IMPACT OF MUTANT COMP ON SECRETION IN CHONDROCYTES

两种人类侏儒疾病，假性软骨发育不全（PSACH）和多发性骨骺发育不良（MED），是由软骨寡聚基质蛋白（COMP）突变引起的。在PSACH软骨细胞中，COMP的突变导致异常大的粗糙内质网（RER）的形成，具有独特的指纹外观。位于这些囊泡内的是COMP和IX型胶原蛋白。尽管这些细胞分泌COMP，但在肌腱成纤维细胞中没有发现这些异常的囊泡。本研究旨在阐明软骨细胞中COMP和IX型胶原异常保留的分子机制，而在肌腱成纤维细胞中却没有。最近的研究提出了以下假设：突变的COMP分子保留在软骨细胞的内质网中，因为III型重复序列不能正常折叠，从而导致形成非天然二硫键。然后这些分子在组装过程中与软骨细胞特异性RER分子和/或与天然IX型胶原和/或IX型胶原链相互作用。这种非天然复合物不能分泌，并导致软骨细胞内质网层状结构的积累。突变型COMP的分泌发生在成纤维细胞和去分化软骨细胞中，因为这些细胞不合成IX型胶原或不含软骨细胞特异性RER蛋白。具体目的是确定正常和突变的血栓反应蛋白III型重复序列的结构，确定这些重复序列中的二硫键和钙对结构的影响，并测试突变的COMP与IX型胶原和未折叠的IX型胶原链以及软骨细胞的REER蛋白的结合相互作用。

Two human dwarfing disorders, pseudoachondroplasia (PSACH) and multiple epiphyseal dysplasia (MED), are caused by mutations in cartilage oligomeric matrix protein (COMP). In PSACH chondrocytes, mutations in COMP lead to the formation of abnormally large rough endoplasmic reticulum (RER) with a unique fingerprint appearance. Located within these vesicles are COMP and type IX collagen. These abnormal vesicles are not found in tendon fibroblasts, although these cells secrete COMP. This proposal is aimed at elucidating the molecular mechanism behind the abnormal retention of COMP and type IX collagen in chondrocytes but not in tendon fibroblasts. The following hypotheses were developed from recent studies: Mutant COMP molecules are retained in the RER of chondrocytes because the type III repeats do not fold properly, which leads to the formation of non-native disulfide bonds. These molecules are then interacting with either chondrocyte specific RER molecules and/or with native type IX collagen and/or type IX collagen chains in the process of assembly. This non-native complex cannot be secreted and leads to the accumulation in lamellar structures within the RER of chondrocytes. Secretion of mutant COMP occurs in fibroblasts and dedifferentiated chondrocytes, because these cells do not synthesize type IX collagen or do not contain chondrocyte specific RER proteins. Specific aims are proposed to determine the structure of normal and mutant thrombospondin type III repeats, to define the disulfide linkages in these repeats and the effects of calcium on structure, and to test binding interactions of mutant COMP with type IX collagen and unfolded type IX collagen chains and with REER proteins of chondrocytes.