Abstract
Osteoarthritis is a debilitating disease of the joint that affects over 230 million people worldwide. Currently there are no treatments that slow the progression of this disease. For these reasons, new biological treatment options are currently being explored. Inorganic polyphosphates are naturally occurring biological molecules that have an anabolic effect on chondrocytes grown in vitro in the presence of Ca2+. We hypothesized that when examining significant changes in protein phosphorylation, key candidates would emerge that could help to elucidate the anabolic effects of polyphosphate on chondrocytes. Therefore, we conducted a large-scale quantitative proteomic and phosphoproteomic study of bovine primary articular chondrocytes after 30-minute treatment with inorganic polyphosphate and Ca2+. Mass spectrometry identified more than 6000 phosphorylation sites on ∼1600 chondrocyte phosphoproteins while proteomic analysis detected approximately 4100 proteins. Analysis of the data revealed a swift and dynamic response to polyphosphate after 30 minutes. What emerged from the list of proteins most affected by the treatment were proteins with key roles in chondrogenesis including TNC, IGFBP-5, and CTGF, indicating that polyphosphate plays an important role in chondrocyte metabolism. This phosphoproteome serves as a meaningful resource to help elucidate the molecular events that contribute to extracellular matrix production in cartilage.
Competing Interest Statement
The authors have declared no competing interest.
Abbreviations
- ACN
- acetonitrile
- CTGF
- connective tissue growth factor
- EIF4B
- eukaryotic translation initiation factor 4B
- FDR
- false discovery rate
- GSEA
- gene set enrichment analysis
- HILIC
- hydrophilic interaction liquid chromatography
- IGFBP5
- insulin-like growth factor binding protein 5
- MS
- mass spectrometry
- PCA
- principal component analysis
- polyP-22
- polyP with chain length of 22 phosphate residues
- polyP-45
- polyP with chain length of 45 phosphate residues
- polyP, PP
- polyphosphate
- RPS6KA1
- ribosomal protein S6 kinase alpha-1
- SRSF6
- serine/arginine-rich splicing factor 6
- TFA
- trifluoroacetic acid
- TiO2
- titanium dioxide
- TNC
- tenascin C