IRF2BP2-FOSL2 interaction regulates SOX9-mediated cartilage regeneration in osteoarthritis models
Hao Zhou, Tiantian Lu, Zhuoqun Gao, Can Cui, Zhiqiang Fang, Yu You, Jing Jiang, Jinjin Xu, Fang Wei, Jing Yu, Hu Zhou, Jinsong Li, Jian Luo, Lei Zhang, Yunyun Jin
Hao Zhou, Tiantian Lu, Zhuoqun Gao, Can Cui, Zhiqiang Fang, Yu You, Jing Jiang, Jinjin Xu, Fang Wei, Jing Yu, Hu Zhou, Jinsong Li, Jian Luo, Lei Zhang, Yunyun Jin
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Research Article Bone biology Cell biology

IRF2BP2-FOSL2 interaction regulates SOX9-mediated cartilage regeneration in osteoarthritis models

Abstract

Osteoarthritis (OA) is the most common degenerative joint disease. However, the molecular mechanisms underlying OA-induced cartilage degeneration remain elusive. Here, we identify IFN regulatory factor 2 binding protein 2 (IRF2BP2) as an important regulator of chondrocyte differentiation and articular cartilage regeneration. Irf2bp2 deficiency promoted chondrogenesis and inhibited chondrocyte terminal differentiation by transcriptionally upregulating the key chondrogenic marker SRY-box transcription factor 9 (SOX9), inducing its ectopic expression in the hypertrophic zone. Cartilage-specific KO of Irf2bp2 elevated SOX9 levels, attenuated OA progression in the destabilization of the medial meniscus (DMM) model, and enhanced cartilage regeneration and hyalinization in the microfracture model. Mechanistically, IRF2BP2 bound to and stabilized Fos-related antigen 2 (FOSL2), an activator protein 1 (AP-1) family member, thereby facilitating FOSL2/c-Jun complex formation and repressing SOX9 expression. Adeno-associated virus–mediated (AAV-mediated) FOSL2 overexpression reversed the protective phenotype observed in Irf2bp2-deficient mice. These findings establish the Hippo/IRF2BP2/FOSL2/SOX9 regulatory axis as critical for cartilage regeneration. Importantly, AAV-mediated IRF2BP2 knockdown, pharmacological targeting of the IRF2BP2/AP-1 axis using T-5224, and delivery of a peptide that disrupts IRF2BP2-FOSL2 binding via intra-articular injection all preserved cartilage integrity, offering potential therapeutic strategies for OA and cartilage regeneration.

Authors

Hao Zhou, Tiantian Lu, Zhuoqun Gao, Can Cui, Zhiqiang Fang, Yu You, Jing Jiang, Jinjin Xu, Fang Wei, Jing Yu, Hu Zhou, Jinsong Li, Jian Luo, Lei Zhang, Yunyun Jin

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Figure 4

IRF2BP2 interacts with and stabilizes FOSL2 to inhibit SOX9 transcriptional activation.

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IRF2BP2 interacts with and stabilizes FOSL2 to inhibit SOX9 transcriptio...
(A and B) Western blot analysis of SOX9 and COL2A1 in ATDC5 cells after IRF2BP2 overexpression (A) or knockdown (B). (C) Sox9 promoter–luciferase reporter in ATDC5 cells. (D) 4 × 48-p89-Luc activity in ATDC5 cells cotransfected with SOX9 and IRF2BP2. (E) A co-IP experiment of IRF2BP2 and FOSL2 was performed using HEK-293T cells. (F) Effects of IRF2BP2, FOSL2, and c-Jun on the Sox9 promoter reporter in ATDC5 cells. (G) Co-IP analysis of the effect of IRF2BP2 on the interaction between c-Jun and FOSL2. (H) ChIP-qPCR analysis of the effect of IRF2BP2 on FOSL2 binding to the promoter region of SOX9 in SW1353 cells. (I and J) Protein levels of FOSL2 were determined in ATDC5 cells after IRF2BP2 overexpression (I) or knockdown (J). (K) IF staining for FOSL2 in WT and Irf2bp2–/– mice at P1. Scale bar: 100 μm. (L) Quantification of FOSL2 staining in K (n = 4). (M) ATDC5 cells were transfected with vector or IRF2BP2, treated with CHX, and harvested at the indicated time points. The protein levels of endogenous FOSL2 were determined. (N) Quantitative analysis of the FOSL2 panels in M using ImageJ. (O) Control (sh-Luc) and IRF2BP2-knockdown ATDC5 cells were treated with CHX and harvested at the indicated time points. The protein levels of FOSL2 were determined. (P) Quantitative analysis of FOSL2 panels in O using ImageJ. In C, D, F, H, and L, data are shown as the mean ± SD. **P < 0.01, ***P < 0.001, and ****P < 0.0001, by 2-tailed, unpaired Student’s t test (C and L), 1-way ANOVA, followed by Tukey’s multiple-comparison test (F), and 2-way ANOVA, followed by Tukey’s multiple-comparison test (D and H). Dotted lines in K indicate the surface of articular cartilage. IB, immunoblot.