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N6-methyladenosine (m6A) is a RNA modification that can regulate post-transcriptional processes including RNA stability, translation, splicing, and nuclear export. In CD4+ lymphocytes, m6A modifications have been demonstrated to play a role in early differentiation processes. The role of m6A in CD4+ T cell activation and effector function remains incompletely understood. To assess the role of m6A in CD4+ T lymphocyte activation and function, we assessed the transcriptome-wide m6A landscape of human primary CD4+ T cells by methylated RNA immunoprecipitation sequencing. Stimulation of the T cells impacted the m6A pattern of hundreds of transcripts including tumor necrosis factor (TNF). m6A methylation was increased on TNF messenger RNA (mRNA) after activation, predominantly in the 3' untranslated region of the transcript. Manipulation of m6A levels in primary human T cells, the directly affected the expression of TNF. Furthermore, we identified that the m6A reader protein YTHDF2 binds m6A-methylated TNF mRNA, and promotes its degradation. Taken together, this study demonstrates that TNF expression in CD4+ T lymphocytes is regulated via m6A and YTHDF2, thereby providing novel insight into the regulation of T cell effector functions., Competing Interests: Conflict of interest statement. None declared., (© The Author(s) 2024. Published by Oxford University Press on behalf of Society for Leukocyte Biology.)

T cell activation is critical for an effective immune response against pathogens. However, dysregulation contributes to the pathogenesis of autoimmune diseases, including Juvenile Idiopathic Arthritis (JIA). The molecular mechanisms underlying T cell activation are still incompletely understood. T cell activation promotes the acetylation of histone 3 at Lysine 27 (H3K27ac) at enhancer and promoter regions of proinflammatory cytokines, thereby increasing the expression of these genes which is essential for T cell function. Co-activators E1A binding protein P300 (P300) and CREB binding protein (CBP), collectively known as P300/CBP, are essential to facilitate H3K27 acetylation. Presently, the role of P300/CBP in human CD4+ T cells activation remains incompletely understood. To assess the function of P300/CBP in T cell activation and autoimmune disease, we utilized iCBP112, a selective inhibitor of P300/CBP, in T cells obtained from healthy controls and JIA patients. Treatment with iCBP112 suppressed T cell activation and cytokine signaling pathways, leading to reduced expression of many proinflammatory cytokines, including IL-2, IFN-γ, IL-4, and IL-17A. Moreover, P300/CBP inhibition in T cells derived from the inflamed synovium of JIA patients resulted in decreased expression of similar pathways and preferentially suppressed the expression of disease-associated genes. This study underscores the regulatory role of P300/CBP in regulating gene expression during T cell activation while offering potential insights into the pathogenesis of autoimmune diseases. Our findings indicate that P300/CBP inhibition could potentially be leveraged for the treatment of autoimmune diseases such as JIA in the future.

Respiratory syncytial virus (RSV) is the most prevalent cause of acute lower respiratory infection in young children. Currently, the first RSV vaccines are approved by the FDA. Recently, N6-methyladenosine (m 6 A) RNA methylation has been implicated in the regulation of the viral life cycle and replication of many viruses, including RSV. m 6 A methylation of RSV RNA has been demonstrated to promote replication and prevent anti-viral immune responses by the host. Whether m 6 A is also involved in viral entry and whether m 6 A can also affect RSV infection via different mechanisms than methylation of viral RNA is poorly understood. Here, we identify m 6 A reader YTH domain-containing protein 1 (YTHDC1) as a novel negative regulator of RSV infection. We demonstrate that YTHDC1 abrogates RSV infection by reducing the expression of RSV entry receptor CX3C motif chemokine receptor 1 (CX3CR1) on the cell surface of lung epithelial cells. Altogether, these data reveal a novel role for m 6 A methylation and YTHDC1 in the viral entry of RSV. These findings may contribute to the development of novel treatment options to control RSV infection.

T cell activation is a highly regulated process, modulated via the expression of various immune regulatory proteins including cytokines, surface receptors and co-stimulatory proteins. N 6 -methyladenosine (m 6 A) is an RNA modification that can directly regulate RNA expression levels and it is associated with various biological processes. However, the function of m 6 A in T cell activation remains incompletely understood. We identify m 6 A as a novel regulator of the expression of the CD40 ligand (CD40L) in human CD4 + lymphocytes. Manipulation of the m 6 A 'eraser' fat mass and obesity-associated protein (FTO) and m 6 A 'writer' protein methyltransferase-like 3 (METTL3) directly affects the expression of CD40L. The m 6 A 'reader' protein YT521-B homology domain family-2 (YTHDF2) is hypothesized to be able to recognize and bind m 6 A specific sequences on the CD40L mRNA and promotes its degradation. This study demonstrates that CD40L expression in human primary CD4 + T lymphocytes is regulated via m 6 A modifications, elucidating a new regulatory mechanism in CD4 + T cell activation that could possibly be leveraged in the future to modulate T cell responses in patients with immune-related diseases.