rnai reduces expression and intracellular retention of mutant cartilage oligomeric matrix proteinrnai减少表达和突变软骨寡聚基质蛋白的细胞内保留.pdfVIP

RNAi Reduces Expression and Intracellular Retention of Mutant Cartilage Oligomeric Matrix Protein 1 1 1 1 1 Karen L. Posey *, Peiman Liu , Huiqiu R. Wang , Alka C. Veerisetty , Joseph L. Alcorn , Jacqueline T. Hecht1,2 1 Department of Pediatrics, University of Texas Medical School at Houston, Houston, Texas, United States of America, 2 Shriners Hospital for Children, Houston, Texas, United States of America Abstract Mutations in cartilage oligomeric matrix protein (COMP), a large extracellular glycoprotein expressed in musculoskeletal tissues, cause two skeletal dysplasias, pseudoachondroplasia and multiple epiphyseal dysplasia. These mutations lead to massive intracellular retention of COMP, chondrocyte death and loss of growth plate chondrocytes that are necessary for linear growth. In contrast, COMP null mice have only minor growth plate abnormalities, normal growth and longevity. This suggests that reducing mutant and wild-type COMP expression in chondrocytes may prevent the toxic cellular phenotype causing the skeletal dysplasias. We tested this hypothesis using RNA interference to reduce steady state levels of COMP mRNA. A panel of shRNAs directed against COMP was tested. One shRNA (3B) reduced endogenous and recombinant COMP mRNA dramatically, regardless of expression levels. The activity of the shRNA against COMP mRNA was maintained for up to 10 weeks. We also demonstrate that this treatment reduced ER stress. Moreover, we show that reducing steady state levels of COMP mRNA alleviates intracellular retention of other extracellular matrix proteins associated with the pseudoachon- droplasia cellular pathology. These findings are a proof of principle and the foundation for the development of a th