Department of Pathology, Barbara Ann Karmanos Cancer Institute, Wayne State School of Medicine, Detroit, MI 48201, USA.
Biochem J. 2012 Feb 1;441(3):909-18. doi: 10.1042/BJ20111020.
The PDGF (platelet-derived growth factor) family members are potent mitogens for cells of mesenchymal origin and serve as important regulators of cell migration, survival, apoptosis and transformation. Tumour-derived PDGF ligands are thought to function in both autocrine and paracrine manners, activating receptors on tumour and surrounding stromal cells. PDGF-C and -D are secreted as latent dimers, unlike PDGF-A and -B. Cleavage of the CUB domain from the PDGF-C and -D dimers is required for their biological activity. At present, little is known about the proteolytic processing of PDGF-C, the rate-limiting step in the regulation of PDGF-C activity. In the present study we show that the breast carcinoma cell line MCF7, engineered to overexpress PDGF-C, produces proteases capable of cleaving PDGF-C to its active form. Increased PDGF-C expression enhances cell proliferation, anchorage-independent cell growth and tumour cell motility by autocrine signalling. In addition, MCF7-produced PDGF-C induces fibroblast cell migration in a paracrine manner. Interestingly, PDGF-C enhances tumour cell invasion in the presence of fibroblasts, suggesting a role for tumour-derived PDGF-C in tumour-stromal interactions. In the present study, we identify tPA (tissue plasminogen activator) and matriptase as major proteases for processing of PDGF-C in MCF7 cells. In in vitro studies, we also show that uPA (urokinase-type plasminogen activator) is able to process PDGF-C. Furthermore, by site-directed mutagenesis, we identify the cleavage site for these proteases in PDGF-C. Lastly, we provide evidence suggesting a two-step proteolytic processing of PDGF-C involving creation of a hemidimer, followed by GFD-D (growth factor domain dimer) generation.
血小板衍生生长因子（PDGF）家族成员是间充质来源细胞的有效有丝分裂原，是细胞迁移、存活、凋亡和转化的重要调节剂。肿瘤衍生的 PDGF 配体被认为以自分泌和旁分泌的方式发挥作用，在肿瘤和周围基质细胞上激活受体。PDGF-C 和 -D 作为潜伏的二聚体分泌，而 PDGF-A 和 -B 则不然。需要从 PDGF-C 和 -D 二聚体中裂解 CUB 结构域，才能发挥其生物学活性。目前，对于 PDGF-C 的蛋白水解加工知之甚少，而 PDGF-C 活性调节的限速步骤。在本研究中，我们表明，过表达 PDGF-C 的乳腺癌细胞系 MCF7 产生能够将 PDGF-C 切割成其活性形式的蛋白酶。增加 PDGF-C 的表达通过自分泌信号增强细胞增殖、锚定非依赖性细胞生长和肿瘤细胞迁移。此外，MCF7 产生的 PDGF-C 以旁分泌方式诱导成纤维细胞迁移。有趣的是，PDGF-C 在存在成纤维细胞的情况下增强肿瘤细胞侵袭，表明肿瘤衍生的 PDGF-C 在肿瘤-基质相互作用中起作用。在本研究中，我们确定 tPA（组织纤溶酶原激活物）和 matriptase 是 MCF7 细胞中 PDGF-C 加工的主要蛋白酶。在体外研究中，我们还表明 uPA（尿激酶型纤溶酶原激活物）能够处理 PDGF-C。此外，通过定点突变，我们确定了这些蛋白酶在 PDGF-C 中的裂解位点。最后，我们提供了证据表明 PDGF-C 的两步蛋白水解加工涉及创建半二聚体，然后是 GFD-D（生长因子域二聚体）生成。
