Type IIA Procollagen Containing the Cysteine-rich Amino Propeptide Is Deposited in the Extracellular Matrix of Prechondrogenic Tissue and Binds to TGF-β1 and BMP-2

Type II procollagen is expressed as two splice forms. One form, type IIB, is synthesized by chondrocytes and is the major extracellular matrix component of cartilage. The other form, type IIA, contains an additional 69 amino acid cysteine-rich domain in the NH2-propeptide and is synthesized by chondrogenic mesenchyme and perichondrium. We have hypothesized that the additional protein domain of type IIA procollagen plays a role in chondrogenesis. The present study was designed to determine the localization of the type IIA NH2-propeptide and its function during chondrogenesis. Immunofluorescence histochemistry using antibodies to three domains of the type IIA procollagen molecule was used to localize the NH2-propeptide, fibrillar domain, and COOH-propeptides of the type IIA procollagen molecule during chondrogenesis in a developing human long bone (stage XXI). Before chondrogenesis, type IIA procollagen was synthesized by chondroprogenitor cells and deposited in the extracellular matrix. Immunoelectron microscopy revealed type IIA procollagen fibrils labeled with antibodies to NH2-propeptide at ∼70 nm interval suggesting that the NH2-propeptide remains attached to the collagen molecule in the extracellular matrix. As differentiation proceeds, the cells switch synthesis from type IIA to IIB procollagen, and the newly synthesized type IIB collagen displaces the type IIA procollagen into the interterritorial matrix. To initiate studies on the function of type IIA procollagen, binding was tested between recombinant NH2-propeptide and various growth factors known to be involved in chondrogenesis. A solid phase binding assay showed no reaction with bFGF or IGF-1, however, binding was observed with TGF-β1 and BMP-2, both known to induce endochondral bone formation. BMP-2, but not IGF-1, coimmunoprecipitated with type IIA NH2-propeptide. Recombinant type IIA NH2-propeptide and type IIA procollagen from media coimmunoprecipitated with BMP-2 while recombinant type IIB NH2-propeptide and all other forms of type II procollagens and mature collagen did not react with BMP-2. Taken together, these results suggest that the NH2-propeptide of type IIA procollagen could function in the extracellular matrix distribution of bone morphogenetic proteins in chondrogenic tissue.

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Type IIA Procollagen Amino Propeptide Is Localized in Human Embryonic Tissues

Journal of Histochemistry & Cytochemistry ◽

10.1177/002215549704501104 ◽

1997 ◽

Vol 45 (11) ◽

pp. 1469-1480 ◽

Cited By ~ 82

Author(s):

Anush Oganesian ◽

Yong Zhu ◽

Linda J. Sandell

Keyword(s):

Extracellular Matrix ◽

Type Ii Collagen ◽

Protein Domain ◽

Collagen Molecule ◽

Endochondral Bone Formation ◽

Type Ii ◽

Exon 2 ◽

Helical Domain ◽

Chondroprogenitor Cells ◽

Triple Helical Domain

Type II procollagen is synthesized in two forms generated by the alternative splicing of its precursor mRNA. The alternatively spliced domain, exon 2, encodes the 69-amino-acid cysteinerich region of the NH2 propeptide. Studies of mRNA expression have shown that the longer form, designated Type IIA procollagen, is synthesized by chondroprogenitor cells and various noncartilaginous tissues. The shorter form, Type IIB procollagen, is synthesized by differentiated chondrocytes. As the initial step in our investigations of the function of the Type IIA procollagen, the protein domain corresponding to exon 2 was created as a recombinant fusion protein and used to raise antibodies in rabbits. The resulting antiserum was specific for Type IIA procollagen NH2 propeptide as shown by ELISA, Western blotting, and immunofluorescent co-localization with the triple-helical domain of Type II collagen. Type IIA procollagen was identified in tissue culture medium of 54-day human fetal ribs. Confocal microscopy was used to localize the Type IIA NH2 propeptide in Day 50 and 53 human embryos. In the digital rays of the developing hand, where only Type IIA procollagen mRNA was detected, Type IIA procollagen NH2 propeptide was observed in the extracellular matrix. The presence of Type IIA procollagen NH2 propeptide was observed in the cartilage of the developing long bones of the lower arm and vertebral bodies even though these tissues synthesize Type IIB mRNA at this developmental stage. Type IIA procollagen NH2 propeptide was localized in the developing trachea, a cartilage that does not undergo endochondral bone formation. Type IIA NH2 propeptide was also localized in noncartilaginous tissues known to synthesize Type IIA mRNA, such as the intervertebral area, perichondrium, notochordal sheath, and neuroepithelium of the otic vesicle. In most tissues, co-localization with antiserum against the triple-helical domain of Type II collagen was observed. Positive immunoreactivity with the Type IIA NH2 propeptide antiserum indicates, for the first time, that this propeptide is present in the tissue. Co-localization of NH2 propeptide antibodies with the triple-helical domain of the collagen molecule suggests that Type IIA procollagen is intact in the extracellular matrix of these tissues. Taken together, these results strongly suggest that around cells that synthesize Type IIA procollagen mRNA, Type IIA procollagen NH2 propeptide is secreted and deposited into the extracellular matrix. In light of these results, we predict that Type IIA procollagen plays a role in differentiation of tissues that augments its purely architectural function. (J Histochem Cytochem 45:1469–1480, 1997)

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Regulation of extracellular matrix synthesis by TNF-α and TGF-β1 in type II cells exposed to coal dust

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.1998.275.4.l637 ◽

1998 ◽

Vol 275 (4) ◽

pp. L637-L644 ◽

Cited By ~ 3

Author(s):

Yu-Chen Lee ◽

D. Eugene Rannels

Keyword(s):

Extracellular Matrix ◽

Cell Cultures ◽

Transforming Growth Factor ◽

Coal Dust ◽

Transforming Growth Factor Β1 ◽

Type Ii ◽

Tnf Α ◽

Type Ii Cell ◽

Primary Type ◽

Tgf Β1

Type II pulmonary epithelial cells respond to anthracite coal dust PSOC 867 with increased synthesis of extracellular matrix (ECM) components. Alveolar macrophages modulate this response by pathways that may involve soluble mediators, including tumor necrosis factor-α (TNF-α) or transforming growth factor-β1 (TGF-β1). The effects of TNF-α (10 ng/ml) and/or TGF-β1 (2 ng/ml) were thus investigated in dust-exposed primary type II cell cultures. In control day 1 or day 3 cultures, TNF-α and/or TGF-β1 had little or no effect on the synthesis of type II cellular proteins, independent of whether the cells were exposed to dust. With PSOC 867 exposure, where ECM protein synthesis is elevated, TNF-α and TGF-β1 further increased both the absolute and relative rates of ECM synthesis on day 3 but had little effect on day 1. Each mediator increased expression of fibronectin mRNA, as well as of ECM fibronectin content, in a manner qualitatively similar to their effects on synthesis. Thus TNF-α and TGF-β1 modulate both ECM synthesis and fibronectin content in coal dust-exposed type II cell cultures.

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On the role of type IX collagen in the extracellular matrix of cartilage: type IX collagen is localized to intersections of collagen fibrils.

The Journal of Cell Biology ◽

10.1083/jcb.102.5.1931 ◽

1986 ◽

Vol 102 (5) ◽

pp. 1931-1939 ◽

Cited By ~ 113

Author(s):

W Müller-Glauser ◽

B Humbel ◽

M Glatt ◽

P Sträuli ◽

K H Winterhalter ◽

...

Keyword(s):

Extracellular Matrix ◽

Bone Formation ◽

Tissue Distribution ◽

Chicken Embryo ◽

Polyclonal Antibodies ◽

Type Ii Collagen ◽

Endochondral Bone Formation ◽

Type Ii ◽

Endochondral Bone

The tissue distribution of type II and type IX collagen in 17-d-old chicken embryo was studied by immunofluorescence using polyclonal antibodies against type II collagen and a peptic fragment of type IX collagen (HMW), respectively. Both proteins were found only in cartilage where they were co-distributed. They occurred uniformly throughout the extracellular matrix, i.e., without distinction between pericellular, territorial, and interterritorial matrices. Tissues that undergo endochondral bone formation contained type IX collagen, whereas periosteal and membranous bones were negative. The thin collagenous fibrils in cartilage consisted of type II collagen as determined by immunoelectron microscopy. Type IX collagen was associated with the fibrils but essentially was restricted to intersections of the fibrils. These observations suggested that type IX collagen contributes to the stabilization of the network of thin fibers of the extracellular matrix of cartilage by interactions of its triple helical domains with several fibrils at or close to their intersections.

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Faculty Opinions recommendation of Prestress in the extracellular matrix sensitizes latent TGF-β1 for activation.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.723349628.793501092 ◽

2014 ◽

Author(s):

Martin A Schwartz

Keyword(s):

Extracellular Matrix ◽

Tgf Β1

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Connexin expression by alveolar epithelial cells is regulated by extracellular matrix

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.2001.280.2.l191 ◽

2001 ◽

Vol 280 (2) ◽

pp. L191-L202 ◽

Cited By ~ 32

Author(s):

Yihe Guo ◽

Cara Martinez-Williams ◽

Clare E. Yellowley ◽

Henry J. Donahue ◽

D. Eugene Rannels

Keyword(s):

Extracellular Matrix ◽

Epithelial Cells ◽

Gap Junction ◽

Intercellular Communication ◽

Translational Regulation ◽

Alveolar Epithelial Cells ◽

Alveolar Type ◽

Type Ii ◽

Gap Junction Intercellular Communication ◽

Type Ii Cell

Extracellular matrix (ECM) proteins promote attachment, spreading, and differentiation of cultured alveolar type II epithelial cells. The present studies address the hypothesis that the ECM also regulates expression and function of gap junction proteins, connexins, in this cell population. Expression of cellular fibronectin and connexin (Cx) 43 increase in parallel during early type II cell culture as Cx26 expression declines. Gap junction intercellular communication is established over the same interval. Cells plated on a preformed, type II cell-derived, fibronectin-rich ECM demonstrate accelerated formation of gap junction plaques and elevated gap junction intercellular communication. These effects are blocked by antibodies against fibronectin, which cause redistribution of Cx43 protein from the plasma membrane to the cytoplasm. Conversely, cells cultured on a laminin-rich ECM, Matrigel, express low levels of Cx43 but high levels of Cx26, reflecting both transcriptional and translational regulation. Cx26 and Cx43 thus demonstrate reciprocal regulation by ECM constituents.

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Expression of alpha-smooth muscle actin, TGF-β1 and TGF-β type II receptor during connective tissue contraction

In Vitro Cellular & Developmental Biology - Animal ◽

10.1007/s11626-997-0112-4 ◽

1997 ◽

Vol 33 (8) ◽

pp. 622-627 ◽

Cited By ~ 6

Author(s):

M. Reza Ghassemifar ◽

Roy W. Tarnuzzer ◽

Nasser Chegini ◽

Erkki Tarpila ◽

Gregory S. Schultz ◽

...

Keyword(s):

Smooth Muscle ◽

Connective Tissue ◽

Smooth Muscle Actin ◽

Type Ii ◽

Muscle Actin ◽

Alpha Smooth Muscle Actin ◽

Tissue Contraction ◽

Tgf Β1

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Activation of transmembrane receptor tyrosine kinase DDR1-STAT3 cascade by extracellular matrix remodeling promotes liver metastatic colonization in uveal melanoma

Signal Transduction and Targeted Therapy ◽

10.1038/s41392-021-00563-x ◽

2021 ◽

Vol 6 (1) ◽

Author(s):

Wei Dai ◽

Shenglan Liu ◽

Shubo Wang ◽

Li Zhao ◽

Xiao Yang ◽

...

Keyword(s):

Extracellular Matrix ◽

Cancer Cells ◽

Uveal Melanoma ◽

Specific Inhibitor ◽

Type I ◽

Matrix Remodeling ◽

Metastatic Colonization ◽

Tgf Β1

AbstractColonization is believed a rate-limiting step of metastasis cascade. However, its underlying mechanism is not well understood. Uveal melanoma (UM), which is featured with single organ liver metastasis, may provide a simplified model for realizing the complicated colonization process. Because DDR1 was identified to be overexpressed in UM cell lines and specimens, and abundant pathological deposition of extracellular matrix collagen, a type of DDR1 ligand, was noted in the microenvironment of liver in metastatic patients with UM, we postulated the hypothesis that DDR1 and its ligand might ignite the interaction between UM cells and their surrounding niche of liver thereby conferring strengthened survival, proliferation, stemness and eventually promoting metastatic colonization in liver. We tested this hypothesis and found that DDR1 promoted these malignant cellular phenotypes and facilitated metastatic colonization of UM in liver. Mechanistically, UM cells secreted TGF-β1 which induced quiescent hepatic stellate cells (qHSCs) into activated HSCs (aHSCs) which secreted collagen type I. Such a remodeling of extracellular matrix, in turn, activated DDR1, strengthening survival through upregulating STAT3-dependent Mcl-1 expression, enhancing stemness via upregulating STAT3-dependent SOX2, and promoting clonogenicity in cancer cells. Targeting DDR1 by using 7rh, a specific inhibitor, repressed proliferation and survival in vitro and in vivo outgrowth. More importantly, targeting cancer cells by pharmacological inactivation of DDR1 or targeting microenvironmental TGF-β1-collagen I loop exhibited a prominent anti-metastasis effect in mice. In conclusion, targeting DDR1 signaling and TGF-β signaling may be a novel approach to diminish hepatic metastasis in UM.

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