Elsevier

Experimental Cell Research

Volume 252, Issue 2, 1 November 1999, Pages 423-431
Experimental Cell Research

Regular Article
Disintegrin-like/Cysteine-Rich Region of ADAM 12 Is an Active Cell Adhesion Domain

https://doi.org/10.1006/excr.1999.4632Get rights and content

Abstract

ADAM (a disintegrin and metalloprotease) proteins contain structural homology to the P-III class of snake venom metalloproteases (SVMPs) and are postulated to function, by analogy to these SMVPs, as cell adhesion molecules. ADAM 12 has been implicated in fusion of myoblasts, but its mechanism of action is not known. Instead of the RGD-like cell-binding motif present in SVMP disintegrins, the disintegrin domain of ADAM 12 contains a unique SNS sequence and therefore its adhesive potential has been controversial. In this report we demonstrate that the disintegrin-like/cysteine-rich (DC) domain of ADAM 12 constitutes a functional cell adhesion domain. We have expressed the DC domain of mouse ADAM 12 in insect cells and shown that the recombinant protein supported adhesion of C2C12 myoblasts and NIH 3T3 fibroblasts in a divalent cation-dependent manner. A sulfhydryl-specific biotinylation reagent revealed, however, that the overall conformation and flexibility of the cell-binding region of ADAM 12 DC domain may be significantly different from those of the SVMP disintegrins. Moreover, the disulfide bond structure of the DC domain was critical for its function, as incubation of the recombinant protein with reducing agents abolished subsequent cell adhesion. Recombinant DC bound to C2C12 cells with high affinity (KD ≈ 0.10 μM, total number of binding sites n ≈ 4.6 × 105/cell). Adhesive properties of the DC domain of ADAM 12 produced in insect cells were further confirmed by cell surface binding of the DC domain expressed in C2C12 cells and secreted to the medium, consistent with the role of ADAM 12 in cell–cell interactions and myoblast fusion.

References (32)

Cited by (52)

  • A disintegrin and metalloproteinase-12 (ADAM12): Function, roles in disease progression, and clinical implications

    2013, Biochimica et Biophysica Acta - General Subjects
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    This binding to C2C12 myoblasts proceeds via the disintegrin-like and cysteine-rich domains of ADAM12. Although they share structural homology with the P-III class of snake venom metalloproteases (SVMPs) [23], they lack the commonly associated RGD integrin-binding motif responsible for binding to structures in the vasculature [24]. Iba et al., further observed that ADAM12 binds to primary murine osteoblasts, fibroblastic cells, osteoblastic cells, and myoblastic cells during the well plate assay in which these cells were seeded to the pre-attached rADAM12 [25].

  • Drosophila metalloproteases in development and differentiation: The role of ADAM proteins and their relatives

    2011, European Journal of Cell Biology
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    While the prodomain is considered to be involved in protein folding and in the regulation of activity (Suzuki et al., 1998), the intracellular domain is apparently required for proper trafficking of these proteins (Cao et al., 2002). The disintegrin-like domain, on the other hand, is known to interact with integrins, thereby mediating cell adhesion (Zolkiewska, 1999) and the ACR domain was reported to be involved in homo-oligomerization (Meyer et al., 2010). With respect to Kuzbanian, the zygotic loss of the active form of the protein results in a dramatic excess of cardiomyocytes in the Drosophila heart, which affects all subtypes of these cells, e.g., the tinman and the seven-up expressing cells.

  • The disintegrin and metalloprotease Meltrin from Drosophila forms oligomers via its protein binding domain and is regulated by the homeobox protein VND during embryonic development

    2010, Insect Biochemistry and Molecular Biology
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    The disintegrin-like, cysteine rich and EGF-like domains on the other hand are considered to be the site of protein-protein interaction. In this context especially the disintegrin-like domain is known to interact with integrins and thereby mediates cell adhesion (Zolkiewska, 1999). In mice, the ADAM protein Meltrin β was shown to be expressed in the peripheral nervous system, skeletal muscles, bones and in the embryonic heart (Kurisaki et al., 1998; Yagami-Hiromasa et al., 1995).

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Correspondence and reprint requests may be addressed to the author at Department of Biochemistry, Kansas State University, 104 Willard Hall, Manhattan, KS 66506. Fax: (785) 532-7278. E-mail: [email protected].

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