CRIP1 exacerbates osteoarthritis progression by recruiting UBE3A to induce the ubiquitination-mediated degradation of MFGE8

Osteoarthritis (OA) is a common degenerative joint disease. Cysteine-rich intestinal protein 1(CRIP1) is involved in many biological processes, but its role in OA remains unclear. OA-related datasets from the GEO database revealed that CRIP1 expression was upregulated in OA patient cartilage. This upregulation was also confirmed in our collected clinical samples and OA mice. CRIP1 knockdown increased the expression of collagen II and tissue inhibitor of metalloproteinases 3, but reduced the levels of matrix metalloproteinase 13 and phosphorylated p65 in IL-1β-induced primary mouse chondrocytes. In contrast, CRIP1 overexpression had the opposite effects. CRIP1 knockdown alleviated cartilage damage, reduced extracellular matrix (ECM) degradation, and inhibited p65 phosphorylation in OA mice. To investigate the CRIP1 mechanism in OA, proteins regulated by CRIP1 were screened by label-free quantitative proteomics, and the milk fat globule-epidermal growth factor 8 (MFGE8) protein was identified in the search for CRIP1-specific binding proteins in chondrocytes using immunoprecipitation/mass spectrometry. CRIP1 knockdown increased MFGE8 expression in OA mouse cartilage and IL-1β-induced chondrocytes. CRIP1 overexpression resulted in a shortened half-life of MFGE8 in chondrocytes. The proteasome inhibitor reversed the MFGE8 downregulation in CRIP1 overexpressing chondrocytes. Immunoprecipitation and rescue experiments verified that CRIP1 regulated MFGE8 degradation by promoting its binding to E3 ubiquitin-protein ligase 3A (UBE3A). Meanwhile, CRIP1 affected ECM degradation and NF-κB pathway through MFGE8. In summary, CRIP1 knockdown attenuated cartilage damage in OA mice and inhibited the ECM degradation and p65 activation in vivo and in vitro . This effect may be mediated by inhibiting the interaction between UBE3A and MFGE8.