Single-Cell Atlas of Common Variable Immunodeficiency reveals germinal center-associated epigenetic dysregulation in B cell responses

Common variable immunodeficiency (CVID), the most prevalent symptomatic primary immunodeficiency, is characterized by impaired terminal B-cell differentiation and defective antibody responses. Incomplete genetic penetrance and a wide range of phenotypic expressivity in CVID suggest the participation of additional pathogenic mechanisms. Monozygotic (MZ) twins discordant for CVID are uniquely valuable for studying the contribution of epigenetics to the disease. We used single-cell epigenomics and transcriptomics to create a cell census of naive-to-memory B cell differentiation in a pair of CVID-discordant MZ twins. Our analysis identifies DNA methylation, chromatin accessibility and transcriptional defects in memory B cells that mirror defective cell-cell communication defects following activation. These findings were validated in a cohort of CVID patients and healthy donors. Our findings provide a comprehensive multi-omics map of alterations in naive-to-memory B-cell transition in CVID and reveal links between the epigenome and immune cell cross-talk. Our resource, publicly available at the Human Cell Atlas, paves the way for future diagnosis and treatments of CVID patients.

Twins with Craniosynostosis: An Unidentified Variant/Twin Research: Kangaroo Care for Premature Twins; Developmental Delay in MZ Twins; Osteosarcoma in One Identical Twin; Controversies in Twin Pregnancy Management/Media Reports: Twin and Triplet Olympians; Twin’s Rescue from a Crocodile; Twin Pandas in Japan; Twin Surrogacy; Identical Twins in Pennsylvania

Craniosynostosis involves the early closure of one or more joints connecting the bones of an infant’s skull. A case of young monozygotic (MZ) male twins with an unidentified variant of this condition is described, followed by a summary of relevant published reports. This overview is followed by descriptions of a kangaroo care program for premature twins, developmental delay in an MZ twin pair, osteosarcoma in one MZ twin and controversial issues in the management of multiple pregnancies. Media reports of twin and triplet Olympic athletes, a twin’s rescue from a crocodile, the birth of twin pandas in Japan, a case of twin surrogacy and the birth of identical triplets are also presented.

Frailty and Processing Speed Performance at the Cusp of Midlife in CATSLife

Frailty is an important multi-domain measure of health status and aging. Processing speed (PS) performance may be predictive of later frailty among older adults, but the interrelation between frailty and PS at the cusp of mid-adulthood is unclear. Using data from the ongoing Colorado Adoption/Twin Study of Lifespan Behavioral Development and Cognitive Aging (CATSLife; N = 1213; Mean age = 33.22 years; SD = 5.0), we constructed a 24-item frailty sum score across anthropomorphic, objective health, and perceived health and engagement measures based on the Accumulation of Deficits model. PS was measured using the Colorado Perceptual Speed (CPS) and WAIS-III Digit Symbol (DS) tests. All mixed-effects regression models accounted for clustering among siblings, and covariates included sex, age, race, ethnicity, and educational attainment. Intraclass correlations (ICCs) [95% CI] for frailty among siblings, adjusted for sex and age, ranged from near zero for siblings in adoptive families, .13 [.08-.30] for nonadoptive siblings/fraternal (DZ) twins, and .44 [.40-.48] for identical (MZ) twins, suggesting possible heritable influences. Poorer PS performance was associated with higher frailty, but was significant for DS only (B: DS = -0.43, p =.005). Furthermore, the DS-frailty association was magnified by age (B: DSxAge = -0.06, p =.025), suggesting that the associations between processing speed and frailty may increase with age. These findings help elucidate the interrelationship between indicators of frailty and cognitive performance for adults approaching midlife, a salient and understudied period within lifespan development.

Nucleic Acid-Sensing and Interferon-Inducible Pathways Show Differential Methylation in MZ Twins Discordant for Lupus and Overexpression in Independent Lupus Samples: Implications for Pathogenic Mechanism and Drug Targeting

Systemic lupus erythematosus (SLE) is a chronic, multisystem, autoimmune inflammatory disease with genomic and non-genomic contributions to risk. We hypothesize that epigenetic factors are a significant contributor to SLE risk and may be informative for identifying pathogenic mechanisms and therapeutic targets. To test this hypothesis while controlling for genetic background, we performed an epigenome-wide analysis of DNA methylation in genomic DNA from whole blood in three pairs of female monozygotic (MZ) twins of European ancestry, discordant for SLE. Results were replicated on the same array in four cell types from a set of four Danish female MZ twin pairs discordant for SLE. Genes implicated by the epigenetic analyses were then evaluated in 10 independent SLE gene expression datasets from the Gene Expression Omnibus (GEO). There were 59 differentially methylated loci between unaffected and affected MZ twins in whole blood, including 11 novel loci. All but two of these loci were hypomethylated in the SLE twins relative to the unaffected twins. The genes harboring these hypomethylated loci exhibited increased expression in multiple independent datasets of SLE patients. This pattern was largely consistent regardless of disease activity, cell type, or renal tissue type. The genes proximal to CpGs exhibiting differential methylation (DM) in the SLE-discordant MZ twins and exhibiting differential expression (DE) in independent SLE GEO cohorts (DM-DE genes) clustered into two pathways: the nucleic acid-sensing pathway and the type I interferon pathway. The DM-DE genes were also informatically queried for potential gene–drug interactions, yielding a list of 41 drugs including a known SLE therapy. The DM-DE genes delineate two important biologic pathways that are not only reflective of the heterogeneity of SLE but may also correlate with distinct IFN responses that depend on the source, type, and location of nucleic acid molecules and the activated receptors in individual patients. Cell- and tissue-specific analyses will be critical to the understanding of genetic factors dysregulating the nucleic acid-sensing and IFN pathways and whether these factors could be appropriate targets for therapeutic intervention.

Spatial Similarity of MRI-Visible Perivascular Spaces in Healthy Young Adult Twins

Purpose This study aims to determine whether genetic factors affect the location of dilated perivascular spaces (dPVS) by comparing healthy young twins and non-twin (NT) siblings. Methods A total of 700 healthy young adult twins and NT siblings (138 monozygotic (MZ) twin pairs, 79 dizygotic (DZ) twin pairs, and 133 NT sibling pairs) were collected from the Human Connectome Project dataset. dPVS was automatically segmented and normalized to standard space. Then, spatial similarity indices (mean squared error [MSE], structural similarity [SSIM], and dice similarity [DS]) were calculated for dPVS in the basal ganglia (BGdPVS) and white matter (WMdPVS) between paired subjects before and after propensity score matching of dPVS volumes between groups. Within-pair correlations for the regional volumes of dVPS were also assessed using the intraclass correlation coefficient (ICC). Results The spatial similarity of dPVS was significantly higher in MZ twins (higher DS [median, 0.382 and 0.310] and SSIM [0.963 and 0.887] and lower MSE [0.005 and 0.005] for BGdPVS and WMdPVS, respectively) than DZ twins (DS [0.121 and 0.119], SSIM [0.941 and 0.868], and MSE [0.010 and 0.011]) and NT siblings (DS [0.106 and 0.097], SSIM [0.924 and 0.848], and MSE [0.016 and 0.017]). No significant difference was found between DZ twins and NT siblings. Similar results were found even after subjects were matched according to dPVS volume. Regional dPVS volumes were also more correlated within pairs in MZ twins than DZ twins and NT siblings. Conclusion Our results suggest that genetic factors affect the location of dPVS.

Identical twins carry a persistent epigenetic signature of early genome programming

Monozygotic (MZ) twins and higher-order multiples arise when a zygote splits during pre-implantation stages of development. The mechanisms underpinning this event have remained a mystery. Because MZ twinning rarely runs in families, the leading hypothesis is that it occurs at random. Here, we show that MZ twinning is strongly associated with a stable DNA methylation signature in adult somatic tissues. This signature spans regions near telomeres and centromeres, Polycomb-repressed regions and heterochromatin, genes involved in cell-adhesion, WNT signaling, cell fate, and putative human metastable epialleles. Our study also demonstrates a never-anticipated corollary: because identical twins keep a lifelong molecular signature, we can retrospectively diagnose if a person was conceived as monozygotic twin.

Examining the Vanishing Twin Hypothesis of Neural Tube Defects: Application of an Epigenetic Predictor for Monozygotic Twinning

Strong associations between neural tube defects (NTDs) and monozygotic (MZ) twinning have long been noted, and it has been suggested that NTD cases who do not present as MZ twins may be the survivors of MZ twinning events. We have recently shown that MZ twins carry a strong, distinctive DNA methylation signature and have developed an algorithm based on genomewide DNA methylation array data that distinguishes MZ twins from dizygotic twins and other relatives at well above chance level. We have applied this algorithm to published methylation data from five fetal tissues (placental chorionic villi, kidney, spinal cord, brain and muscle) collected from spina bifida cases (n = 22), anencephalic cases (n = 15) and controls (n = 19). We see no difference in signature between cases and controls, providing no support for a common etiological role of MZ twinning in NTDs. The strong associations therefore continue to await elucidation.

POS0361 DNA METHYLATION SIGNATURES CHARACTERIZE PSORIASIS AND PSORIATIC ARTHRITIS IN MONOZYGOTIC TWINS DISCORDANT FOR THE DISEASE

Background:Psoriatic disease is a chronic inflammatory disorder spanning from skin disease (psoriasis) to psoriatic arthritis (PsA). The genetic background is insufficient to explain disease onset as illustrated by not very informative Genome Wide Association Studies and monozygotic (MZ) twin studies recently performed. It is strongly assumed that epigenetics may contribute to disease susceptibility modulating gene expression. DNA methylation has been found involved in several autoimmune inflammatory rheumatic diseases. Here we have analysed the DNA methylation profile of a selected cohort of MZ twins discordant for psoriasis/PsA.Objectives:To identify the methylome associated with psoriasis and PsA in the peripheral blood of MZ twins discordant for these conditions.Methods:Peripheral blood from 7 couples of MZ twins discordant for psoriatic disease was collected and DNA extracted for a genome-wide evaluation of the DNA methylation profile, with the Infinium MethylationEPIC BeadChip. Minfi and the packages of the Bioconductor were used to analyse the data obtained. Quality control and exclusion criteria were applied to the raw data having a final number of 762.451 probes, which accounts for 88% of the total.Results:The approach first identified 2564 differentially methylated positions (DMPs; *p<0.005) with 19 genes potentially affected (with at least two DMPs within 1 kb of distance), including SMAD3 and SMARCA4/BRG1 involved in the Interferon and TGFβ pathways. Gene Ontology (GO) analysis of DMP-associated genes showed a significative enrichment (*p<0.005) in transcription factor binding, transcription corepressor and transcription coactivator activity, SMAD binding and histone -lysine-N-methyltransferase activity. To further validate the results, 5’-methylcytosine immunoprecipitation (MedIP) followed by Real Time PCR was performed to assess the methylation level of SMAD3 and SMARCA4/BRG1 promoters in the same cohort of MZ twins. We found significantly DNA methylation enrichment in SMARCA4/BRG1 promoter in psoriatic disease twins (p<0.05). SMAD3 and SMARCA4/BRG1 mRNA expression was also assessed to evaluate any inverse correlation with promoter methylation level, on the MZ cohort used for the EPIC array (n=4) and on a cohort of PsA/Ps patients (n=8) and appropriate healthy controls (n=3). Reduced mRNA expression (p<0.05) was demonstrated for SMARCA4/BRG1 (n=4). Conversely, no changes were found for SMAD3.Conclusion:We report the first DNA methylation approach in MZ twins discordant for psoriatic disease. We believe that the observed changes in SMAD3 and SMARCA/BRG1 genes may suggest an epigenetic imbalance of chromatin remodelling factors involved in inflammation pathways with a potential role in PsA/psoriasis immunopathogenesis.Disclosure of Interests:None declared

Genome-wide associations between alcohol consumption and blood DNA methylation: evidence from twin study

Aim: Alcohol intake alters DNA methylation profiles and methylation might mediate the association between alcohol and disease, but limited number of positive CpG sites repeatedly replicated. Materials & methods: In total, 57 monozygotic (MZ) twin pairs discordant for alcohol drinking from the Chinese National Twin Registry and 158 MZ and dizygotic twin pairs in the Swedish Adoption/Twin Study of Aging were evaluated. DNA methylation was detected using the Infinium HumanMethylation450 BeadChip. Results: Among candidate CpG sites, cg07326074 was significantly correlated with drinking after adjusting for covariates in MZ twins in both datasets but not in the entire sample or dizygotic twins. Conclusion: The hypermethylation of cg07326074, located in the tumor-promoting gene C16orf59, was associated with alcohol consumption.

DNA Methylation Variation Is Identified in Monozygotic Twins Discordant for Non-syndromic Cleft Lip and Palate

Non-syndromic cleft lip with or without cleft palate (NSCLP) is the most common craniofacial birth defect. The etiology of NSCLP is complex with multiple genes and environmental factors playing causal roles. Although studies have identified numerous genetic markers associated with NSCLP, the role of epigenetic variation remains relatively unexplored. Because of their identical DNA sequences, monozygotic (MZ) twins discordant for NSCLP are an ideal model for examining the potential contribution of DNA methylation to non-syndromic orofacial clefting. In this study, we compared the patterns of whole genome DNA methylation in six MZ twin pairs discordant for NSCLP. Differentially methylated positions (DMPs) and regions (DMRs) were identified in NSCLP candidate genes, including differential methylation in MAFB and ZEB2 in two independent MZ twin pairs. In addition to DNA methylation differences in NSCLP candidate genes, we found common differential methylation in genes belonging to the Hippo signaling pathway, implicating this mechanosensory pathway in the etiology of NSCLP. The results of this novel approach using MZ twins discordant for NSCLP suggests that differential methylation is one mechanism contributing to NSCLP, meriting future studies on the role of DNA methylation in familial and sporadic NSCLP.