m6A methyltransferase METTL3 programs CD4+ T-cell activation and effector T-cell differentiation in systemic lupus erythematosus

Abstract

Background: Systemic lupus erythematosus (SLE) is an autoimmune disorder in which excessive CD4⁺ T-cell activation and imbalanced effector T-cell differentiation play critical roles. Recent studies have implied a potential association between posttranscriptional N6-methyladenosine (m⁶A) modification and CD4⁺ T-cell-mediated humoral immunity. However, how this biological process contributes to lupus is not well understood. In this work, we investigated the role of the m⁶A methyltransferase like 3 (METTL3) in CD4⁺ T-cell activation, differentiation, and SLE pathogenesis both in vitro and in vivo.

Methods: The expression of METTL3 was knocked down and METTL3 enzyme activity was inhibited using siRNA and catalytic inhibitor, respectively. In vivo evaluation of METTL3 inhibition on CD4⁺ T-cell activation, effector T-cell differentiation, and SLE pathogenesis was achieved using a sheep red blood cell (SRBC)-immunized mouse model and a chronic graft versus host disease (cGVHD) mouse model. RNA-seq was performed to identify pathways and gene signatures targeted by METTL3. m⁶A RNA-immunoprecipitation qPCR was applied to confirm the m⁶A modification of METTL3 targets.

Results: METTL3 was defective in the CD4⁺ T cells of SLE patients. METTL3 expression varied following CD4⁺ T-cell activation and effector T-cell differentiation in vitro. Pharmacological inhibition of METTL3 promoted the activation of CD4⁺ T cells and influenced the differentiation of effector T cells, predominantly Treg cells, in vivo. Moreover, METTL3 inhibition increased antibody production and aggravated the lupus-like phenotype in cGVHD mice. Further investigation revealed that catalytic inhibition of METTL3 reduced Foxp3 expression by enhancing Foxp3 mRNA decay in a m⁶A-dependent manner, hence suppressing Treg cell differentiation.

Conclusion: In summary, our findings demonstrated that METTL3 was required for stabilizing Foxp3 mRNA via m⁶A modification to maintain the Treg differentiation program. METTL3 inhibition contributed to the pathogenesis of SLE by participating in the activation of CD4⁺ T cells and imbalance of effector T-cell differentiation, which could serve as a potential target for therapeutic intervention in SLE.

Keywords: Autoimmune disorders; METTL3; N6-Methyladenosine; SLE; mRNA methylation.