Biology subjects

Castillo-Fernandez, J.

Publications and source records attributed to Castillo-Fernandez, J..

6 recordsLinked to original sources

bioRxiv2025

Genetic impacts on within-pair DNA methylation variance in monozygotic twins capture gene-environment interactions and cell-type effects

BackgroundGenetic variants that are associated with phenotypic variability, or variance quantitative trait loci (vQTLs), have been detected for multiple human traits. Gene-environment interactions can lead to differential phenotypic variability across genotype groups, therefore, genetic variants that interact with environmental exposures can manifest as vQTLs. Although changes in DNA methylation variability have been observed in several diseases, vQTLs for methylation levels (vmeQTL) have not yet been explored in depth. ResultsWe optimize the value of monozygotic (MZ) twin studies to identify and replicate vmeQTLs for blood DNA methylation variance at 358 CpGs in 988 adult MZ twin pairs from two European twin registries. Over a third of vmeQTLs captured identical vmeQTL-environmental factor interactions in both datasets, and the majority of interactions were observed with blood cell counts. Correspondingly, over 60% of CpGs affected by genotype-monocyte and genotype-T cell interactions replicated as CpGs affected by genetic effects in the relevant cell type in an independent dataset. Most vmeQTLs also replicated in 1,348 UK non-twin adults and showed longitudinal stability in a sample subset. Integrating gene expression and phenotype association results identified multiple vmeQTLs that capture GxE effects relevant to human health. Examples include vmeQTLs interacting with blood cell type to influence DNA methylation in FAM65A, NAPRT, and CSGALNACT1 underlying immune disease susceptibility and progression. ConclusionOur findings identify novel genetic effects on human DNA methylation variability within a unique MZ twin study design. The results show the potential of vmeQTLs to identify gene-environment interactions and provide novel insights into complex traits.

genetics↗

bioRxiv2023

Genetic impacts on DNA methylation help elucidate regulatory genomic processes

Pinpointing genetic impacts on DNA methylation can improve our understanding of pathways that underlie gene regulation and disease risk. We report heritability and methylation quantitative trait locus (meQTL) analysis at 724,499 CpGs profiled with the Illumina Infinium MethylationEPIC array in 2,358 blood samples from three UK cohorts, with replication. Methylation levels at 34.2% of CpGs were affected by SNPs, and 98% of effects were cis-acting or within 1 Mbp of the tested CpG. Our results are consistent with meQTL analyses based on the former Illumina Infinium HumanMethylation450 array. Both meQTL SNPs and CpGs with meQTLs were overrepresented in enhancers, which have improved coverage on this platform compared to previous approaches. Co-localisation analyses across genetic effects on DNA methylation and 56 human traits identified 1,520 co-localisations across 1,325 unique CpGs and 34 phenotypes, including in disease-relevant genes, such ICOSLG (inflammatory bowel disease), and USP1 and DOCK7 (total cholesterol levels). Enrichment analysis of meQTLs and integration with expression QTLs gave insights into mechanisms underlying cis-meQTLs, for example through disruption of transcription factor binding sites for CTCF and SMC3, and trans-meQTLs, for example through regulating the expression of ACD and SENP7 which can modulate DNA methylation at distal sites. Our findings improve the characterisation of the mechanisms underlying DNA methylation variability and are informative for prioritisation of GWAS variants for functional follow-ups. A results database and viewer are available online.

genomics↗

bioRxiv2021

GLP is critical for oogenesis exhibiting a G9A-independent role in transcriptional repression

GLP (EHMT1) is a multifunctional protein, best known for its role as an H3K9me1 and H3K9me2 methyltransferase through its reportedly obligatory dimerization with G9A (EHMT2). Here, we investigate the role of GLP in the oocyte in comparison to G9A using oocyte-specific conditional knockout mouse models (G9a cKO, Glp cKO, G9a-Glp cDKO). Loss of GLP in Glp cKO and G9a-Glp cDKO oocytes re-capitulated meiotic defects observed in the G9a cKO; however, there was a significant impairment in oocyte maturation and developmental competence in Glp cKO and G9a-Glp cDKO oocytes beyond that observed in the G9a cKO. Consequently, loss of GLP in oogenesis results upon fertilisation in mid-gestation embryonic lethality. To assess the molecular functions of GLP and G9A, we applied a multi-omics approach, supported by immunofluorescence, to identify changes in epigenomic, transcriptomic and proteomic signatures in cKO oocytes. H3K9me2 was equally depleted in all cKO oocytes, whereas H3K9me1 was decreased only upon loss of GLP. The transcriptome, DNA methylome and proteome were markedly more affected in G9a-Glp cDKO than G9a cKO oocytes, with transcriptional de-repression associated with increased protein abundance and gains in genic DNA methylation in G9a-Glp cDKO oocytes. Together, our findings suggest that GLP contributes to transcriptional repression in the oocyte, independent of G9A, and is critical for oogenesis and oocyte developmental competence.

developmental biology↗

bioRxiv2021

Leptin Signalling in the Ovary of Diet-Induced Obese Mice Regulates Activation of Nod-Like Receptor Protein 3 Inflammasome

1Obesity leads to ovarian dysfunction and the establishment of local leptin resistance. The aim of our study was to characterise levels of Nod-Like Receptor Protein 3 (NLRP3) inflammasome activation during obesity progression in the mouse ovaries and liver and test the putative role of leptin on its regulation. C57BL/6J mice were treated with equine chorionic gonadotropin (eCG) or human chorionic gonadotropin (hCG) for oestrous cycle synchronisation and ovaries collection. In diet-induced obesity (DIO) model, mice were fed chow diet (CD) or high fat diet (HFD) for 4 or 16 weeks (wk), whereas in hyperleptinemic model (LEPT), mice were injected with leptin for 16 days (16L) or saline (16C) and in the genetic obese leptin-deficient ob/ob (+/? and -/-) animals were fed CD for 4wk. Either ovaries and liver were collected, as well as cumulus cells (CCs) after superovulation from DIO and LEPT. In DIO protocol, protein expression of NLRP3 inflammasome components was increased in 4wk HFD, but decreased in 16wk HFD. Moreover LEPT and ob/ob models revealed NLRP3 and IL-1{beta} upregulation in 16L and downregulation in ob/ob. Transcriptome analysis of CC showed common genes between LEPT and 4wk HFD modulating NLRP3 inflammasome. Moreover analysis in the liver showed upregulation of NLRP3 protein only after 16wk HFD, but also the downregulation of NLRP3 protein in ob/ob-/-. We showed the link between leptin signalling and NLRP3 inflammasome activation in the ovary throughout obesity progression in mice, elucidating the molecular mechanisms underpinning ovarian failure in maternal obesity.

physiology↗

bioRxiv2020

Epigenetic change induced by in utero dietary challenge provokes phenotypic variability across multiple generations of mice

Transmission of epigenetic information between generations occurs in nematodes, flies and plants, mediated by specialised small RNA pathways, histone H3K9me3, H3K27me3, H4K16ac and DNA methylation1-3. In higher vertebrates, epidemiological and experimental evidence supports similar trans-generational effects4,5 although the mechanisms that underpin these are incompletely understood6-9. We generated a luciferase reporter knock-in mouse for the imprinted Dlk1 locus, to visualise and track epigenetic fidelity across generations. We showed that exposure to high-fat diet (HFD) in pregnancy provokes sustained re-expression of the normally silent maternal Dlk1 allele in offspring, coincident with increased DNA methylation at the Dlk1 sDMR. Interestingly, maternal Dlk1 mis-expression was also evident in the next generation (F2), exclusively in animals derived from F1-exposed females. Oocytes from these females showed altered microRNA and gene expression, without any major changes in underlying DNA methylation, and correctly imprinted Dlk1 expression resumed in subsequent generations (F3 onwards). Our results reveal how canonical and non-canonical imprinting mechanisms enable the foetal epigenome to adapt to in utero challenge to modulate the properties of two successive generations of offspring.

developmental biology↗

bioRxiv2019

Leptin resistance establishment in the ovary of diet-induced obese mice and its relationship with cumulus cells transcriptome

Background/AimsObesity is associated with infertility, decreased ovarian performance and lipotoxicity. However, little is known about the aetiology of these reproductive impairments. Here, we hypothesise that the majority of changes in ovarian physiology in diet-induced obesity (DIO) are a consequence of transcriptional changes downstream of altered leptin signalling. Therefore, we investigated the extent to which leptin signalling is altered in the ovary upon obesity with particular emphasis on effects on cumulus cells (CCs), the intimate functional companions of the oocyte. Furthermore, we used the pharmacological hyperleptinemic (LEPT) mouse model to compare transcriptional profiles to DIO. MethodsMice were subjected to DIO for 4 and 16 weeks (wk) and leptin treatment for 16 days, to study effects in the ovary in components of leptin signalling at the transcript and protein levels, using Western blot, Real-time PCR and immunostaining. Furthermore, we used low-cell RNA sequencing to characterise changes in the transcriptome of CCs in these models. ResultsIn the DIO model, obesity led to establishment of ovarian leptin resistance after 16 wk high fat diet (HFD), as evidenced by increases in the feedback regulator suppressor of cytokine signalling 3 (SOCS3) and decreases in the positive effectors phosphorylation of tyrosine 985 of leptin receptor (ObRb-pTyr985) and Janus kinase 2 (pJAK2). Transcriptome analysis of the CCs revealed a complex response to DIO, with large numbers and distinct sets of genes deregulated at early and late stages of obesity; in addition, there was a striking correlation between body weight and global transcriptome profile of CCs. Further analysis indicated that the transcriptome profile in 4 wk HFD CCs resembled that of LEPT CCs, in the upregulation of cellular trafficking and impairment in cytoskeleton organisation. Conversely, after 16 wk HFD CCs showed expression changes indicative of augmented inflammatory responses, cell morphogenesis, and decreased metabolism and transport, mainly as a consequence of the physiological changes of obesity. ConclusionObesity leads to ovarian leptin resistance and major time-dependent changes in gene expression in CCs, which in early obesity may be caused by increased leptin signalling in the ovary, whereas in late obesity are likely to be a consequence of metabolic changes taking place in the obese mother.

physiology↗