Procollagen C-Proteinase Enhancers: In vivo Roles

描述（由申请人提供）：前胶原C-蛋白酶（PCP）裂解前胶原MII的C-前肽，产生形成ECM主要纤维组分的单体。两种PCP增强蛋白（PCOLCE 1和2）不是蛋白酶，但它们在体外以某种方式增强PCP活性约10倍。PCOLCE 1 C-末端结构域可以抑制肿瘤细胞分泌的基质金属蛋白酶（MMP），并提供从软骨分离的抗血管生成活性。PCOLCE 1也可能在生长控制中发挥作用，因为同源基因的破坏导致培养的成纤维细胞的锚定非依赖性生长。我们已经产生了小鼠与无效等位基因的PCOLCE 1基因PColce，以确定其在体内的作用。PCOLCE-/-小鼠是可行的和可生育的，但在骨和软组织中具有深刻的ECM扰动，因此将PCOLCE 1鉴定为骨强度和ECM完整性的决定因素。最近，我们成功地产生了具有PCOLCE 2基因Pcolce 2空等位基因的小鼠，并发现Pcolce 2-/-小鼠既能存活又能生育。Pcolce 2在软骨中的主要发育表达，加上Pcolce-/-小鼠的骨骼表型，表明Pcolce 2在软骨内骨形成和关节炎发病机制中的作用。Pcolce和Pcolce 2在心脏中的表达水平特别高，并且Pcolce 2在眼睛中也以高水平表达。我们建议充分表征成年和胚胎Pcolce-/-和Pcolce 2-/-小鼠和衍生细胞，以确定骨骼和软组织异常的分子基础和完整程度。Pcolce-/-和Pcolce 2-/-小鼠杂交以产生Pcolce/Pcolce 2双纯合无效后代将去除由于Pcolce和Pcolce 2之间可能的功能冗余而导致的任何残余活性。对PColce-/-、PColce 2-/-和双缺陷小鼠、胚胎和细胞的研究将使PCPE在哺乳动物发育、体内平衡和病理学中的作用得以清晰描绘。根据我们实验室最近的PCOLCE 1和BMP 1样PCP之间的物理相互作用的证据，我们还提出了一些研究，这些研究有望使我们对PCP和PCOLCEs功能的理解发生范式转变。

DESCRIPTION (provided by applicant): Procollagen C-proteinases (PCPs) cleave the C-propetides of procollagens Mil, to yield monomers that form the major fibrous components of ECM. The two PCP enhancer proteins (PCOLCE1 and 2) are not proteinases, yet they somehow enhance PCP activities approximately 10-fold in vitro. The PCOLCE1 C-terminal domain can inhibit matrix metalloproteinases (MMPs) secreted by tumor cells, and provides an anti-angiogenic activity isolated from cartilage. PCOLCE1 may also play a role in growth control, as disruption of the cognate gene results in anchorage-independent growth in cultured fibroblasts. We have generated mice with null alleles for the PCOLCE1 gene Pcolce, to ascertain its in vivo roles. Pcolce-/- mice are viable and fertile, but have profound ECM perturbations in bone and soft tissues, thus identifying PCOLCE1 as a determinant of bone strength and ECM integrity. More recently, we have succeeded in generating mice with null-alleles for the PCOLCE2 gene, Pcolce2, and have found Pcolce2-/- mice to be both viable and fertile. Predominant developmental expression of Pcolce2 in cartilage, plus the skeletal phenotype of Pcolce-/- mice, suggest role(s) for Pcolce2 in endochondral bone formation and in pathogenesis of arthridities. Both Pcolce and Pcolce2 are notable for particularly high expression levels in heart and Pcolce2 is also expressed at high levels in eye. We propose fully characterizing adult and embryo Pcolce-/- and Pcolce2-/- mice and derived cells, to determine the molecular bases and full extents of skeletal and soft tissue abnomalities. Crosses of Pcolce-/- and Pcolce2-/- mice to produce Pcolce/Pcolce2 doubly homozygous null progeny will remove any residual activity due to possible functional redundancy between Pcolce and Pcolce2. Studies of Pcolce-/-, Pcolce2-/- and doubly null mice, embryos and cells will enable clear delineation of PCPE roles in mammalian development, homeostasis and pathology. Based on recent evidience in our lab of physical interactions between PCOLCE1 and BMP1-like PCPs, we also propose studies that promise a paradigmatic shift in our understanding of how PCPs and PCOLCEs function.