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Angiopoietin-2 (ANGPT2) is a context-dependent TIE2 agonistic or antagonistic ligand that induces diverse responses in cancer. Blocking ANGPT2 provides a promising strategy for inhibiting tumor growth and metastasis, yet variable effects of targeting ANGPT2 have complicated drug development. ANGPT2443 is a naturally occurring, lower oligomeric protein isoform whose expression is increased in cancer. Here, we use a knock-in mouse line (mice expressing Angpt2443), a genetic model for breast cancer and metastasis (MMTV-PyMT), a syngeneic melanoma lung colonization model (B16F10), and orthotopic injection of E0771 breast cancer cells to show that alternative forms increase the diversity of Angpt2 function. In a mouse retina model of angiogenesis, expression of Angpt2443 caused impaired venous development, suggesting enhanced function as a competitive antagonist for Tie2. In mammary gland tumor models, Angpt2443 differentially affected primary tumor growth and vascularization; these varying effects were associated with Angpt2 protein localization in the endothelium or in the stromal extracellular matrix as well as the frequency of Tie2-positive tumor blood vessels. In the presence of metastatic cells, Angpt2443 promoted destabilization of pulmonary vasculature and lung metastasis. In vitro, ANGPT2443 was susceptible to proteolytical cleavage, resulting in a monomeric ligand (ANGPT2DAP) that inhibited ANGPT1- or ANGPT4-induced TIE2 activation but did not bind to alternative ANGPT2 receptor α5β1 integrin. Collectively, these data reveal novel roles for the ANGPT2 N-terminal domain in blood vessel remodeling, tumor growth, metastasis, integrin binding, and proteolytic regulation. Significance: This study identifies the role of the N-terminal oligomerization domain of angiopoietin-2 in vascular remodeling and lung metastasis and provides new insights into mechanisms underlying the versatile functions of angiopoietin-2 in cancer. See related commentary by Kamiyama and Augustin, p. 35

The extracellular matrix (ECM) is ubiquitously involved in neoplastic transformation, tumour growth and metastatic dissemination, and the interplay between tumour and stromal cells and the ECM is now considered crucial for the formation of a tumour-supporting microenvironment. The 28 different collagens (Col) form a major ECM protein family and display extraordinary functional diversity in tissue homeostasis as well as in pathological conditions, with functions ranging from structural support for tissues to regulatory binding activities and storage of biologically active cryptic domains releasable through ECM proteolysis. Two subfamilies of collagens, namely the plasma membrane-associated collagens with interrupted triple-helices (MACITs, including ColXIII, ColXXIII and ColXXV) and the basement membrane-associated collagens with multiple triple-helix domains with interruptions (multiplexins, including ColXV and ColXVIII), have highly interesting regulatory functions in tissue and organ development, as well as in various diseases, including cancer. An increasing, albeit yet sparse, data suggest that these collagens play crucial roles in conveying regulatory signals from the extracellular space to cells. We summarize here the current knowledge about MACITs and multiplexins as regulators of stemness and oncogenic processes, as well as their roles in influencing cell fate decisions in healthy and cancerous tissues. In addition, we present a bioinformatic analysis of the impacts of MACITs and multiplexins transcript levels on the prognosis of patients representing a wide array of malignant diseases, to aid future diagnostic and therapeutic efforts.

Collagen XIII is a conserved transmembrane collagen mainly expressed in mesenchymal tissues. Previously, we have shown that collagen XIII modulates tissue development and homeostasis. Integrins are a family of receptors that mediate signals from the environment into the cells and vice versa. Integrin α11β1 is a collagen receptor known to recognize the GFOGER (O=hydroxyproline) sequence in collagens. Interestingly, collagen XIII and integrin α11β1 both have a role in the regulation of bone homeostasis. To study whether α11β1 is a receptor for collagen XIII, we utilized C2C12 cells transfected to express α11β1 as their only collagen receptor. The interaction between collagen XIII and integrin α11β1 was also confirmed by surface plasmon resonance and pull-down assays. We discovered that integrin α11β1 mediates cell adhesion to two collagenous motifs, namely GPKGER and GF(S)QGEK, that were shown to act as the recognition sites for the integrin α11-I domain. Furthermore, we studied the in vivo significance of the α11β1-collagen XIII interaction by crossbreeding α11 null mice (Itga11−/−) with mice overexpressing Col13a1 (Col13a1oe). When we evaluated the bone morphology by microcomputed tomography, Col13a1oe mice had a drastic bone overgrowth followed by severe osteoporosis, whereas the double mutant mouse line showed a much milder bone phenotype. To conclude, our data identifies integrin α11β1 as a new collagen XIII receptor and demonstrates that this ligand-receptor pair has a role in the maintenance of bone homeostasis.

The binomial nervous system involves the central nervous system (CNS), comprising the brain and the spinal cord, and the peripheral nervous system (PNS). Both divisions of the nervous system contain electrically excitable neurons as well as a number of supporting neuroglial cells, which include oligodendrocytes, astrocytes, microglia, choroid plexus ependymal cells in the CNS, and satellite and Schwann cells in the PNS. Connective tissues rich in fibrillar collagens form the outermost cover for the nervous system proper. Moreover, there are rich basement membranes (BM) surrounding all nervous system tissues and vessels within these structures. BMs compartmentalize nervous tissues and contribute to selective barrier and filtration functions essential for brain homeostasis. While BMs are absent from the brain parenchyma, there are extracellular matrices (ECM) in these regions that remain under-explored. The composition and types of matrices differ substantially in different parts of the nervous system. ECMs are significantly abundant during development, guiding cellular migration and differentiation as well as axon navigation and synaptogenesis. Additionally, ECMs promote neuronal health, contribute to synaptic homeostasis and plasticity, and are upregulated in response to disease and trauma. It is for these reasons that the mutation and malfunction of collagens have been linked to neurodevelopmental, degenerative, and psychiatric disorders as well as motor and sensory dysfunction. In recent years, it has become clear that collagens, constituting a major family of ECM proteins, and other extracellular components are generated not only by glial cells but also by neurons. The functions and expression patterns of the collagen superfamily members in the nervous system are summarized in this chapter, where we focus on their roles in vitro and in vivo in a number of animal models, and in human diseases of the nervous system.

Introduction: Myasthenia Gravis (MG) is a neuromuscular junction (NMJ) disorder caused by autoantibodies against NMJ proteins. Collagen XIII is a muscle-derived transmembrane protein required for NMJ maturation. The objective of this study is to explore existence of autoantibodies to collagen XIII in MG patients. Methods: Seventy MG patient sera and 61 human healthy controls were screened for collagen XIII autoantibodies by enzyme-linked immunosorbent assay (ELISA). The collagen XIII autoantibody-positive sera were further analyzed by a cell-based assay together with western blotting and immunofluorescence staining of cells expressing recombinant collagen XIII. Results: Five of the 70 MG patients were found to have autoantibodies against collagen XIII. All the five patients were young women with no or low levels of autoantibodies to acetylcholine receptor (AChR). Discussion: Collagen XIII is associated with MG but it is unknown if the collagen XIII autoantibodies are pathogenic. This article is protected by copyright. All rights reserved.

Both transmembrane and extracellular cues, one of which is collagen XIII, regulate the formation and function of the neuromuscular synapse, and their absence results in myasthenia. We show that the phenotypical changes in collagen XIII knock-out mice are milder than symptoms in human patients, but the Col13a1-/- mice recapitulate major muscle findings of congenital myasthenic syndrome type 19 and serve as a disease model. In the lack of collagen XIII neuromuscular synapses do not reach full size, alignment, complexity and function resulting in reduced muscle strength. Collagen XIII is particularly important for the preterminal integrity, and when absent, destabilization of the motor nerves results in muscle regeneration and in atrophy especially in the case of slow muscle fibers. Collagen XIII was found to affect synaptic integrity through binding the ColQ tail of acetylcholine esterase. Although collagen XIII is a muscle-bound transmembrane molecule, it also undergoes ectodomain shedding to become a synaptic basal lamina component. We investigated the two forms' roles by novel Col13a1tm/tm mice in which ectodomain shedding is impaired. While postsynaptic maturation, terminal branching and neurotransmission was exaggerated in the Col13a1tm/tm mice, the transmembrane form's presence sufficed to prevent defects in transsynaptic adhesion, Schwann cell invagination/retraction, vesicle accumulation and acetylcholine receptor clustering and acetylcholinesterase dispersion seen in the Col13a1-/- mice, pointing to the transmembrane form as the major conductor of collagen XIII effects. Altogether, collagen XIII secures postsynaptic, synaptic and presynaptic integrity, and it is required for gaining and maintaining normal size, complexity and functional capacity of the neuromuscular synapse.

Alongside playing structural roles, the extracellular matrix (ECM) acts as an interaction platform for cellular homeostasis, organ development, and maintenance. The necessity of the ECM is highlighted by the diverse, sometimes very serious diseases that stem from defects in its components. The neuromuscular junction (NMJ) is a large peripheral motor synapse differing from its central counterparts through the ECM included at the synaptic cleft. Such synaptic basal lamina (BL) is specialized to support NMJ establishment, differentiation, maturation, stabilization, and function and diverges in molecular composition from the extrasynaptic ECM. Mutations, toxins, and autoantibodies may compromise NMJ integrity and function, thereby leading to congenital myasthenic syndromes (CMSs), poisoning, and autoimmune diseases, respectively, and all these conditions may involve synaptic ECM molecules. With neurotransmission degraded or blocked, muscle function is impaired or even prevented. At worst, this can be fatal. The article reviews the synaptic BL composition required for assembly and function of the NMJ molecular machinery through the lens of studies primarily with mouse models but also with human patients. In-depth focus is given to collagen XIII, a postsynaptic-membrane-spanning but also shed ECM protein that in recent years has been revealed to be a significant component for the NMJ. Its deficiency in humans causes CMS, and autoantibodies against it have been recognized in autoimmune myasthenia gravis. Mouse models have exposed numerous details that appear to recapitulate human NMJ phenotypes relatively faithfully and thereby can be readily used to generate information necessary for understanding and ultimately treating human diseases. Anat Rec, 2019. © 2019 Wiley Periodicals, Inc.

Chemical synapses allow neurons to perform complex computations and regulate other systems of the body. At a chemical synapse, pre- and postsynaptic sites are separated by a small space (the synaptic cleft) and surrounded by astrocytes. The basement membrane (BM), a sheetlike, specialized extracellular matrix (ECM), is found ubiquitously in the PNS. It has become clear that the ECMs not only play a structural role but also serve as barriers and filters in the PNS and CNS. Moreover, proteoglycans and tenascin family proteins in the ECM regulate synapse formation and synaptic plasticity. Although CNS synapses lack the BMs, recent results indicate that the BM-associated collagens are also present in the CNS synaptic cleft and affect synaptogenesis in both the CNS and the PNS. The C1q domain-containing family proteins are important components of the CNS synaptic cleft in regulating synapse formation, maintenance, and the pruning process. The ECM is regarded as a crucial component of the tetrapartite synapse, consisting of pre- and postsynaptic neurons, astrocyte, and ECM.