-like responses to cells cultured in these matrices. In this review, we focus on mechanisms through which the distinct molecular and topographical composition of cell-derived matrices directs cellular behavior, specifically through regulation of cell-matrix adhesions and subsequent contributions to the process of cell migration.     

Patterning of the membrane cytoskeleton by the extracellular matrix

The extracellular matrices of different tissues contain components which affect the migration, morphology and differentiation of many types of cells. These forms of cell behavior often involve dramatic changes in cytoskeletal organization. Extracellular matrix components are recognized by specific cell surface receptors which span the membrane and interact with the actin cytoskeleton. In cultured cells, the matrix receptors are concentrated in sites of cell attachment called focal adhesions. Information that is conveyed from the extracellular matrix to the cytoskeleton may involve matrix components, cell surface receptors, as well as the proteins at the cytoplasmic face of the focal adhesion which link the receptors to the actin cytoskeleton.     

Sulfated glycoproteins and extracellular matrix of cultured human pulmonary endothelial cells

Endothelial cells derived from human pulmonary arteries incorporate (³H)-glucosamine and ³⁵SO₄ into glycosaminoglycans and into the carbohydrate side chains of glycoproteins. These ³H/³⁵S-carbohydrate chains were isolated from cells and culture medium after Pronase digestion. The ³H/³⁵S-glycosaminoglycans were separated from the ³H/³⁵S glycopeptides by chromatography on Sephadex G-50. The distribution of cellular glycosaminoglycans and glycopeptides indicated that 30–60% of the cellular ³⁵S-glycopeptides may be associated with the matrix components that are synthesized by the cell and attached to a plastic substratum. Human pulmonary arterial endothelial cells were grown on collagen or on a matrix derived from vascular smooth muscle cells in order to investigate how smooth muscle cell extracellular matrix components may regulate the synthesis of endothelial cell glycoconjugates. Endothelial cells grown on plastic release various proportions of the glycoconjugates they synthesize into the culture medium. However, these same cells, when grown on substratum composed of extracellular matrix materials, synthesized altered proportions of cell-associated glycosaminoglycans and reduced the levels of total glycosaminoglycans they released into the culture medium. Thus the growth of endothelial cells on a matrix of smooth muscle cell components indicates that the glycosaminoglycan materials released into the culture medium by cells grown on a plastic substratum may not be an accurate reflection of the levels or composition of extracellular matrix materials made by endothelial cells in vivo.     

Extracellular matrix formation by epithelial cells from human polycystic kidney cysts in culture

Cells from the cysts of patients with autosomal dominant polycystic kidney disease (PKD) were grown in vitro under standard conditions without the aid of collagen-pretreated surfaces, and both the synthesis and composition of the extracellular matrix were investigated. At confluence, PKD cells presented the typical features of epithelial cells, but showed a different collagen composition from fibroblasts. Compared with normal tubular epithelia (NTE), PKD monolayers produced an excess of extracellular matrix, which accounted for 30% of the total incorporation of [³H] proline, although this value was considerably lower (by a factor of 10) in the case of NTE. Immunohistochemical and electrophoretic techniques revealed a complex collagen composition in the extracellular matrix which included [α(III)]₃ and collagen IV. However, part of the collagen components remained unidentified in spite of the fact that they exhibited a typical M_r of α₁(I) and α₂(I) in the presence of urea. Immunoprecipitation with monospecific antibodies and Northern blotting with specific probes failed to recognize α₁(I) and α₂(I), but demonstrated their presence in fibroblasts. Purification and cyanogen bromide digestion demonstrated a strong interhomology in fingerprint peptide composition among the uncharacterized collagens synthesized by PKD cells, thus suggesting a common identity. These observations document a markedly augmented production of extracellular matrix by PKD cultured cells in vitro, and show the presence of collagens which do not share homologies with the major collagen molecules. A better characterization of extracellular matrix composition is central to any comprehension of the cystogenetic mechanisms in vivo.     

Connective tissue growth factor coordinates chondrogenesis and angiogenesis during skeletal development

Coordinated production and remodeling of the extracellular matrix is essential during development. It is of particular importance for skeletogenesis, as the ability of cartilage and bone to provide structural support is determined by the composition and organization of the extracellular matrix. Connective tissue growth factor (CTGF, CCN2) is a secreted protein containing several domains that mediate interactions with growth factors, integrins and extracellular matrix components. A role for CTGF in extracellular matrix production is suggested by its ability to mediate collagen deposition during wound healing. CTGF also induces neovascularization in vitro, suggesting a role in angiogenesis in vivo. To test whether CTGF is required for extracellular matrix remodeling and/or angiogenesis during development, we examined the pattern of Ctgf expression and generated Ctgf-deficient mice. Ctgf is expressed in a variety of tissues in midgestation embryos, with highest levels in vascular tissues and maturing chondrocytes. We confirmed that CTGF is a crucial regulator of cartilage extracellular matrix remodeling by generating Ctgf(-/-) mice. Ctgf deficiency leads to skeletal dysmorphisms as a result of impaired chondrocyte proliferation and extracellular matrix composition within the hypertrophic zone. Decreased expression of specific extracellular matrix components and matrix metalloproteinases suggests that matrix remodeling within the hypertrophic zones in Ctgf mutants is defective. The mutant phenotype also revealed a role for Ctgf in growth plate angiogenesis. Hypertrophic zones of Ctgf mutant growth plates are expanded, and endochondral ossification is impaired. These defects are linked to decreased expression of vascular endothelial growth factor (VEGF) in the hypertrophic zones of Ctgf mutants. These results demonstrate that CTGF is important for cell proliferation and matrix remodeling during chondrogenesis, and is a key regulator coupling extracellular matrix remodeling to angiogenesis at the growth plate.     

The Biofilm Matrix

The extracellular matrix is a complex and extremely important component of all biofilms, providing architectural structure and mechanical stability to the attached population. The matrix is composed of cells, water and secreted/released extracellular macromolecules. In addition, a range of enzymic and regulatory activities can be found within the matrix. Together, these different components and activities are likely to interact and in so doing create a series of local environments within the matrix which co-exist as a functional consortium. The matrix architecture is also subject to a number of extrinsic factors, including fluctuations in nutrient and gaseous levels and fluid shear. Together, these intrinsic and extrinsic factors combine to produce a dynamic, heterogeneous microenvironment for the attached and enveloped cells.     

Fibroblasts retain their tissue phenotype when grown in three-dimensional collagen gels

Fibroblasts are responsible for the synthesis, assembly, deposition, and organization of extracellular matrix molecules, and thus determine the morphology of connective tissues. Deposition of matrix molecules occurs in extracellular compartments, where the sequential stages are under cellular control. Cell orientation/polarity is important in determining how the cell orients these extracytoplasmic compartments and therefore how the matrix is assembled and oriented. However, the control of cell orientation is not understood. Fibroblasts from three tissues with different morphologies were studied to determine whether cells maintained their characteristic phenotype. Fibroblasts from cornea, which in vivo are oriented in orthogonal layers along with their matrix; from tendon, a uniaxial connective tissue, where cells orient parallel to each other; and from dermis, a connective tissue with no apparent cellular orientation, were used to study cell morphology and orientation in three-dimensional collagen gels. The different cells were grown for 3 and 7 days in identical three-dimensional collagen gels with a nonoriented matrix. Confocal fluorescence microscopy demonstrated that corneal fibroblasts oriented perpendicular to one another at 3 days, and after 7 days in hydrated gels these cells formed orthogonal sheets. Tendon fibroblasts were shown by the same methods to orient parallel to one another in bundles at both 3 and 7 days, throughout the depth of the gel. Dermal fibroblasts showed no apparent orientation throughout the hydrated gels at either time point examined. The organization of these different cell types was consistent with their tissue of origin as was the cell structure and polarity. These studies imply that cellular and tissue phenotype is innate to differentiated fibroblasts and that these cells will orient in a tissue-specific manner regardless of the extracellular matrix present.     

Sturctural analysis of cell:matrix association during the morphogenesis of atrioventricular cushion tissue.

Translocation of an endocardially seeded cushion cell progeny across a broad acellular expanse of extracellular matrix (ECM) constitutes a fundamental morphogenetic event in the development of atrioventricular (AV) cushion pads, the primordia of membraneous septa and cardiac valves. Transmission, scanning, and high-voltage electron microscopy together with light microscopic examination of living or fixed tissues were utilized to determine if (1) one component of the ECM more than any other interacted with the motility-like appendages of cushion cells in such a manner as to suggest a physical substratum; (2) any ECM components were organized into polarized “tracks” which could serve to guide cells centrifugally; and (3) cell:ECM associations varied among the cells comprising the migratory wave. Results indicated that two morphologically identifiable matrix components, microfibrils and a continuum of solid pleomorphic strands of heterogeneous composition called cetylpyridinium chloride (CPCL)-dependent matrix, comprised the bulk of the premigratory ECM. Contact of the premigratory matrix by cushion cells at the leading edge (pioneer cells) of the migratory wave coincided with modification in composition of the CPCL matrix and alignment of microfibrils into polarized tracks (an event seemingly dependent on motility appendage formation, since cells lacking processes after cytochalasin B treatment had altered track associations). Trailing cushion cells uniformly populated the ECM, never piled up against the myocardium, had no track associations, formed numerous cell to cell associations, and were coated with a granular remnant of disrupted CPCL-dependent matrix. The foregoing data suggest that active in vivo translocation and subsequent stabilization of cushion cells involve alignment and compositional changes in the premigratory ECM, events linked temporally with the passage of pioneer cells.     

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.     

Metalloproteinases in disease: identification of biomarkers of tissue damage through proteomics

ABSTRACT

Introduction: Metalloproteinases play key roles in health and disease, by generating novel proteoforms with variable structure and function.

Areas covered: This review focuses on the role of endogenous [a Disintegrin and Metalloproteinase (ADAMs), ADAMs with thrombospondin motifs (ADAMTS), and matrix metalloproteinases (MMPs)] and exogenous metalloproteinases in various disease conditions, and describes the application of mass spectrometry-based proteomics to detect qualitative and quantitative changes in protein profiles in tissues and body fluids in disease. Emphasis is placed on the proteomic analysis of exudates collected from affected tissues, including methods that enrich newly generated protein fragments derived from proteolysis in cells, stroma, or extracellular matrix. The use of proteomic analysis of exudates in the study of the local tissue damage induced by metalloproteinases derived from viperid snake venoms is discussed, particularly in relation to extracellular matrix degradation and to the overall pathology of these envenomings.

Expert commentary: The information provided by these proteomics approaches is paving the way for the identification of biomarkers based on particular proteolytic signatures associated with different pathologies. Together with other methodological approaches, a comprehensive view of the mechanisms and dynamics of diseases can be achieved. Such basis of knowledge allows for the design of novel diagnostic and therapeutic approaches within the frame of ‘precision’ or ‘personalized’ medicine.     

Extracellular matrix organization in developing muscle: correlation with acetylcholine receptor aggregates

Monoclonal antibodies recognizing laminin, heparan sulfate proteoglycan, fibronectin, and two apparently novel connective tissue components have been used to examine the organization of extracellular matrix of skeletal muscle in vivo and in vitro. Four of the five monoclonal antibodies are described for the first time here. Immunocytochemical experiments with frozen-sectioned muscle demonstrated that both the heparan sulfate proteoglycan and laminin exhibited staining patterns identical to that expected for components of the basal lamina. In contrast, the remaining matrix constituents were detected in all regions of muscle connective tissue: the endomysium, perimysium, and epimysium. Embryonic muscle cells developing in culture elaborated an extracellular matrix, each antigen exhibiting a unique distribution. Of particular interest was the organization of extracellular matrix on myotubes: the build-up of matrix components was most apparent in plaques overlying clusters of an integral membrane protein, the acetylcholine receptor (AChR). The heparan sulfate proteoglycan was concentrated at virtually all AChR clusters and showed a remarkable level of congruence with receptor organization; laminin was detected at 70-95% of AChR clusters but often was not completely co-distributed with AChR within the cluster; fibronectin and the two other extracellular matrix antigens occurred at approximately 20, 8, and 2% of the AChR clusters, respectively, and showed little or no congruence with AChR. From observations on the distribution of extracellular matrix components in tissue cultured fibroblasts and myogenic cells, several ideas about the organization of extracellular matrix are suggested. (a) Congruence between AChR clusters and heparan sulfate proteoglycan suggests the existence of some linkage between the two molecules, possibly important for regulation of AChR distribution within the muscle membrane. (b) The qualitatively different patterns of extracellular matrix organization over myotubes and fibroblasts suggest that each of these cell types uses somewhat different means to regulate the assembly of extracellular matrix components within its domain. (c) The limited co-distribution of different components within the extracellular matrix in vitro and the selective immune precipitation of each antigen from conditioned medium suggest that each extracellular matrix component is secreted in a form that is not complexed with other matrix constituents.     

Synthesis and characterization of silver nanoparticles decorated polydopamine coated hexagonal boron nitride and its effect on wound healing

BackgroundWound healing is an essential physiological process involving many cell types and their products acting in a marvellous harmony to repair damaged tissues. During the healing process, cellular proliferation and extracellular matrix remodelling stages could be interrupted by undesired factors including microorganisms and altered metabolic activities. In such a case, the process requires some external stimulants to accelerate or remediate the healing stages.MethodsIn this study, we report a multifunctional wound healing stimulating agent. In this context, hexagonal boron nitride (hBN) nanoparticles, silver nanoparticles (AgNPs) and polydopamine(pdopa) were used through mussel-inspired chemistry of dopamine to obtain pdopa coated hBN (hBN@pdopa) and AgNPs decorated hBN@pdopa (hBN@pdopa-AgNPs). These two nanostructures were investigated to observe stages of healing.ResultsAgNPs were chosen for inflammation reduction and hBN for induced cell proliferation and migration. In in vitro experiments, firstly, high cellular uptake capacity and biocompatibility of hBN@pdopa and hBN@pdopa-AgNPs were evaluated. They were also tested for their reaction against increased concentration of reactive oxygen species (ROS) in injured cells. Finally, their effect on cellular migration, intracellular tube formation and F-actin organization were monitored by light and confocal microscopy, respectively.ConclusionThe results clearly indicate that the hBN@pdopa-AgNPs significantly decrease ROS production, promote wound closure, and reorganize tube formation in cells.     

Interactions of Human Skin Fibroblasts with Monomeric or Fibrillar Collagens Induce Different Organization of the Cytoskeleton

Fibrillar collagens represent the most abundant extracellular matrix components surrounding fibroblasts. Although there is a large heterogeneity in the collagen composition and in the physiological functions of different tissues, interactions between cells and native collagens monomers are mediated by only two integrins, the α1β1 and α2β1 integrins. In tissue, fibroblasts are exposed to collagen polymers, supramolecular assemblies which might play a role on the availability of the cell-binding sites at the surface of the fibrils. We have addressed this issue by investigating the patterns of adhesion structures in normal human skin fibroblasts exposed to collagen monomers or polymers. Our results showed that cell morphology, cell adhesion pattern, actin organization, and distribution of integrin subunits, talin, vinculin, and phosphotyrosine-containing proteins are dependent on the supramolecular organization of the collagens. In particular, compared to monomers, collagen polymers induced a looser organization of the actin network and a linear clustering of integrins, talin, vinculin, and phosphotyrosine-containing proteins. These results emphasize the role of the physical state of collagen on cellular interactions and underline the role of the extracellular matrix in the phenotypic modulation of fibroblasts. Furthermore, our studies suggest the existence of a local heterogeneity in the biological activity of collagen fibrils.     

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.     

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.     

The spatial structures of hypolithic communities in the Dry Valleys of East Antarctica

Hypolithic communities represent important reservoirs of microbial life in hyper-arid deserts. A number of studies on the diversity and ecology of these communities from different geographic areas have been reported in the past decade, but the spatial distribution of the different components of these communities is still not understood. Moss- and cyanobacteria-dominated hypolithic community morphotypes from Miers Valley (McMurdo Dry Valleys, East Antarctica) were analyzed by electron microscopy in order to characterize the microscale spatial structure. The two communities showed a high degree of internal organization, but differing according to the biological composition. In moss-dominated hypoliths, the moss plantlets are intermixed with mineral fragments of soil origin. However, in cyanobacteria-dominated hypoliths, a layered spatial organization was structured by filamentous cyanobacteria and associated extracellular polymeric components. While moss cells were lacking in cyanobacteria-dominated communities, biofilms formed by cyanobacteria and heterotrophic bacteria were observed in both community morphotypes. The water-holding capacity of both live and dead moss cells and the associated organic matrix, together with the protective properties of the extracellular polymeric substances, could facilitate the survival and activity of these communities. Similar structural strategies can favor the survival of microbial communities in different extreme environments.     

Assessment of hyaline cartilage matrix composition using near infrared spectroscopy

Changes in the composition of the extracellular matrix (ECM) are characteristic of injury or disease in cartilage tissue. Various imaging modalities and biochemical techniques have been used to assess the changes in cartilage tissue but lack adequate sensitivity, or in the case of biochemical techniques, result in destruction of the sample. Fourier transform near infrared (FT-NIR) spectroscopy has shown promise for the study of cartilage composition. In the current study NIR spectroscopy was used to identify the contributions of individual components of cartilage in the NIR spectra by assessment of the major cartilage components, collagen and chondroitin sulfate, in pure component mixtures. The NIR spectra were obtained using homogenous pellets made by dilution with potassium bromide. A partial least squares (PLS) model was calculated to predict composition in bovine cartilage samples. Characteristic absorbance peaks between 4000 and 5000 cm⁻¹ could be attributed to components of cartilage, i.e. collagen and chondroitin sulfate. Prediction of the amount of collagen and chondroitin sulfate in tissues was possible within 8% (w/dw) of values obtained by gold standard biochemical assessment. These results support the use of NIR spectroscopy for in vitro and in vivo applications to assess matrix composition of cartilage tissues, especially when tissue destruction should be avoided.     

The Gyc76C Receptor Guanylyl Cyclase and the Foraging cGMP-Dependent Kinase Regulate Extracellular Matrix Organization and BMP Signaling in the Developing Wing of Drosophila melanogaster

The developing crossveins of the wing of Drosophila melanogaster are specified by long-range BMP signaling and are especially sensitive to loss of extracellular modulators of BMP signaling such as the Chordin homolog Short gastrulation (Sog). However, the role of the extracellular matrix in BMP signaling and Sog activity in the crossveins has been poorly explored. Using a genetic mosaic screen for mutations that disrupt BMP signaling and posterior crossvein development, we identify Gyc76C, a member of the receptor guanylyl cyclase family that includes mammalian natriuretic peptide receptors. We show that Gyc76C and the soluble cGMP-dependent kinase Foraging, likely linked by cGMP, are necessary for normal refinement and maintenance of long-range BMP signaling in the posterior crossvein. This does not occur through cell-autonomous crosstalk between cGMP and BMP signal transduction, but likely through altered extracellular activity of Sog. We identify a novel pathway leading from Gyc76C to the organization of the wing extracellular matrix by matrix metalloproteinases, and show that both the extracellular matrix and BMP signaling effects are largely mediated by changes in the activity of matrix metalloproteinases. We discuss parallels and differences between this pathway and other examples of cGMP activity in both Drosophila melanogaster and mammalian cells and tissues.