Genomic Imprinting Research Articles

Distinct melanocyte subpopulations defined by stochastic expression of proliferation or maturation programs enable a rapid and sustainable pigmentation response.

The ultraviolet (UV) radiation triggers a pigmentation response in human skin, wherein, melanocytes rapidly activate divergent maturation and proliferation programs. Using single-cell sequencing, we demonstrate that these 2 programs are segregated in distinct subpopulations in melanocytes of human and zebrafish skin. The coexistence of these 2 cell states in cultured melanocytes suggests possible cell autonomy. Luria-Delbrück fluctuation test reveals that the initial establishment of these states is stochastic. Tracking of pigmenting cells ascertains that the stochastically acquired state is faithfully propagated in the progeny. A systemic approach combining single-cell multi-omics (RNA+ATAC) coupled to enhancer mapping with H3K27 acetylation successfully identified state-specific transcriptional networks. This comprehensive analysis led to the construction of a gene regulatory network (GRN) that under the influence of noise, establishes a bistable system of pigmentation and proliferation at the population level. This GRN recapitulates melanocyte behaviour in response to external cues that reinforce either of the states. Our work highlights that inherent stochasticity within melanocytes establishes dedicated states, and the mature state is sustained by selective enhancers mark through histone acetylation. While the initial cue triggers a proliferation response, the continued signal activates and maintains the pigmenting subpopulation via epigenetic imprinting. Thereby our study provides the basis of coexistence of distinct populations which ensures effective pigmentation response while preserving the self-renewal capacity.

Widespread natural selection on metabolite levels in humans.

Natural selection acts ubiquitously on complex human traits, predominantly constraining the occurrence of extreme phenotypes (stabilizing selection). These constraints propagate to DNA sequence variants associated with traits under selection. The genetic signatures of such evolutionary events can thus be detected via combining effect size estimates from genetic association studies and the corresponding allele frequencies. Although this approach has been successfully applied to high-level traits, the prevalence and mode of selection acting on molecular traits remain poorly understood. Here, we estimate the action of natural selection on genetic variants associated with metabolite levels, an important layer of molecular traits. By leveraging summary statistics of published genome-wide association studies with large sample sizes, we find strong evidence of stabilizing selection for 15 out of 97 plasma metabolites, with nonessential amino acids displaying especially strong selection signatures. Mendelian randomization analysis reveals that metabolites under stronger stabilizing selection display larger effects on a range of clinically relevant complex traits, suggesting that maintaining a disease-free profile may be an important source of selective constraints on the metabolome. Metabolites under strong stabilizing selection in humans are also more conserved in their concentrations among diverse mammalian species, suggesting shared selective forces across micro- and macroevolutionary timescales. Overall, this study demonstrates that variation in metabolite levels among humans is frequently shaped by natural selection and this may act through their causal impact on disease susceptibility.

The Genetic Puzzle of Cerebral Palsy: Results of a Monocentric Study

BackgroundCerebral palsy (CP) is the most frequent cause of motor impairment in children. Although perinatal asphyxia was long considered to be the leading cause of CP, recent studies demonstrate its causation in only around one in 10 individuals with CP. Instead, genetic causes are increasingly demonstrated. We systematically performed clinical phenotyping and genetic investigations in a monocentric CP cohort, aiming to gain insight into the contribution of genetic variants in CP and its different subtypes. MethodsChromosomal microarray and/or trio exome sequencing were systematically performed in 337 individuals with CP between September 2017 and August 2022. Deep phenotyping was performed through clinical multidisciplinary evaluation and review of medical files. ResultsGenetic analyses resulted in an overall diagnostic yield of 38.3% (129 of 337). In cases with one or more comorbidities (intellectual disability, epilepsy, autism spectrum disorder), the yield increased to almost 50%. Functional enrichment analysis showed over-representation of the following pathways: genetic imprinting, DNA modification, liposaccharide metabolic process, neuron projection guidance, and axon development. ConclusionsGenetic analyses in our CP cohort, the largest monocentric study to date, demonstrated a diagnostic yield of 38.3%, highlighting the importance of genetic testing in CP. The diagnosis of a genetic disorder is essential for prognosis and clinical follow-up, as well as for family counseling. Pathway analysis points to dysregulation of general developmental and metabolic processes as well as neuronal development and function. Unraveling the role of these pathways in CP pathogenesis is instrumental for the identification of CP candidate genes as well as potential therapeutic targets.

Disentangling cell-intrinsic and extrinsic factors underlying evolution.

A key goal of developmental biology is to determine the extent to which cells and organs develop autonomously, as opposed to requiring interactions with other cells or environmental factors. Chimeras have played a foundational role in this by enabling qualitative classification of cell-intrinsically vs. extrinsically driven processes. Here, we extend this framework to precisely decompose evolutionary divergence in any quantitative trait into cell-intrinsic, extrinsic, and intrinsic-extrinsic interaction components. Applying this framework to thousands of gene expression levels in reciprocal rat-mouse chimeras, we found that the majority of their divergence is attributable to cell-intrinsic factors, though extrinsic factors also play an integral role. For example, a rat-like extracellular environment extrinsically up-regulates the expression of a key transcriptional regulator of the endoplasmic reticulum (ER) stress response in some but not all cell types, which in turn strongly predicts extrinsic up-regulation of its target genes and of the ER stress response pathway as a whole. This effect is also seen at the protein level, suggesting propagation through multiple regulatory levels. Applying our framework to a cellular trait, neuronal differentiation, revealed a complex interaction of intrinsic and extrinsic factors. Finally, we show that imprinted genes are dramatically mis-expressed in species-mismatched environments, suggesting that mismatch between rapidly evolving intrinsic and extrinsic mechanisms controlling gene imprinting may contribute to barriers to interspecies chimerism. Overall, our conceptual framework opens new avenues to investigate the mechanistic basis of developmental processes and evolutionary divergence across myriad quantitative traits in any multicellular organism.

Loss-of-Imprinting of HM13 Leads to Poor Prognosis in Clear Cell Renal Cell Carcinoma.

Genomic imprinting refers to the epigenetic silencing of one of both alleles in a parent-of-origin-specific manner, particularly in genes regulating growth and development. Impaired genomic imprinting leading to the activation of the silenced allele, also called canonical loss-of-imprinting (LOI), is considered an early factor in oncogenesis. As LOI studies in clear cell renal cell carcinoma (ccRCC) are limited to IGF2, we performed a genome-wide analysis in 128 kidney normal solid tissue and 240 stage 1 ccRCC samples (TCGA RNA-seq data) to screen for canonical LOI in early oncogenesis. In ccRCC, we observed LOI (adj. p = 2.74 × 10-3) of HM13 (Histocompatibility Minor 13), a signal peptide peptidase involved in epitope generation. HM13 LOI samples featured HM13 overexpression, both compared to normal solid tissues (p = 3.00 × 10-7) and non-LOI (p = 1.27 × 10-2) samples. Upon adjustment for age and sex, HM13 expression was significantly associated with poor survival (p = 7.10 × 10-5). Moreover, HM13 overexpression consistently exacerbated with increasing tumor stage (p = 2.90 × 10-8). For IGF2, LOI was observed in normal solid tissues, but the prevalence did not increase in cancer. In conclusion, HM13 LOI is an early event in ccRCC, causing overexpression leading to poor prognosis.

A case of an Angelman-syndrome caused by an intragenic duplication of UBE3A uncovered by adaptive nanopore sequencing

Adaptive nanopore sequencing as a diagnostic method for imprinting disorders and episignature analysis revealed an intragenic duplication of Exon 6 and 7 in UBE3A (NM_000462.5) in a patient with relatively mild Angelman-like syndrome. In an all-in-one nanopore sequencing analysis DNA hypomethylation of the SNURF:TSS-DMR, known contributing deletions on the maternal allele and point mutations in UBE3A could be ruled out as disease drivers. In contrast, breakpoints and orientation of the tandem duplication could clearly be defined. Segregation analysis in the family showed that the duplication derived de novo in the maternal grandfather. Our study shows the benefits of an all-in-one nanopore sequencing approach for the diagnostics of Angelman syndrome and other imprinting disorders.

Genome-Scale Analyses Reveal Roadblocks to Monkey Cloning.

Cloning by somatic cell nuclear transfer (SCNT) remained challenging for Rhesus monkeys, mostly due to its low efficiency and neonatal death. Genome-scale analyses revealed that monkey SCNT embryos displayed widespread DNA methylation and transcriptional alterations, thus including loss of genomic imprinting that correlated with placental dysfunction. The transfer of inner cell masses (ICM) from cloned blastocysts into ICM-depleted fertilized embryos rescued placental insufficiency and gave rise to a cloned Rhesus monkey that reached adulthood without noticeable abnormalities.

Genomic imprinting and random monoallelic expression

In this review we consider mechanisms of monoallelic expression: genomic imprinting, in which gene transcription depends on the parental origin of the allele, and random monoallelic transcription. We summarize evidence for the regulation of activity in imprinted regions. Particular attention is paid to areas that control imprinting. Factors that may influence the variability of the imprintome are considered. The perspectives behind monoallelic expression studies are also discussed.

Exploring the Stability of Genomic Imprinting and X-Chromosome Inactivation in the Aged Brain

Exploring the Stability of Genomic Imprinting and X-Chromosome Inactivation in the Aged Brain

Multilayered epigenetic control of persistent and stage-specific imprinted genes in rice endosperm.

In angiosperms, epigenetic profiles for genomic imprinting are established before fertilization. However, the causal relationships between epigenetic modifications and imprinted expression are not fully understood. In this study, we classified 'persistent' and 'stage-specific' imprinted genes on the basis of time-course transcriptome analysis in rice (Oryza sativa) endosperm and compared them to epigenetic modifications at a single time point. While the levels of epigenetic modifications are relatively low in stage-specific imprinted genes, they are considerably higher in persistent imprinted genes. Overall trends revealed that the maternal alleles of maternally expressed imprinted genes are activated by DNA demethylation, while the maternal alleles of paternally expressed imprinted genes with gene body methylation (gbM) are silenced by DNA demethylation and H3K27me3 deposition, and these regions are associated with an enriched motif related to Tc/Mar-Stowaway. Our findings provide insight into the stability of genomic imprinting and the potential variations associated with endosperm development, different cell types and parental genotypes.

Aberrant DNA methylation associated with the development of metabolic dysfunction-associated fatty liver disease

The literature review deals with DNA methylation, a key epigenetic mechanism that controls the activity of gene transcription, plays a decisive role in the formation of genomic imprinting, gene silencing, X-chromosome inactivation, RNA splicing, DNA repair, cell differentiation and cell reprogramming, and also determines the occurrence and development of liver steatotic lesions and metabolic disorders. Methylation of DNA cytosine dinucleotide (CpG) can be represented in two types: de novo CpG methylation, which is carried out by 5mC DNA writers — DNA-(cytosine-5)-methyltransferase (DNMT) 3a and 3b, and suppor­ting DNA methylation, which is performed by DNMT1 during DNA replication. It has been found that the maintenance DNA methylation allows the preservation of the methylation pattern characteristic of progenitor cells in the cells of the new generation, and the DNA methylation of the gene body is associated with its increased expression. Active demethylation of 5mC is carried out by TET dioxygenases, including three enzymatic representatives: TET1, TET2 and TET3. It has been demonstrated that aberrant methylation of DNA nucleotides is directly related to the activity of lipid synthesis, the degree of oxidative stress, the development of liver steatosis, low-grade inflammation, insulin resistance, and the progression of liver fibrosis. The authors presented in detail the functions and features of DNA methyltransferases, erasers, and readers of 5mC sites; possible violations of the balance of activity of writers and erasers of 5mC DNA; DNA methylation landscape and patterns; clinical significance of DNA methylation signatures in metabolic dysfunction-associated fatty liver disease. Global hypomethylation of genome, at least 55 genes, is observed in patients with metabolic dysfunction-associated fatty liver disease. The authors emphasize that the use of DNA methylation signatures is a promising direction for early diagnosis and prognosis of the course of metabolic dysfunction-associated fatty liver disease, while the study of molecular components of DNA methylation mechanisms involved in the regulation of gene expression, the dependence of their activity on exposure to the exposome will allow to persona­lize and improve recommendations for lifestyle and diet modification in patients with metabolic dysfunction-associated fatty liver disease.

Epigenetic Modifications of Hormonal Signaling Pathways in Plant Drought Response and Tolerance for Sustainable Food Security.

Drought significantly challenges global food security, necessitating a comprehensive understanding of plant molecular responses for effective mitigation strategies. Epigenetic modifications, such as DNA methylation and histone modifications, are key in regulating genes and hormones essential for drought response. While microRNAs (miRNAs) primarily regulate gene expression post-transcriptionally, they can also interact with epigenetic pathways as potential effectors that influence chromatin remodeling. Although the role of miRNAs in epigenetic memory is still being explored, understanding their contribution to drought response requires examining these indirect effects on epigenetic modifications. A key aspect of this exploration is epigenetic memory in drought-adapted plants, offering insights into the transgenerational inheritance of adaptive traits. Understanding the mechanisms that govern the maintenance and erasure of these epigenetic imprints provides nuanced insights into how plants balance stability and flexibility in their epigenomes. A major focus is on the dynamic interaction between hormonal pathways-such as those for abscisic acid (ABA), ethylene, jasmonates, and salicylic acid (SA)-and epigenetic mechanisms. This interplay is crucial for fine-tuning gene expression during drought stress, leading to physiological and morphological adaptations that enhance plant drought resilience. This review also highlights the transformative potential of advanced technologies, such as bisulfite sequencing and CRISPR-Cas9, in providing comprehensive insights into plant responses to water deficit conditions. These technologies pave the way for developing drought-tolerant crops, which is vital for sustainable agriculture.

Ornaments for efficient allele-specific expression estimation with bias correction

Allele-specific expression plays a crucial role in unraveling various biological mechanisms, including genomic imprinting and gene expression controlled by cis-regulatory variants. However, existing methods for quantification from RNA-sequencing (RNA-seq) reads do not adequately and efficiently remove various allele-specific read mapping biases, such as reference bias arising from reads containing the alternative allele that do not map to the reference transcriptome or ambiguous mapping bias caused by reads containing the reference allele that map differently from reads containing the alternative allele. We present Ornaments, a computational tool for rapid and accurate estimation of allele-specific transcript expression at unphased heterozygous loci from RNA-seq reads while correcting for allele-specific read mapping biases. Ornaments removes reference bias by mapping reads to a personalized transcriptome and ambiguous mapping bias by probabilistically assigning reads to multiple transcripts and variant loci they map to. Ornaments is a lightweight extension of kallisto, a popular tool for fast RNA-seq quantification, that improves the efficiency and accuracy of WASP, a popular tool for bias correction in allele-specific read mapping. In experiments with simulated and human lymphoblastoid cell-line RNA-seq reads with the genomes of the 1000 Genomes Project, we demonstrate that Ornaments improves the accuracy of WASP and kallisto, is nearly as efficient as kallisto, and is an order of magnitude faster than WASP per sample, with the additional cost of constructing a personalized index for multiple samples. Additionally, we show that Ornaments finds imprinted transcripts with higher sensitivity than WASP, which detects imprinted signals only at gene level.

Clinical utility of regions of homozygosity (ROH) identified in exome sequencing: when to pursue confirmatory uniparental disomy testing for imprinting disorders?

Regions of homozygosity (ROH) could implicate uniparental disomy (UPD) on specific chromosomes associated with imprinting disorders. Though the algorithms for ROH detection in exome sequencing (ES) have been developed, optimal reporting thresholds and when to pursue confirmatory UPD testing for imprinting disorders remain in ambiguity. This study used a data-driven approach to assess optimal reporting thresholds of ROH in clinical practice. ROH analysis was performed using Automap in a retrospective cohort of 8,219 patients and a prospective cohort of 1,964 patients with ES data. Cases with ROH on imprinting-disorders related chromosomes were selected for additional methylation-specific confirmatory testing. The diagnostic yield, the ROH pattern of eventually diagnosed cases and optimal thresholds for confirmatory testing were analyzed. In the retrospective analysis, 15 true UPD cases of imprinting disorders were confirmed among 51 suspected cases by ROH detection. Pattern of ROH differed between confirmed UPD and non-UPD cases. Maximized yield and minimized false discovery rate of confirmatory UPD testing was achieved at the thresholds of >20 Mb or >25 % chromosomal coverage for interstitial ROH, and >5 Mb for terminal ROH. Current recommendation by ACMG was nearly optimal, though refined thresholds as proposed in this study could reduce the workload by 31 % without losing any true UPD diagnosis. Our refined thresholds remained optimal after independent evaluation in a prospective cohort. ROH identified in ES could implicate the presence of clinically relevant UPD. This study recommended size and coverage thresholds for confirmatory UPD testing after ROH detection in ES, contributing to the development of evidence-based reporting guidelines.

Gestational Trophoblastic Disease: Complete versus Partial Hydatidiform Moles.

Hydatidiform moles, including both complete and partial moles, constitute a subset of gestational trophoblastic diseases characterized by abnormal fertilization resulting in villous hydrops and trophoblastic hyperplasia with or without embryonic development. This involves chromosomal abnormalities, where one or two sperms fertilize an empty oocyte (complete hydatidiform mole (CHM); mostly 46,XX) or two sperms fertilize one oocyte (partial hydatidiform mole (PHM); mostly 69,XXY). Notably, recurrent occurrences are associated with abnormal genomic imprinting of maternal effect genes such as NLRP7 (chromosome 19q13.4) and KHDC3L (chromosome 6q1). Ongoing efforts to enhance identification methods have led to the identification of growth-specific markers, including p57 (cyclin-dependent kinase inhibitor 1C; CDKN1C), which shows intact nuclear expression in the villous cytotrophoblast and villous stromal cells in PHMs and loss of expression in CHMs. Treatment of hydatidiform moles includes dilation and curettage for uterine evacuation of the molar pregnancy followed by surveillance of human chorionic gonadotropin (HCG) levels to confirm disease resolution and rule out the development of any gestational trophoblastic neoplasia. In this review, we provide a synopsis of the existing literature on hydatidiform moles, their diagnosis, histopathologic features, and management.

Automatized detection of uniparental disomies in a large cohort

Uniparental disomy (UPD) is the inheritance of both homologues of a chromosome from only one parent. The detection of UPDs in sequencing data is not well established and a common gap in genetic diagnostics. We applied our in-house UPD detection pipeline to evaluate a cohort of 9212 samples, including multigene panels as well as exome sequencing data in a single, duo or trio constellation. We used the results to inform the design of our publicly available web app altAFplotter. UPDs categorized as heterodisomy, whole chromosome or segmental isodisomy were identified and validated with microsatellites, multiplex ligation-dependent probe amplification as well as Sanger sequencing. We detected 14 previously undiagnosed UPDs including nine isodisomies, four segmental isodisomies as well as one heterodisomy on chromosome 22. We characterized eight findings as potentially causative through homozygous pathogenic variants or imprinting disorders. Overall, our study demonstrates the utility of our UPD detection pipeline with our web app, altAFplotter, to reliably identify UPDs. This not only increases the diagnostic yield of cases with growth and metabolic disturbances, as well as developmental delay, but also enhances the understanding of UPDs that may be relevant for recurrence risks and genetic counseling.

Aspects of genetic imprinting: implications, syndromes and therapies

Aspects of genetic imprinting: implications, syndromes and therapies

Hypothetical involvement of stress hormones-induced reprograming of adult stem/progenitor cells in tumorigenesis

Stress is a state of threatened or perceived as threatened homeostasis that can be induced by various external and internal stimuli such as psychosocial factors, inflammatory or injurious conditions, and infections. In order to restore body homeostasis, adrenal glands produce and secrete glucocorticoids (GCs) and catecholamines (CAs), which are the main stress hormones that support the survival and adaptation of the organisms to the new environment. In contrast to the rather beneficial impact of acute and short-lasting stress, chronic stress and related dysregulation of the stress system is implicated in the development of many non-communicable diseases, including cancer. Particularly, ever-increasing experimental and clinical evidence implicates the involvement of CAs and GCs as well as the overexpression of their receptors in the activation of the major pathways involved in tumour development, metastasis, and resistance to various therapies. More importantly, results of experimental and epidemiological studies revealed that overexposure to stress hormones during pre- and early postnatal life might induce life-long or even transgenerational dysregulation of the stress system and predispose it to the development of various tumours. Although the exact mechanisms involved in the latter process are not yet fully known, it has been demonstrated that GC-induced epigenetic modifications can change the expression of several key genes involved in the regulation of the stress system, tumour initiation, and epigenetic imprinting. When such alterations occur in stem/progenitor cells (SPCs), this might not only lead to long-term dysfunction of the stress system but might promote the generation of cancer stem cells (CSCs). This review article discusses a hypothesis that stress hormones-mediated epigenetic reprograming of various SPCs during sensitive developmental periods, might contribute to their dysfunction and increased sensitivity to malignant transformation, thereby promoting tumorigenesis.

Creation and validation of the first infinium DNA methylation array for the human imprintome

BackgroundDifferentially methylated imprint control regions (ICRs) regulate the monoallelic expression of imprinted genes. Their epigenetic dysregulation by environmental exposures throughout life results in the formation of common chronic diseases. Unfortunately, existing Infinium methylation arrays lack the ability to profile these regions adequately. Whole genome bisulfite sequencing (WGBS) is the unique method able to profile the ICRs. However, it is very expensive and it requires not only a high coverage, but it is also computationally intensive to assess these regions.FindingsTo address this deficiency, we developed a custom methylation array containing 22,819 probes. Among them, 10,438 are CG probes targeting unique CpG sites, with 9,757 probes successfully mapping to 1,088 out of the 1,488 candidate ICRs recently described. To assess the performance of the array, we created matched samples processed with the Human Imprintome array and WGBS, which is the current standard method for assessing the methylation of the Human Imprintome. We compared the methylation levels from the shared CpG sites, and obtained a mean R2 = 0.569. We also created matched samples processed with the Human Imprintome array and the Infinium Methylation EPIC v2 array, and obtained a mean R2 = 0.796. Furthermore, replication experiments demonstrated high reliability (ICC: 0.799–0.945).ConclusionsOur custom array will be useful for replicable and accurate assessment, mechanistic insight, and targeted investigation of ICRs. This tool should accelerate the discovery of ICRs associated with a wide range of diseases and exposures, and advance our understanding of genomic imprinting and its relevance in development and disease formation throughout the life course.

Epigenetic and transcriptional consequences in the endosperm of chemically induced transposon mobilization in Arabidopsis.

Genomic imprinting, an epigenetic phenomenon leading to parent-of-origin-specific gene expression, has independently evolved in the endosperm of flowering plants and the placenta of mammals-tissues crucial for nurturing embryos. While transposable elements (TEs) frequently colocalize with imprinted genes and are implicated in imprinting establishment, direct investigations of the impact of de novo TE transposition on genomic imprinting remain scarce. In this study, we explored the effects of chemically induced transposition of the Copia element ONSEN on genomic imprinting in Arabidopsis thaliana. Through the combination of chemical TE mobilization and doubled haploid induction, we generated a line with 40 new ONSEN copies. Our findings reveal a preferential targeting of maternally expressed genes (MEGs) for transposition, aligning with the colocalization of H2A.Z and H3K27me3 in MEGs-both previously identified as promoters of ONSEN insertions. Additionally, we demonstrate that chemically-induced DNA hypomethylation induces global transcriptional deregulation in the endosperm, leading to the breakdown of MEG imprinting. This study provides insights into the consequences of chemically induced TE remobilization in the endosperm, revealing that chemically-induced epigenome changes can have long-term consequences on imprinted gene expression.