ENAapplication/xmlftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/078/SRR26445378/SRR26445378_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/083/SRR26445383/SRR26445383_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/082/SRR26445382/SRR26445382_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/070/SRR26445370/SRR26445370_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/071/SRR26445371/SRR26445371_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/074/SRR26445374/SRR26445374_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/075/SRR26445375/SRR26445375_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/077/SRR26445377/SRR26445377_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/070/SRR26445370/SRR26445370_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/082/SRR26445382/SRR26445382_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/081/SRR26445381/SRR26445381_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/072/SRR26445372/SRR26445372_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/079/SRR26445379/SRR26445379_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/076/SRR26445376/SRR26445376_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/075/SRR26445375/SRR26445375_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/073/SRR26445373/SRR26445373_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/076/SRR26445376/SRR26445376_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/077/SRR26445377/SRR26445377_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/071/SRR26445371/SRR26445371_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/078/SRR26445378/SRR26445378_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/083/SRR26445383/SRR26445383_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/081/SRR26445381/SRR26445381_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/080/SRR26445380/SRR26445380_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/072/SRR26445372/SRR26445372_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/080/SRR26445380/SRR26445380_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/079/SRR26445379/SRR26445379_1.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/074/SRR26445374/SRR26445374_2.fastq.gzftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR264/073/SRR26445373/SRR26445373_1.fastq.gzprimaryOK200GenomicsSaarland Universityhttps://www.ebi.ac.uk/ena/browser/view/PRJNA1030038Homo sapiensxref:PubMed:39325839Epigenetic mechanisms stabilize gene expression patterns during CD8+ T cell differentiation. Although adoptive transfer of virus-specific T cells is clinically applied to reduce the risk of virus infection or reactivation in immunocompromised individuals, the DNA methylation pattern of virus-specific CD8+ T cells is largely unknown. Hence, we here performed whole-genome bisulfite sequencing of cytomegalovirus-specific human CD8+ T cells and found that they display a unique DNA methylation pattern consisting of 79 differentially methylated regions (DMRs) when compared to memory CD8+ T cells. Among the top demethylated DMRs in cytomegalovirus-specific CD8+ T cells was TBKBP1, coding for TBK-binding protein 1 that can interact with TANK-binding kinase 1 (TBK1) and mediate pro-inflammatory responses in innate immune cells downstream of intracellular virus sensing. Since TBKBP1 has not yet been reported in T cells, we aimed to unravel its role in virus-specific CD8+ T cells. TBKBP1 demethylation in terminal effector CD8+ T cells correlated with higher TBKBP1 expression at both mRNA and protein level, independent of alternative splicing of TBKBP1 transcripts. Notably, the distinct DNA methylation patterns in CD8+ T cell subsets was stable upon long-term in vitro culture. TBKBP1 overexpression resulted in enhanced TBK1 phosphorylation upon stimulation of CD8+ T cells and significantly improved their virus neutralization capacity. Collectively, our data demonstrate that TBKBP1 modulates virus-specific CD8+ T cell responses and could be exploited as therapeutic target to improve adoptive T cell therapies. Overall design: Peripheral blood mononuclear cells (PBMCs) from CMV-seropositive donors were were isolated, stimulated with CMV-pp65 overlapping peptide pool and subjected to flow cytometric sorting. Genomic DNA was isolated from sorted naive CD8+ T cells, interferon gamma-negative memory CD8+ T cells and interferon gamma-positive CD8+ T cells, and used for Bisulfite-Seq. Differentially methylated regions were identified by pairwise comparison of the samples with the software BSmooth.ENAThymus-Dependent Lymphocyte., 3110043L15Rik, cucumber mosaic virus CMV, Thymus-Dependent Lymphocytes, human being, Man (Taxonomy), Activity, T-Cells, ProSAPiP2, p32, Modern, Thymus-Dependent, Leu2, Prosapip2, T, Lymphocytes, cucumber mosaic virus, T-Lymphocyte, DNA methylation profiling, human, Cell, SINTBAD, Human, T Cells, Cytomegalovirus Infections, Homo sapiens, T Lymphocyte, T-Cell, CMV, Modern Man, Cells, T Cell, CMV Infections, cucumber mosaic cucumovirus CMV, CD8, MAL, T Lymphocytes, General activity, cucumber mosaic cucumovirus, Man, Thymus Dependent Lymphocytes, humans, LymphocyteThymus-Dependent Lymphocyte., 3110043L15Rik, cucumber mosaic virus CMV, Thymus-Dependent Lymphocytes, human being, Man (Taxonomy), Activity, T-Cells, ProSAPiP2, p32, Modern, Thymus-Dependent, Leu2, Prosapip2, T, Lymphocytes, cucumber mosaic virus, T-Lymphocyte, DNA methylation profiling, human, Cell, SINTBAD, Human, T Cells, Cytomegalovirus Infections, Homo sapiens, T Lymphocyte, T-Cell, CMV, Modern Man, Cells, T Cell, CMV Infections, cucumber mosaic cucumovirus CMV, CD8, MAL, T Lymphocytes, General activity, cucumber mosaic cucumovirus, Man, Thymus Dependent Lymphocytes, humans, LymphocytefalseDNA methylation profiling identifies TBKBP1 as potent amplifier of cytotoxic activity in CMV-specific human CD8+ T cellsDNA methylation profiling identifies TBKBP1 as potent amplifier of cytotoxic activity in CMV-specific human CD8+ T cells2025-09-242024-09-15PRJNA1030038GSE245832960639325839