Impaired chondrocyte U3 snoRNA expression in osteoarthritis impacts the chondrocyte protein translation apparatus

Abstract

Although pathways controlling ribosome activity have been described to regulate chondrocyte homeostasis in osteoarthritis, ribosome biogenesis in osteoarthritis is unexplored. We hypothesized that U3 snoRNA, a non-coding RNA involved in ribosomal RNA maturation, is critical for chondrocyte protein translation capacity in osteoarthritis. U3 snoRNA was one of a number of snoRNAs with decreased expression in osteoarthritic cartilage and osteoarthritic chondrocytes. OA synovial fluid impacted U3 snoRNA expression by affecting U3 snoRNA gene promoter activity, while BMP7 was able to increase its expression. Altering U3 snoRNA expression resulted in changes in chondrocyte phenotype. Interference with U3 snoRNA expression led to reduction of rRNA levels and translational capacity, whilst induced expression of U3 snoRNA was accompanied by increased 18S and 28S rRNA levels and elevated protein translation. Whole proteome analysis revealed a global impact of reduced U3 snoRNA expression on protein translational processes and inflammatory pathways. For the first time we demonstrate implications of a snoRNA in osteoarthritis chondrocyte biology and investigated its role in the chondrocyte differentiation status, rRNA levels and protein translational capacity. Osteoarthritis (OA) is a chronic debilitating joint disease that is strongly associated with ageing 1,2. OA involves pathological cellular processes in all joint structures and affects articular cartilage integrity, leading to dysfunctional joint articulation 2. During OA development and progression, the articular chondrocyte's phenotype changes 3-5 and presents with disturbed cellular homeostasis characterized by abnormal expression of (pre-) hypertrophic-[RUNX2 (runt-related transcription factor 2); COL10A1 (type X collagen)], catabolic-[ALPL (alkaline phosphatase); MMP13 (matrix metallopeptidase 13) and ADAMTS5 (a disintegrin and metalloproteinase with thrombospondin motifs 5) and inflammatory (COX2 (cyclooxygenase 2) and IL-6 (interleukin 6)] genes, while chondrogenic gene expression [SOX9 (SRY-box transcription factor 9); COL2A1 (type 2 collagen); ACAN (aggrecan) and NKX3-2 (NK3 homeobox 2)] is attenuated 3,4. The biomolecular processes that catalyze disturbances in the articular chondrocyte phenotype leading to OA are poorly understood, and it is expected that a comprehensive understanding of the avenues leading to disruption of articular chondrocyte homeostasis will provide important clues for future treatments. Chondrocytes are specialized secretory cells, enabling the synthesis and maintenance of the protein-rich cartilage extracellular matrix (ECM). Disturbances in chondrocyte protein translation in cartilage development and OA are connected to mTOR (mammalian target of rapamycin) activity 6 , endoplasmic reticulum stress 7 , unfolded protein response and apoptosis 8. These responses change the downstream translational activity of the

Ribosome dysfunction in osteoarthritis

“…Another key factor in the endoribonucleolytic processing of the 47S prerRNA is U3 snoRNA [ 18 ]. The expression of U3 snoRNA was reduced in human osteoarthritis cartilage and chondrocytes, as well as in murine joints in which experimental osteoarthritis [destabilization of the medial meniscus (DMM)] was induced [ 19 ▪▪ ]. Osteoarthritis-dependent inhibition of U3 snoRNA transcription was identified as one of the causes of reduced U3 snoRNA expression in osteoarthritis chondrocytes and resulted in a decrease of chondrocyte rRNA levels [ 19 ▪▪ ].…”

“…The finding that the mTORC1-mediated inhibition of 4E-BP1 precedes cartilage degeneration in rat osteoarthritis knees [ 43 ] strongly suggests its involvement in early osteoarthritis. In contrast to an overall increased level of chondrocyte protein translation activity, work from our group demonstrated that protein translation activity was reduced in chondrocytes isolated from end-stage knee osteoarthritis cartilage [ 19 ▪▪ ]. This was accompanied by lower levels of rRNA in osteoarthritis chondrocytes and chondrocytes treated with osteoarthritis synovial fluid.…”

Small-RNA Sequencing Reveals Altered Skeletal Muscle microRNAs and snoRNAs Signatures in Weanling Male Offspring from Mouse Dams Fed a Low Protein Diet during Lactation

“…In cartilage, we demonstrated the global impact of reduced U3 expression on protein translational processes and inflammatory pathways and found that altering U3 expression has a direct effect on rRNA expression and translational capacity of chondrocytes. We showed that the extracellular environment is capable of controlling cellular U3 levels, thereby tuning the cell’s capacity to generate mature rRNA species [ 110 ]. The increase in U3 in NL may be a compensatory method to protein restriction.…”

Small-RNA sequencing reveals altered skeletal muscle microRNAs and snoRNAs signatures in weanling male offspring from mouse dams fed a low protein diet during lactation

“…In cartilage, we have demonstrated the global impact of reduced U3 expression on protein translational processes and inflammatory pathways and found that altering U3 expression has a direct effect on rRNA expression and translational capacity of chondrocytes. We showed that the extracellular environment is capable of controlling cellular U3 levels, thereby tuning the cell’s capacity to generate mature rRNA species (109). The increase in U3 in NL may be a compensatory method to protein restriction.…”