Project description:We designed and introduced a new methylation array concentrating on human trait screening and discovery. The new MSA (Methylation Screening Array) leveraged the massive Infinium platform-based data from epigenome-wide association studies, combined with updated knowledge from the latest single cell and cell type-resolution whole genome methylome profiles, to achieve scalable screening of epigenetics-trait association in an ultra-high sample-throughput. Our design encompassed diverse human trait associations, including those with genetic, biological, and demographical variables, environmental exposures, and common human diseases such as neurodegenerative, genetic, cardiovascular, infectious, and immune diseases. We comprehensively evaluated this array's reproducibility, accuracy, and capacity in supporting 5-hydroxymethylation profiling and comprehensive cell-type deconvolution in diverse human tissues. Our first data using this platform uncovered dynamic chromatin and tissue contexts of DNA modification variations and genetic variants with human trait associations.
Project description:Primary objectives: Avaliar a eficácia da abordagem profilática com minociclina no desenvolvimento de toxicidade dermatológica secundária ao cetuximab
Primary endpoints: Percentagem de doentes que desenvolvem rash associado ao tratamento com cetuximab para o cancro colo-rectal, no grupo experimental comparativamente com o grupo controlo
Project description:DNA methylation serves a stable gene regulatory function in mature somatic cells. In the germ line and during early embryogenesis, however, DNA methylation undergoes global erasure and re-establishment to support germ cell and embryonic development. While de novo DNA methylation during male germ cell development is essential for setting genomic imprints, possible other intergenerational roles for paternal DNA methylation following fertilization are unknown. To address this question, we reduced the level of DNA methylation in developing male germ cells through conditional gene deletion of the de novo DNA methyltransferases DNMT3A and DNMT3B in undifferentiated spermatogonia. Mutant male germ cells nevertheless completed their differentiation to sperm. We observed that DNMT3A serves a largely maintenance-like methylation function at many intragenic sites in undifferentiated spermatogonia while DNMT3B catalyzes de novo methylation during spermatogonial differentiation. In spermatogonia, the acquisition of DNA methylation and deposition of H3K4me3 occur mutually exclusive. Failing de novo DNA methylation in spermatogonia leads to increased nucleosome occupancy in mature sperm at sites with high CpG content, reinforcing the model that DNA methylation constrains nucleosome retention in sperm. To assess the impact of altered sperm chromatin in the formation of embryonic chromatin, we measured H3K4me3 occupancy at paternal and maternal alleles in 2-cell embryos using a highly sensitive transposon-based tagging assay for modified chromatin. Our data show that reduced DNA methylation in sperm renders paternal alleles permissive for H3K4me3 establishment in early embryos, independently from paternal inheritance of sperm born H3K4me3. Together, this study provides first evidence that paternally inherited DNA methylation directs chromatin formation during early embryonic development.
Project description:The link between DNA methylation and neurodevelopmental disorders is well established. However, how DNA methylation is fine-tuned – ensuring precise gene expression and developmental fidelity – remains poorly understood. PROSER1, a known TET2 interactor, was recently linked to a severe neurodevelopmental disorder. Here, we demonstrate that PROSER1 interacts with all TET enzymes and stabilizes chromatin-bound TET-OGT-PROSER1-DBHS (TOPD) complexes, which regulate DNA demethylation and developmental gene expression. Surprisingly, we find that PROSER1 also sequesters TET enzymes, preventing widespread demethylation and transposable element de-repression. Our findings identify PROSER1 as a key factor which both positively and negatively regulates DNA demethylation essential for mammalian neurodevelopment.
Project description:ObjectivesThis study aims to investigate the association of the tumor necrosis factor-alpha inducible protein 3 (TNFAIP3) (rs5029939) gene single nucleotide polymorphism (SNP) with the risk of systemic lupus erythematosus (SLE) and its clinical manifestations in a cohort of SLE patients.Patients and methodsThis study included a total of 180 participants (18 males, 72 females; mean age: 30.9±10.1 years; range 17 to 59 years) including 90 SLE patients and 90 healthy controls between March 2017 and February 2020. The TNFAIP3 rs5029939 gene polymorphism was identified by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) in all participants.ResultsThere was a significant difference in genotype distribution of the TNFAIP3 rs5029939 SNP between SLE patients and healthy controls, where CG genotype was more common in SLE patients (53.3%) than controls (11.1%) (p=0.001). We found a significant difference in G allele frequency of TNFAIP3 (rs5029939) (37.8% with SLE vs. 5.6% with controls, p=0.001). Genotype CG was significantly associated with lupus nephritis and neuropsychiatric manifestations (p<0.05). Although the response to treatment was numerically higher with the genotype CC, it did not reach statistical significance (p=0.4).ConclusionOur study suggests that TNFAIP3 rs5029939 gene polymorphism is associated with SLE susceptibility and may have an impact on its clinical phenotype. As such association differs among populations of diverse ethnic backgrounds, larger genome-wide association studies are warranted to further elucidate genetic associations.
Project description:Light triggers chloroplast differentiation whereby the etioplast transforms into a photosynthesizing chloroplast and the thylakoid rapidly emerges. However, the sequence of events during chloroplast differentiation remains poorly understood. Here we used whole-seedling proteome data to quantify changes in protein abundances during the course of de-etiolation within the first four days of light exposure. This data complements quantitative lipid and (ultra)structural data described in Pipitone et al. (doi: https://doi.org/10.1101/2020.08.30.274043).
Project description:Long known as the site of ribosome biogenesis, the nucleolus is increasingly recognized for its role in shaping 3D genome organization. Still, the mechanisms governing the targeting of selected regions of the genome to nucleolus-associated domains (NADs) remain enigmatic. Here we reveal the essential role of ZNF274, a SCAN-bearing member of the Krüppel-associated box (KRAB)-containing zinc finger proteins (KZFP) family, in sequestering lineage-specific gene clusters within NADs. Ablation of ZNF274 triggers transcriptional activation across entire genomic neighborhoods – encompassing, among others, protocadherin and KZFP-encoding genes – with loss of repressive chromatin marks, altered 3D genome architecture and de novo CTCF binding. Mechanistically, ZNF274 anchors target DNA sequences at the nucleolus and facilitates their compartmentalization via a previously uncharted function of the SCAN domain. Our findings illuminate the mechanisms underlying NADs organization and suggest that perinucleolar entrapment into repressive hubs constrains the activation of tandemly arrayed genes to enable selective expression and modulate cell differentiation programs during development.
Project description:Epigenetic reprogramming resets parental epigenetic memories and differentiates primordial germ cells (PGCs) into mitotic pro-spermatogonia or oogonia, ensuring sexually dimorphic germ-cell development for totipotency. However, the mechanism of epigenetic reprogramming in humans remains unknown. Here, we establish a robust strategy for inducing epigenetic reprogramming and differentiation of pluripotent stem cell (PSC)-derived human PGC-like cells (hPGCLCs) into mitotic pro-spermatogonia or oogonia, coupled with their extensive amplification (~>10(10)-fold). Strikingly, bone morphogenetic protein (BMP) signaling is the key driver of these processes. Mechanistically, BMP signaling attenuates the mitogen-activated protein kinase/extracellular-regulated kinase (MAPK/ERK) pathway and both de novo and maintenance DNA methyltransferase (DNMT) activities, promoting replication-coupled, passive DNA demethylation. On the other hand, tens-eleven translocation (TET) 1, an active DNA demethylase abundant in human germ cells, plays a dual role in hPGCLC differentiation: safeguarding hPGCLCs against differentiation into amnion-like cells by repressing key genes with bivalent promoters, and facilitating coordinated activation of genes vital for spermatogenesis and oogenesis by demethylating their promoters. Our study uncovers the principle of epigenetic reprogramming in humans, making a fundamental advance in human biology, and through the generation of abundant mitotic pro-spermatogonia and oogonia-like cells, represents a milestone for human in vitro gametogenesis (IVG) research and its potential translation into reproductive medicine.
Project description:Accurate HPV genotyping is crucial in facilitating epidemiology studies, vaccine trials, and HPV-related cancer research. Contemporary HPV genotyping assays only detect < 25% of all known HPV genotypes and are not accurate for low-risk or mixed HPV genotypes. Current genomic HPV genotyping algorithms use a simple read-alignment and filtering strategy that has difficulty handling repeats and homology sequences. Therefore, we have developed an optimized expectation-maximization algorithm, designated HPV-EM, to address the ambiguities caused by repetitive sequencing reads. HPV-EM achieved 97-100% accuracy when benchmarked using cell line data and TCGA cervical cancer data. We also validated HPV-EM using DNA tiling data on an institutional cervical cancer cohort (96.5% accuracy). Using HPV-EM, we demonstrated HPV genotypic differences in recurrence and patient outcomes in cervical and head and neck cancers.
Project description:Interventions: ultra low dose CT colonography
Primary outcome(s): Per-patient performance of CT ultra low dose colonography in the detection of neoplasms, with colonoscopy results as the reference standard.
Study Design: Single arm Non-randomized