Characterization of the DNA Methylome during Human B-Cell Differentiation
Authors: Kulis Marta , Heath Simon , Castellano Giancarlo , Beekman Renée , Merkel Angelika , Raineri Emanuele , Esteve Anna , Queirós Ana C , Clot Guillem , Schuyler Ronald , Ecker Simone , Pancaldi Vera , Rico Daniel , Agueda Lidia , Blanc Julie , Richardson David , Clarke Laura , Datta Avik , Russiñol Nuria , Pascual Marien , Aguirre Xavier , Prosper Felipe , Alignani Diego , Paiva Bruno , Caron Gersende , Fest Thierry , Muench Marcus O. , Fomin Marina , Lee Seung-Tae , Wiemels Joseph L. , Valencia Alfonso , Gut Marta , Flicek Paul , Stunnenberg Hendrik G. , Siebert Reiner , Küppers Ralf , Gut Ivo G. , Campo Elias , Martín-Subero José I. 
Published: October 2019

Abstract
Introduction: Modulation of the DNA methylation landscape during cell differentiation is a well-established phenomenon. The B-cell lineage represents a paradigmatic cellular model to study the dynamic epigenome during cell development and specification because major B-cell maturation stages are well defined and display differential phenotypic and gene expression features. Furthermore, different B-cell subpopulations show different proliferation abilities, microenvironmental influences and life spans, providing a window of opportunity to study the epigenome in the context of multiple processes.
Methods: We performed whole-genome bisulfite sequencing (WGBS), high-density methylation microarrays and gene expression profiling of ten purified human B-cell subpopulations spanning the entire differentiation program, ranging from uncommitted progenitors to terminally-differentiated plasma cells.
Results: The results of both WGBS and methylation microarrays indicate that B-cell ontogenesis involves an extensive and gradual reconfiguration of the DNA methylome. We uncovered that non-CpG methylation at CpApC trinucleotides is present in progenitor cells and disappears upon B-cell commitment independently of CpG demethylation. CpG methylation, in contrast, changed extensively during the entire B-cell maturation program, with one quarter of all measured CpGs showing dynamic methylation. B-cell enhancers suffered more extensive methylation changes than promoter regions, especially in the early differentiation steps up to the germinal center B-cell (gcBC) stage, and their demethylation seemed to be mediated by binding of lineage-specific transcription factors. Enhancers with dynamic methylation were related to genes involved in a large B-cell network that showed high gene expression variability throughout differentiation. In highly proliferative gcBCs, we observed a shift of dynamic methylation from regulatory towards non-functional elements; gcBCs start to undergo global demethylation of late-replicating heterochromatic regions and methylation of polycomb-repressed regions. This signature becomes particularly extensive in long-lived memory B cells and plasma cells, indicating that these changes start in highly proliferative cells and then accumulate in non-proliferative cells with extended lifespan.
Conclusion: Our epigenomic analysis of the B-cell differentiation program extends our knowledge on how the DNA methylome is modulated during cell specification and maturation and offers a resource for researchers in the field, both at global and single gene levels.

Disclosures
No relevant conflicts of interest to declare.