Gaultier Alban, Cousin Hélène, Darribère Thierry, Alfandari Dominique
Department of Cell Biology, University of Virginia School of Medicine, Charlottesville, Virginia 22908, USA.
J Biol Chem. 2002 Jun 28;277(26):23336-44. doi: 10.1074/jbc.M201792200. Epub 2002 Apr 19.
ADAM13 is a member of the disintegrin and metalloprotease protein family that is expressed on cranial neural crest cells surface and is essential for their migration. ADAM13 is an active protease that can cleave fibronectin in vitro and remodel a fibronectin substrate in vivo. Using a recombinant secreted protein containing both disintegrin and cysteine-rich domains of ADAM13, we show that this "adhesive" region of the protein binds directly to fibronectin. Fibronectin fusion proteins corresponding to the various functional domains were used to define the second heparin-binding domain as the ADAM13 binding site. Mutation of the syndecan-binding site (PPRR --> PPTM) within this domain abolishes binding of the recombinant disintegrin and cysteine-rich domains of ADAM13. We further show that the adhesive disintegrin and cysteine-rich domain of ADAM13 can promote cell adhesion via beta(1) integrins. This adhesion requires integrin activation and can be prevented by antibodies to the cysteine-rich domain of ADAM13 and beta(1) integrin. Finally, wild type, but not the E/A mutant of ADAM13 metalloprotease domain, can be shed from the cell surface, releasing the metalloprotease domain associated with the disintegrin and cysteine-rich domains. This suggests that ADAM13 shedding may involve its own metalloprotease activity and that the released protease may interact with both integrins and extracellular matrix proteins.
ADAM13是解整合素和金属蛋白酶蛋白家族的成员，在颅神经嵴细胞表面表达，对其迁移至关重要。ADAM13是一种活性蛋白酶，可在体外切割纤连蛋白，并在体内重塑纤连蛋白底物。使用包含ADAM13解整合素和富含半胱氨酸结构域的重组分泌蛋白，我们发现该蛋白的这一“黏附”区域直接与纤连蛋白结合。对应于各种功能结构域的纤连蛋白融合蛋白被用于确定第二个肝素结合结构域为ADAM13结合位点。该结构域内syndecan结合位点（PPRR --> PPTM）的突变消除了ADAM13重组解整合素和富含半胱氨酸结构域的结合。我们进一步表明，ADAM13的黏附性解整合素和富含半胱氨酸结构域可通过β(1)整合素促进细胞黏附。这种黏附需要整合素激活，并且可被针对ADAM13富含半胱氨酸结构域和β(1)整合素的抗体阻断。最后，野生型而非ADAM13金属蛋白酶结构域的E/A突变体可从细胞表面脱落，释放与解整合素和富含半胱氨酸结构域相关的金属蛋白酶结构域。这表明ADAM13的脱落可能涉及其自身的金属蛋白酶活性，并且释放的蛋白酶可能与整合素和细胞外基质蛋白相互作用。
