DYNAMIC RIBOSOMES IN OSTEOARTHRITIS

Purpose: Osteoarthritis (OA) is characterized by progressive loss and destruction of articular cartilage. Articular chondrocytes produce maintain the cartilage extracellular matrix (ECM), which requires continuous translation mRNAs ribosomes. Recent studies demonstrate that ribosomes can be dynamic heterogeneous. Post-transcriptional modifications (PTMs) ribosomal RNAs (rRNAs), such as 2’-O-ribose methylation (2’-O-Me), are a major source ribosome heterogeneity fine-tune translational activity. These accurately guided specific small nucleolar (snoRNAs). Here we hypothesized chondrocyte rRNA 2’-O-Me profiles altered in osteoarthritis contributes to OA progression. Methods: Non-OA human primary (HACs, 5 donors) were treated with end-stage synovial fluid (OA-SF) for 14 days. COL1A1 protein expression was evaluated immunoblotting. levels on measured quantified RiboMethSeq. CRISPR/Cas9 used knockdown snoRNAs SW1353 cells. CRISPR/Cas9-treated cells control. The efficiency genomic modification determined gDNA analysis (surveyor nuclease) mRNA (RT-qPCR). Cells transfected bicistronic IRES (Internal Ribosome Entry Site) reporters (CrPV IGR, P53, or HCV IRES) assess modus, UAA 0 frame, HIV-1 -1 PRF (programmed frameshifting) determine fidelity. Gene RT-qPCR. Proteomic performed LC-MS/MS. Results: We set up an OA-mimicking vitro model using non-OA HACs prolonged time OA-SF. In found decreased COL2A1 increased COL1A1. Immunoblotting confirmed increase type I collagen after OA-SF treatment (Figure 1A). To evaluate PTMs this model, profiling rRNAs. Five out 109 known sites significantly altered, including U14 5.8S 1B). relevance U14, targeted responsible SNORD71 snoRNA gene CRISPR/Cas9. As expected, led 1C) well 1D). Measurements function uncovered significant changes modus 1E) fidelity (data not shown) KD when compared CRISPR controls. Moreover, showed higher 1F), agreement our results obtained Simultaneously, unchanged 1G). Conclusions: Exposure from patients suffering knee fibrotic phenotype, deposition. This accompanied epitranscriptome RNAs, rRNA. addition, functional impairment SNORD71, mediates modification, affected expression. fact changed hints at preferential translation. demonstrated respond OA-associated their environment adjusting altering characteristics. impacts important components ECM. Thereby linked pathophysiology.