ANKRD55 is a key regulator of T cell inflammation in multiple sclerosis
Chuyu Wu, Meiling Jiang, Xue Yang, Yixuan Liu, Bin Huang, Yi Guo, Runjing Cao, Zhihui Cui, Guozhen Deng, Weiyan Wang, Mengdi Guo, Zhiyong Lin, Jiahui Fan, Lin-ming Zhang, Lorenzo Di Cesare Mannelli, Tao Pang, Chenhui Wang, Cun-Jin Zhang
Chuyu Wu, Meiling Jiang, Xue Yang, Yixuan Liu, Bin Huang, Yi Guo, Runjing Cao, Zhihui Cui, Guozhen Deng, Weiyan Wang, Mengdi Guo, Zhiyong Lin, Jiahui Fan, Lin-ming Zhang, Lorenzo Di Cesare Mannelli, Tao Pang, Chenhui Wang, Cun-Jin Zhang
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Research Article Autoimmunity Immunology

ANKRD55 is a key regulator of T cell inflammation in multiple sclerosis

Abstract

Multiple sclerosis (MS) is a progressive, chronic, and highly disabling neuroinflammatory disorder characterized by demyelination and T cell–driven inflammation. Pathogenic T cells play a central role in MS, but effective therapeutic targeting remains challenging. Here, we identified ankyrin repeat domain–containing protein 55 (ANKRD55) as a key regulator of T cell function by single-cell transcriptomic analysis of cerebrospinal fluid and blood from MS patients. ANKRD55 was predominantly expressed in CD4+ T cells in both compartments. Genetic ablation of Ankrd55 led to a robustly reduced disease severity and neuroinflammation in experimental autoimmune encephalomyelitis (EAE), a widely used animal model for MS. Furthermore, T cell–specific deficiency of Ankrd55 significantly impaired Th1 polarization and Th17 differentiation, reducing EAE pathogenicity. Mechanistically, we found that Ankrd55 deficiency disrupted T cell receptor (TCR) signaling integrity. We demonstrated that ANKRD55 regulates the formation of the immune synapse, an essential prerequisite for TCR activation, by interacting with subunits of the chaperonin-containing TCP1 (CCT) complex and modulating its activity, enhancing its assembly by competing with CCT5 for binding to TCP1, CCT3, and CCT6. This facilitates proper microtubule organization and TCR activation. These findings establish ANKRD55 as a critical regulator of TCR signaling and highlight its therapeutic potential in pathogenic T cell–driven autoimmune diseases.

Authors

Chuyu Wu, Meiling Jiang, Xue Yang, Yixuan Liu, Bin Huang, Yi Guo, Runjing Cao, Zhihui Cui, Guozhen Deng, Weiyan Wang, Mengdi Guo, Zhiyong Lin, Jiahui Fan, Lin-ming Zhang, Lorenzo Di Cesare Mannelli, Tao Pang, Chenhui Wang, Cun-Jin Zhang

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Figure 6

ANKRD55 interacts with multiple subunits of the CCT complex required for TCR activation.

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ANKRD55 interacts with multiple subunits of the CCT complex required for...
(A) Overexpression of ANKRD55 and TCP1 in HEK293T cells followed by co-IP analysis to determine their interaction. (B) Protein extraction from Jurkat cells with subsequent co-IP assay to determine the interaction between ANKRD55 and TCP1. (C) PLA experiment in Jurkat cells to assess the interaction between ANKRD55 and TCP1. Cells were fixed and incubated with primary antibodies against ANKRD55 and TCP1, followed by PLA probe ligation and amplification. Red fluorescent puncta indicate close proximity (<40 nm) between ANKRD55 and TCP1, suggesting a direct or complex-mediated interaction. Nuclei were counterstained with DAPI (blue). Representative images are shown. Scale bars: 2 μm (top), 5 μm (bottom). (D) Immunofluorescent staining of Jurkat cells to analyze the colocalization of ANKRD55 and TCP1. Scale bars: 2 μm. (E–K) Co-IP assays were performed using HEK293T cells to investigate the interactions between ANKRD55 and individual CCT subunits, including CCT2, CCT3, CCT4, CCT5, CCT6, CCT7, and CCT8. (L) Immunofluorescent staining of Jurkat cells to assess colocalization between ANKRD55 and specific CCT subunits (CCT2, CCT3, CCT4, and CCT7). Scale bars: 2 μm (first 4 panels), 1 μm (fifth panel). (M) Immunofluorescent staining of Jurkat cells to visualize the subcellular localization of ANKRD55, TCP1, and pericentrin. Scale bars: 5 μm. (N–P) Immunoblotting analysis of TCP1 (N), CCT3 (O), and CCT6 (P) expression levels in Jurkat cells following ANKRD55 knockdown.