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Integrative transcriptome and chromatin landscape analysis reveals distinct epigenetic regulations in human memory B cells.

ABSTRACT: Memory B cells (MBCs) are long-lived and produce high-affinity, generally, class-switched antibodies. Here, we use a multiparameter approach involving CD27 to segregate naïve B cells (NBC), IgD+ unswitched (unsw)MBCs and IgG+ or IgA+ class-switched (sw)MBCs from humans of different age, sex and race. Conserved antibody variable gene expression indicates that MBCs emerge through unbiased selection from NBCs. Integrative analyses of mRNAs, miRNAs, lncRNAs, chromatin accessibility and cis-regulatory elements uncover a core mRNA-ncRNA transcriptional signature shared by IgG+ and IgA+ swMBCs and distinct from NBCs, while unswMBCs display a transitional transcriptome. Some swMBC transcriptional signature loci are accessible but not expressed in NBCs. Profiling miRNAs reveals downregulated MIR181, and concomitantly upregulated MIR181 target genes such as RASSF6, TOX, TRERF1, TRPV3 and ROR?, in swMBCs. Finally, lncRNAs differentially expressed in swMBCs cluster proximal to the IgH chain locus on chromosome 14. Our findings thus provide new insights into MBC transcriptional programs and epigenetic regulation, opening new investigative avenues on these critical cell elements in human health and disease.

SUBMITTER: Moroney JB

PROVIDER: S-EPMC7595102 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature

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Integrative transcriptome and chromatin landscape analysis reveals distinct epigenetic regulations in human memory B cells.

Moroney Justin B JB Vasudev Anusha A Pertsemlidis Alexander A Zan Hong H Casali Paolo P

Nature communications 20201028 1

Memory B cells (MBCs) are long-lived and produce high-affinity, generally, class-switched antibodies. Here, we use a multiparameter approach involving CD27 to segregate naïve B cells (NBC), IgD+ unswitched (unsw)MBCs and IgG+ or IgA+ class-switched (sw)MBCs from humans of different age, sex and race. Conserved antibody variable gene expression indicates that MBCs emerge through unbiased selection from NBCs. Integrative analyses of mRNAs, miRNAs, lncRNAs, chromati ...[more]

PMID: 33116135

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Project description:Memory B cells (MBCs) are long-lived and rapidly respond to cognate antigen with production of high-affinity, generally, class-switched antibodies. Here, we have developed an integrative analysis of differentially expressed (DE) mRNAs, miRNAs, lncRNAs, chromatin profiles and cis-regulatory elements to determine how gene expression intersects with epigenetic regulatory elements in human MBCs. Using a multiparameter cell sorting approach, we isolated CD27-IgD+NBCs, CD27+IgD+unswMBCs, CD27+IgG+swMBCs and CD27+IgA+swMBCs as well as total CD27-NBCs and CD27+MBCs from peripheral blood of healthy human subjects of different age, sex and ethnic background. Total RNA was used to construct RNA-Seq libraries and subjected to next generation sequencing for in-depth analysis of B cell transcriptome, including mRNAs, miRNAs and lncRNAs, as well as Ig heavy chain (H) V(D)J, and Ig light chain (L) gene transcripts. Differential expression analysis identified shared mRNA, miRNA and lncRNA profiles that distinguished CD27+IgG+swMBCs and CD27+IgA+swMBCs from CD27-IgD+NBCs, and stratified CD27+IgD+unswMBCs between NBCs and swMBCs. Further, targeted integration using Ingenuity Pathway Analysis (IPA) outlined distinct signaling pathways in human MBCs, while the activity of TFs was inferred from global changes in transcriptional activation. ATAC-Seq was integrated with RNA-Seq to correlate DE RNAs with chromatin accessibility, thereby uncovering distinct profiles of cis-regulatory elements in human MBCs. A deep downregulation of MIR181 was concomitant with upregulation of this microRNA target genes, which accounted for 11 percent of the DE mRNAs in swMBCs, indicating an important role for MIR181 in MBC differentiation and/or maintenance. Further, lncRNA MIAT, a sponge of MIR181, was upregulated in MBCs and inversely correlated with MIR181a and MIR181b expression. This along with chromatin accessibility contributes downregulation of MIR181 and promotes MBC-specific gene expression. Overall, our findings provide evidence for overlapping layers of regulation, including chromatin remodeling, cis-regulatory elements and distinct sets of miRNAs and lncRNAs, which integrate to dictate gene expression profiles and activation pathways characteristic of human MBCs.

Project description:Metazoan genomes are spatially organized at multiple scales, from packaging of DNA around individual nucleosomes to segregation of whole chromosomes into distinct territories. At the intermediate scale of kilobases to megabases, which encompasses the sizes of genes, gene clusters and regulatory domains, the three-dimensional (3D) organization of DNA is implicated in multiple gene regulatory mechanisms, but understanding this organization remains a challenge. At this scale, the genome is partitioned into domains of different epigenetic states that are essential for regulating gene expression. Here we investigate the 3D organization of chromatin in different epigenetic states using super-resolution imaging. We classified genomic domains in Drosophila cells into transcriptionally active, inactive or Polycomb-repressed states, and observed distinct chromatin organizations for each state. All three types of chromatin domains exhibit power-law scaling between their physical sizes in 3D and their domain lengths, but each type has a distinct scaling exponent. Polycomb-repressed domains show the densest packing and most intriguing chromatin folding behaviour, in which chromatin packing density increases with domain length. Distinct from the self-similar organization displayed by transcriptionally active and inactive chromatin, the Polycomb-repressed domains are characterized by a high degree of chromatin intermixing within the domain. Moreover, compared to inactive domains, Polycomb-repressed domains spatially exclude neighbouring active chromatin to a much stronger degree. Computational modelling and knockdown experiments suggest that reversible chromatin interactions mediated by Polycomb-group proteins play an important role in these unique packaging properties of the repressed chromatin. Taken together, our super-resolution images reveal distinct chromatin packaging for different epigenetic states at the kilobase-to-megabase scale, a length scale that is directly relevant to genome regulation.

Dataset Information

Integrative transcriptome and chromatin landscape analysis reveals distinct epigenetic regulations in human memory B cells.

Publications

Integrative transcriptome and chromatin landscape analysis reveals distinct epigenetic regulations in human memory B cells.

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