Procollagen C-Proteinase Enhancers: In vivo Roles

描述（由申请人提供）：前胶原c -蛋白酶（pcp）切割前胶原Mil的c -基，产生形成ECM主要纤维成分的单体。两种PCP增强蛋白（PCOLCE1和2）不是蛋白酶，但它们以某种方式在体外将PCP活性提高了约10倍。PCOLCE1 c端结构域可以抑制肿瘤细胞分泌的基质金属蛋白酶（MMPs），并具有抗软骨血管生成的活性。PCOLCE1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成纤维细胞不依赖锚定生长。我们产生了PCOLCE1基因无等位基因的小鼠，以确定其在体内的作用。PCOLCE1 -/-小鼠可以存活和生育，但在骨骼和软组织中有严重的ECM扰动，因此确定PCOLCE1是骨强度和ECM完整性的决定因素。最近，我们成功地培养了PCOLCE2基因的零等位基因小鼠，并发现PCOLCE2 -/-小鼠既能存活又能生育。Pcolce2在软骨中的主要发育表达，加上Pcolce2 -/-小鼠的骨骼表型，表明Pcolce2在软骨内骨形成和关节炎发病中的作用。Pcolce和Pcolce2在心脏中均有高表达，Pcolce2在眼睛中也有高表达。我们建议充分表征成年和胚胎Pcolce-/-和Pcolce2-/-小鼠及其衍生细胞，以确定骨骼和软组织异常的分子基础和全面程度。Pcolce-/-和Pcolce2-/-小鼠杂交产生Pcolce/Pcolce2双纯合零后代，将消除Pcolce和Pcolce2之间可能存在的功能冗余所导致的任何残留活性。对Pcolce-/-、Pcolce2-/-和双空小鼠、胚胎和细胞的研究将有助于清楚地描述PCPE在哺乳动物发育、体内平衡和病理中的作用。基于我们实验室最近关于PCOLCE1和bmp1样pcp之间物理相互作用的证据，我们还提出了一些研究，以期在我们对pcp和PCOLCEs如何发挥作用的理解上有一个范式转变。

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.