ANALYSIS OF THE REGULATORY NETWORK UNDERLYING THE DIFFERENTIATION OF ADAPTIVE IMMUNE RESPONSES

CD8+ T lymphocytes play a key role in the adaptive immune responses. Upon antigen recognition, they differentiate from naive to memory and effector cells. This fate commitment is guided by changes in gene expression patterns, which result in the acquisition of different cellular properties and functions. These changes are regulated by the activity of transcription factors, variations in the epigenetic landscapes and in chromatin structure. In this context, heterochromatin dynamic plays a key role in silencing phenotype-specific genes. So far, the mechanisms underlying heterochromatin and structural remodeling during T cell differentiation are poorly understood. A better understanding of the epigenetic regulation of lymphocyte differentiation may provide new tools for the improvement of immunotherapy design and efficacy. In this study, we used a multi-omic approach, combining the analysis of surface marker expression and gene expression profile at a single-cell (sc) level, to dissect tumor-infiltrating lymphocytes (1) heterogeneity and identify, through differential expression analysis, potential heterochromatin factors driving T cell differentiation. Among others, we found Lmnb1, coding for nuclear lamin B1, which is involved in the structural organization of heterochromatin inside the nucleus. To assess its role in CD8+ T cell differentiation, we generated Lmnb1 KO T cells. Through flow cytometry and sc-multi-omic analysis, we showed that lamin B1 regulates early CD8+ T cell differentiation and proliferation. Indeed, lamin B1 deficiency enhanced central memory T (TCM) cell accumulation while slowing down the acquisition of more differentiated transcriptional programs. Moreover, Lmnb1 disruption in antigen-specific T cells increased their ability to control tumor growth in vivo. To gain more insights into lamin B1-regulated regions, we explored the organization of laminaassociated domains (LADs) in CD8+ naïve T (TNaive) cells. We found that most lamin B1/chromatin interactions take place in chromatin intergenic regions, but we also found interactions at gene loci, including immune genes. In particular, we identified two categories of lamin B1/DNA interactions associated with different levels of activator (H3K4me3 and H3K9ac) and repressive (H3K9me3) histone marks, and inversely correlated with the transcriptional activity. In conclusion, we identify lamin B1 as a new regulatory factor of CD8+ T cell differentiation. We report its involvement in modulating proliferation and acquiring memory properties, both in vitro and in vivo. At the molecular level, we show that lamin B1 synergizes with the histone code to regulate gene expression profiles, including those involved in immune responses. A deeper understanding of the underlying mechanisms can improve our knowledge of immunological memory and exhaustion in cancer, with implications for cancer treatment and vaccination.