Leitinger Birgit, Kwan Alvin P L
Division of Biomedical Sciences, Imperial College London, South Kensington Campus, London SW7 2AZ, UK.
Matrix Biol. 2006 Aug;25(6):355-64. doi: 10.1016/j.matbio.2006.05.006. Epub 2006 May 26.
During endochondral ossification, collagen X is deposited in the hypertrophic zone of the growth plate. Our previous results have shown that collagen X is capable of interacting directly with chondrocytes, primarily via integrin alpha2beta1. In this study, we determined whether collagen X could also interact with the non-integrin collagen receptors, discoidin domain receptors (DDRs), DDR1 or DDR2. The widely expressed DDRs are receptor tyrosine kinases that are activated by a number of different collagen types. Collagen X was found to be a much better ligand for DDR2 than for DDR1. Collagen X bound to the DDR2 extracellular domain with high affinity and stimulated DDR2 autophosphorylation, the first step in transmembrane signalling. Expression of DDR2 in the epiphyseal plate was confirmed by RT-PCR and immunohistochemistry. The spatial expression of DDR2 in the hypertrophic zone of the growth plate is consistent with a physiological interaction of DDR2 with collagen X. Surprisingly, the discoidin domain of DDR2, which fully contains the binding sites for the fibrillar collagens I and II, was not sufficient for collagen X binding. The nature of the DDR2 binding site(s) within collagen X was further analysed. In addition to a collagenous domain, collagen X contains a C-terminal NC1 domain. DDR2 was found to recognise the triple-helical region of collagen X as well as the NC1 domain. Binding to the collagenous region was dependent on the triple-helical conformation. DDR2 autophosphorylation was induced by the collagen X triple-helical region but not the NC1 domain, indicating that the triple-helical region of collagen X contains a specific DDR2 binding site that is capable of receptor activation. Our study is the first to describe a non-fibrillar collagen ligand for DDR2 and will form the basis for further studies into the biological function of collagen X during endochondral ossification.
在软骨内骨化过程中，Ⅹ型胶原蛋白沉积在生长板的肥大带。我们之前的研究结果表明，Ⅹ型胶原蛋白能够主要通过整合素α2β1直接与软骨细胞相互作用。在本研究中，我们确定Ⅹ型胶原蛋白是否也能与非整合素胶原蛋白受体盘状结构域受体（DDRs）、DDR1或DDR2相互作用。广泛表达的DDRs是受体酪氨酸激酶，可被多种不同类型的胶原蛋白激活。研究发现，Ⅹ型胶原蛋白对DDR2而言是比DDR1更好的配体。Ⅹ型胶原蛋白以高亲和力结合到DDR2的胞外结构域，并刺激DDR2自身磷酸化，这是跨膜信号传导的第一步。通过逆转录聚合酶链反应（RT-PCR）和免疫组织化学证实了DDR2在骨骺板中的表达。DDR2在生长板肥大带的空间表达与DDR2与Ⅹ型胶原蛋白的生理相互作用一致。令人惊讶的是，完全包含与Ⅰ型和Ⅱ型纤维状胶原蛋白结合位点的DDR2盘状结构域不足以与Ⅹ型胶原蛋白结合。进一步分析了Ⅹ型胶原蛋白内DDR2结合位点的性质。除了一个胶原结构域外，Ⅹ型胶原蛋白还包含一个C端NC1结构域。发现DDR2可识别Ⅹ型胶原蛋白的三螺旋区域以及NC1结构域。与胶原区域的结合依赖于三螺旋构象。Ⅹ型胶原蛋白的三螺旋区域而非NC1结构域诱导DDR2自身磷酸化，这表明Ⅹ型胶原蛋白的三螺旋区域包含一个能够激活受体的特定DDR2结合位点。我们的研究首次描述了DDR2的一种非纤维状胶原蛋白配体，并将为进一步研究Ⅹ型胶原蛋白在软骨内骨化过程中的生物学功能奠定基础。
