Gene activation precedes DNA demethylation in response to infection in human dendritic cells Alain Pacis, Florence Mailhot-Leonard, Ludovic Tailleux, Haley E Randolph, Vania Yotova, Anne Dumaine, Jean-Christophe Grenier, Luis B Barreiro doi: https://doi.org/10.1101/358531 This article is a preprint and has not been peer-reviewed [what does this mean?]. DNA methylation is considered to be a relatively stable epigenetic mark. Yet, a growing body of evidence indicates that DNA methylation levels can change rapidly, for example, in innate immune cells facing an infectious agent. Nevertheless, the causal relationship between changes in DNA methylation and gene expression during infection remains to be elucidated. Here, we generated time-course data on DNA methylation, gene expression, and chromatin accessibility patterns during infection of human dendritic cells with Mycobacterium tuberculosis. We found that the immune response to infection is accompanied by active demethylation of thousands of CpG sites overlapping distal enhancer elements. However, virtually all changes in gene expression in response to infection occur prior to detectable changes in DNA methylation, indicating that the observed losses in methylation are a downstream consequence of transcriptional activation. Footprinting analysis revealed that immune-related transcription factors (TF), such as NF-κB/Rel, are recruited to enhancer elements prior to the observed losses in methylation, suggesting that DNA demethylation is mediated by TF binding to cis-acting elements. Collectively, our results show that DNA demethylation is not required for the establishment of the core regulatory program engaged upon infection. Copyright The copyright holder for this preprint is the author/funder. It is made available under a CC-BY-NC-ND 4.0 International license.