Biology subjects

Naval-Sanchez, M.

Publications and source records attributed to Naval-Sanchez, M..

8 recordsLinked to original sources

bioRxiv2026

A scalable MNase-seq framework for reproducible nucleosome profiling across pluripotent stem cell and cardiomyocyte models

Micrococcal nuclease (MNase) digestion is widely used to profile chromatin accessibility and nucleosome footprinting. However, its application is often limited by sensitivity to reaction conditions, high cell input requirements, and the lack of standardized protocols across cell types. Here we developed a robust MNase workflow encompassing buffer composition, DNA purification chemistry, fixation and decrosslinking parameters, cell input scalability, and an in-house yeast spike-in for quantitative normalization. We validated this unified framework across human induced pluripotent stem cells (hiPSCs), hiPSC-derived cardiomyocytes at multiple differentiation stages, primary murine embryonic cardiac cells, and adult mouse cardiomyocytes, and demonstrated comparable digestion efficiencies and kinetics despite marked differences in cellular architecture and chromatin organization. Genome-wide MNase-seq in hiPSCs, combined with the nucMACC bioinformatic pipeline, resolved concentration-dependent nucleosomal occupancy and precise nucleosome positioning at pluripotency-related regulatory elements. This modular, end-to-end, and scalable workflow provides a standardized platform for reproducible MNase-based chromatin profiling across diverse in vitro and in vivo models. TEASERA unified, rapid, and scalable MNase-seq workflow for reproducible mononucleosomal and subnucleosomal profiling from stem cells to adult cardiomyocytes. HIGHLIGHTSO_LISystematic MNase optimization across buffer, DNA purification, and cell input variables C_LIO_LIUnified workflow validated in hiPSCs, hiPSC-CMs, embryonic, and adult cardiomyocytes C_LIO_LIFixed-cell protocol enables weeks of storage without loss of DNA quality C_LIO_LICost-effective yeast spike-in ensures quantitative normalization for MNase-seq C_LIO_LIGenome-wide analyses confirm robust and precise nucleosome positioning at regulatory elements C_LI

genomics↗

bioRxiv2021

Micrococcal nuclease sequencing of porcine sperm suggests a nucleosomal involvement on semen quality and early embryo development

BackgroundThe mammalian mature spermatozoon has a unique chromatin structure in which the vast majority of histones are replaced by protamines during spermatogenesis and a small fraction of nucleosomes are retained at specific locations of the genome. The chromatin structure of sperm remains unresolved in most livestock species, including the pig. However, its resolution could provide further light into the identification of the genomic regions related to sperm biology and embryo development and it could also help identifying molecular markers for sperm quality and fertility traits. Here, for the first time in swine, we performed Micrococcal Nuclease coupled with high throughput sequencing on pig sperm and characterized the mono-nucleosomal (MN) and sub-nucleosomal (SN) chromatin fractions. ResultsWe identified 25,293 and 4,239 peaks in the mono-nucleosomal and sub-nucleosomal fractions, covering 0.3% and 0.02% of the porcine genome, respectively. A cross-species comparison of nucleosome-associated DNAs in sperm revealed positional conservation of the nucleosome retention between human and pig. Gene ontology analysis of the genes mapping nearby the mono-nucleosomal peaks and identification of putative transcription factor binding motifs within the mono-nucleosomal peaks showed enrichment for sperm function and embryo development related processes. We found motif enrichment for the transcription factor Znf263, which in humans was suggested to be a key regulator of the genes with paternal preferential expression during early embryo development. Moreover, we found enriched co-occupancy between the RNAs present in pig sperm and the RNA related to sperm quality, and the mono-nucleosomal peaks. We also found preferential co-location between GWAS hits for semen quality in swine and the mono-nucleosomal sites identified in this study. ConclusionsThese results suggest a clear relationship between nucleosome positioning in sperm and sperm and embryo development.

genomics↗

bioRxiv2020

Analysis Of Polycerate Mutants Reveals The Evolutionary Co-Option Of HOXD1 To Determine The Number And Topology Of Horns In Bovidae

In the course of evolution, pecorans (i.e. higher ruminants) developed a remarkable diversity of osseous cranial appendages, collectively referred to as headgear, which likely share the same origin and genetic basis. However, the nature and function of the genetic determinants underlying their number and position remain elusive. Jacob and other rare populations of sheep and goats, are characterized by polyceraty, the presence of more than two horns. Here, we characterize distinct POLYCERATE alleles in each species, both associated with defective HOXD1 function. We show that haploinsufficiency at this locus results in the splitting of horn bud primordia, likely following the abnormal extension of an initial morphogenetic field. These results highlight the key role played by this gene in headgear patterning and illustrate the evolutionary co-option of a gene involved in the early development of bilateria to properly fix the position and number of these distinctive organs of Bovidae.

genomics↗

bioRxiv2020

Integrated transcriptome, DNA methylome and chromatin state accessibility landscapes reveal regulators of Atlantic salmon maturation

Despite sexual development being ubiquitous to vertebrates, the epigenetic mechanisms controlling this fundamental transition remain largely undocumented in many organisms. Through whole-methylome, whole-transcriptome and chromatin landscape sequencing, we discovered global control mechanisms as well as specific regulators of sexual maturation in Atlantic salmon. This large integrated study was based on an experimental time course that successfully sampled the period when Atlantic salmon commence their trajectory towards sexual maturation. Co-analysis of DNA methylome and gene expression changes revealed chromatin remodelling genes arid1b and smarca2 were both significantly hypermethylated and upregulated in the ovary during the onset of maturation. We also observed changes in chromatin state landscape occurred early in the transition and were strongly correlated with fundamental remodelling of gene expression. Finally, we integrated our multiomics datasets to identify trim25 and znf423 as key regulators in the pituitary that underwent 60 fold change in connectivity during the transition to sexual maturation. The study provides a comprehensive view of the spatiotemporal changes involved in a complex trait and opens the door to future efforts aiming to manipulate puberty in an economically important aquaculture species.

genomics↗

bioRxiv2020

Genome-wide co-expression distributions as a metric to prioritize genes of functional importance

Genome-wide gene expression is routinely used as a tool to gain a systems-level understanding of complex, biological processes. Numerical approaches that have been used to highlight influential genes include abundance, differential expression, differential variation, network connectivity and differential connectivity. Network connectivity tends to be built on a small subset of extremely high co-expression signals that are deemed significant, but this overlooks the vast majority of pairwise signals. Here, we aimed to assess a complementary strategy, namely whether the entire shape of the distribution of genome-wide co-expression values contains a meaningful biological signal that has hitherto remained hidden from view. We have developed a computational pipeline to assign one of 8 distributions (including normal, skewed, bimodal, kurtotic, inverted) to every gene. We then used a hypergeometric enrichment process to determine if particular genes (regulators versus non-regulators) and properties (differentially expressed or not) tend to be associated with particular distributions greater than would be expected by chance. Examination of several distinct data sets spanning 4 species indicates that there is indeed an additional biological signal present in the genome-wide distribution of co-expression values which would be overlooked by currently adopted approaches. Author summaryHigh-throughput technologies, such as RNA-Seq, enables access to a vast amount of data. Here, we describe a new approach to interrogate these data and extract further information to help researchers to understand complex phenotypes. Our method is based on gene-level co-expression distributions which were compared to eight possible template shapes to group genes with similar behaviours. The method was tested using five different datasets and the consistency of the results indicate it can be used as a complementary strategy to analyse transcriptomic data.

bioinformatics↗

bioRxiv2019

Association analysis of loci implied in “buffering” epistasis

The existence of buffering mechanisms is an emerging property of biological networks, and this results in the possible existence of \"buffering\" loci, that would allow buildup of robustness through evolution. So far, there are no explicit methods to find loci implied in buffering mechanisms. However, buffering can be seen as interaction with genetic background. Here we develop this idea into a tractable model for quantitative genetics, in which the buffering effect of one locus with many other loci is condensed into a single (statistical) effect, multiplicative on the total (statistical) additive genetic effect. This allows easier interpretation of the results, and it also simplifies the problem of detecting epistasis from quadratic to linear in the number of loci. Armed with this formulation, we construct a linear model for genome-wide association studies that estimates, and declares significance, of multiplicative epistatic effects at single loci. The model has the form of a variance components, norm reaction model and likelihood ratio tests are used for significance. This model is a generalization and explanation of previous ones. We then test our model using bovine data: Brahman and Tropical Composite animals, phenotyped for body weight at yearling and genotyped up to [~]770,000 Single Nucleotide Polymorphisms (SNP). After association analysis and based on False Discovery Rate rules, we find a number of loci with buffering action in one, the other, or both breeds; these loci do not have significant statistical additive effect. Most of these loci have been reported in previous studies, either with an additive effect, or as footprints of selection. We identify epistatic SNPs present in or near genes encoding for proteins that are functionally enriched for peptide activity and transcription factors reported in the context of signatures of selection in multi-breed cattle population studies. These include loci known to be associated with coat color, fertility and adaptation to tropical environments. In these populations we found loci that have a non-significant statistical additive effect but a significant epistatic effect. We argue that the discovery and study of loci associated with buffering effects allows attacking the difficult problems, among others, of release of maintenance variance in artificial and natural selection, of quick adaptation to the environment, and of opposite signs of marker effects in different backgrounds. We conclude that our method and our results generate promising new perspectives for research in evolutionary and quantitative genetics based on the study of loci that buffer effect of other loci.

genetics↗

bioRxiv2018

Systems biology reveals NR2F6 and TGFB1 as key regulators of feed efficiency in beef cattle

Systems biology approaches are used as strategy to uncover tissue-specific perturbations and regulatory genes related to complex phenotypes. We applied this approach to study feed efficiency (FE) in beef cattle, an important trait both economically and environmentally. Poly-A selected RNA of five tissues (adrenal gland, hypothalamus, liver, skeletal muscle and pituitary) of eighteen young bulls, selected for high and low FE, were sequenced (100bp, pared-end). From the 17,354 expressed genes, 1,317 were prioritized by five selection categories (differentially expressed, harbouring SNPs associated with FE, tissue-specific, secreted in plasma and key regulators) and used for network construction. NR2F6 and TGFB were identified and validated by motif discovery as key regulators of hepatic inflammatory response and muscle tissue development, respectively, two biological processes demonstrated to be associated to FE. Moreover, we indicated potential biomarkers of FE which are related to hormonal control of metabolism and sexual maturity. By using robust methodologies and validation strategies, we confirmed main biological processes related to FE in Bos indicus and indicated candidate genes as regulators or biomarkers of superior animals.

systems biology↗

bioRxiv2017

Mammalian genomic regulatory regions predicted by utilizing human genomics, transcriptomics and epigenetics data

Genome sequences for hundreds of mammalian species are available, but an understanding of their genomic regulatory regions, which control gene expression, is only beginning. A comprehensive prediction of potential active regulatory regions is necessary to functionally study the roles of the majority of genomic variants in evolution, domestication, and animal production. We developed a computational method to predict regulatory DNA sequences (promoters, enhancers and transcription factor binding sites) in production animals (cows and pigs) and extended its broad applicability to other mammals. The method utilizes human regulatory features identified from thousands of tissues, cell lines, and experimental assays to find homologous regions that are conserved in sequences and genome organization and are enriched for regulatory elements in the genome sequences of other mammalian species. Importantly, we developed a filtering strategy, including a machine learning classification method, to utilize a very small number of species-specific experimental datasets available to select for the likely active regulatory regions. The method finds the optimal combination of sensitivity and accuracy to unbiasedly predict regulatory regions in mammalian species. Furthermore, we demonstrated the utility of the predicted regulatory datasets in cattle for prioritizing variants associated with multiple production and climate change adaptation traits, and identifying potential genome editing targets.

genomics↗