Efficient immune responses orchestrated by CD4⁺ T lymphocytes require both lineage commitment and phenotypic flexibility, allowing the development of responses tailored to invading pathogens. With this project we aim at comprehensively investigating the role of DNA modifications and DNA-modifying enzymes in human T cell responses. Specifically, we want to address fundamental questions about the gene regulatory networks controlling the balance between commitment, phenotypic stability and plasticity of T cells. This will be performed by combining genome-wide analyses of DNA modifications and genetic manipulation of primary human T cells. Indeed, the stability of DNA methylation and its heritability across mitosis make it particularly apt to mediate the maintenance of transcriptional networks and cellular phenotypes. In the case of T cells, however, stability in the expression of subset-selective genes (notably cytokine genes) must be reconciled with mechanisms enabling plastic phenotypic changes in response to environmental clues. The recent discovery that methylated DNA can be dynamically modified, impacting gene expression directly or via erasure of DNA methylation, suggests its possible role in T cell plasticity. Our study will thoroughly describe dynamics in the methylation landscape in primary human T cells in response to specific pathogens and antigens, and assess the effects of methylation dynamics in T cell functions, leading to novel insights in immunity against pathogens and in disease.

Mast cell activation is involved in the response to a variety of pathogens and allergens, making these cells an important effector type not only in innate immunity but also in allergic reactions and asthma. In addition, alterations in the number, localization, and reactivity of mast cells are typical features of systemic mastocytosis, a myeloproliferative disorder characterized by an increase in mast cell burden.

Mast cell responses in health and disease. Mast cells act as important sentinels against danger signals in innate immunity, but alterations in the number, localization, and reactivity of mast cells are features of mast cell-related diseases such as systemic mastocytosis or allergy and asthma.

Multiple genetic and epigenetic mechanisms can contribute to the onset and severity of all types of mast cell-related diseases. Methylation of the genomic DNA is an epigenetic process in which a methyl group is covalently linked to a cytosine base in the DNA, and such modification in our genome has a critical impact in the control of gene expression. Indeed, enzymes involved in catalyzing this process are implicated in the pathogenesis of a variety of diseases and in regulating the function of immune cells. The enzyme TET2 is responsible for the oxidation of 5-methylcytosine (5mC) in genomic DNA to 5-hydroxymethylcytosine (5hmC), and such modification contributes to gene transcriptional regulation and in some cases to tumorigenic transformation (Leoni, 2015).

We found that overall levels of genomic 5hmC and the activity of the TET enzymes were crucial in regulating mast cell differentiation from hematopoietic progenitors (Montagner, 2016), while appropriate patterns of DNA methylation and sufficient levels of DNA methyltransferase enzyme activity were critical to restrain mast cell inflammatory responses in vivo and in vitro, in response to both acute and chronic stimulation (Leoni, 2017). In other words, mast cells with a normal hydroxymethylation/ methylation pattern can differentiate and respond adequately to stimuli from the environment, while mast cells with an altered methylation pattern show abnormal proliferation and respond with exaggerated responses to normal stimuli, leading to unrestrained inflammation.