Biology subjects

Ang, C. E.

Publications and source records attributed to Ang, C. E..

5 recordsLinked to original sources

bioRxiv2022

Spatially resolved epigenomic profiling of single cells in complex tissues

The recent development of spatial omics methods enables single-cell profiling of the transcriptome and the 3D genome organization in a spatially resolved manner. Expanding the repertoire of spatial omics tools, a spatial epigenomics method will accelerate our understanding of the spatial regulation of cell and tissue functions. Here, we report a method for spatially resolved profiling of epigenomes in single cells using in-situ tagmentation and transcription followed by highly multiplexed imaging. We profiled histone modifications marking active promoters and enhancers, H3K4me3 and H3K27ac, and generated high-resolution spatial atlas of hundreds of active promoters and putative enhancers in embryonic and adult mouse brains. Our results further revealed putative promoter-enhancer pairs and enhancer hubs regulating the expression of developmentally important genes. We envision this approach will be generally applicable to spatial profiling of epigenetic modifications and DNA-binding proteins, advancing our understanding of how gene expression is spatiotemporally regulated by the epigenome.

genomics↗

bioRxiv2021

Direct induction of human neurons from fibroblasts carrying the neuropsychiatric 22q11.2 microdeletion reveals transcriptome- and epigenome-wide alterations

Standard methods for the creation of neuronal cells via direct induction from primary tissue use perinatal fibroblasts, which hinders the important study of patient specific genetic lesions such as those underlying neuropsychiatric disorders. To address this we developed a novel method for the direct induction of neuronal cells (induced neuronal cells, iN cells) from adult human fibroblast cells. Reprogramming fibroblasts into iN cells via recombinant virus resulted in cells that stain for markers such as MAP2 and PSA-NCAM and exhibit electrophysiological properties such as action potentials and voltage dependent sodium- and potassium currents that reveal a neuronal phenotype. Transcriptome and chromatin analysis using RNA-Seq, microRNA-Seq and ATAC-Seq, respectively, further confirm neuronal character. 22q11.2 Deletion-Syndrome (22q11DS) is caused by a large 3 million base-pair heterozygous deletion on human chromosome 22 and is strongly associated with neurodevelopmental, neuropsychiatric phenotypes such as schizophrenia and autism. We leverage the direct-iN cell model for the study of genetic neurodevelopmental conditions by presenting gene-by-gene as well as networkwide effects of the 22q11DS deletion on gene expression in human neuronal cells, on several levels of functional genomics analysis. Some of the genes within the 22q11DS deletion boundary exhibit unexpected cell-type-specific changes in transcript levels, and genome-wide we can detect dysregulation of calcium channel subunit genes and other genes known to be involved in autism or schizophrenia, such as NRXN1, as well synaptic pathways. This genome-wide effect on gene expression can also be observed at the microRNA and chromatin levels, showing that the iN cells have indeed converted to a neuronal phenotype at several regulatory levels: chromatin, protein-coding RNAs and microRNAs, revealing relevant disease pathways and genes. We present this model of inducing neurons from fibroblasts as a useful general resource to study the genetic and molecular basis of normal and abnormal brain development and brain function.

genomics↗

bioRxiv2020

THE DYNAMIC INTERPLAY BETWEEN HOMEODOMAIN TRANSCRIPTION FACTORS AND CHROMATIN ENVIRONMENT REGULATES PRONEURAL FACTOR OUTCOMES

Generation of neurons of vast diversity involves early spatial and temporal patterning of the neuronal precursors by morphogenic gradients and combinatorial expression of transcription factors. While the proneuronal function of the basic-helix-loop-helix (bHLH) transcription factor Ngn2 is well established, its role in neuronal subtype specification remains unclear. Here, we found that coexpressing NGN2 with the forebrain homeobox factor EMX1 converts human pluripotent stem cells into a highly homogeneous glutamatergic forebrain neurons without partial cholinergic and monoaminergic gene programs observed in cells infected with NGN2 only. Our molecular characterization revealed that transcriptional output and genomic targeting of Ngn2 is altered by co-factors such as EMX1 explaining the more focused subtype specification. Ngn2 function is less modified by the chromatin environment and does not affect regionalization of pre-patterned neural progenitors. These results enable improved strategies for generating a plethora of defined neuronal subpopulations from pluripotent stem cells for therapeutic or disease-modeling purposes. HighlightsO_LINGN2 converts human ES cells into glutamatergic neurons some of which co-express a partial cholinergic program C_LIO_LINGN2 directly binds to and activates ISL1 in ES cells which together with PHOX2A/B induce cholinergic genes C_LIO_LIAnterior-posterior regionalization affects NGN2 binding and transcriptional output but does not focus subtype specification C_LIO_LIForebrain homeobox factors including EMX1 and FOXG1 redirect NGN2 chromatin binding and repress posterior and cholinergic genes, resulting in homogeneous forebrain excitatory neurons C_LI

genomics↗

bioRxiv2018

Old fibroblasts secrete inflammatory cytokines that drive variability in reprogramming efficiency and may affect wound healing between old individuals

Age-associated chronic inflammation (inflammaging) has emerged as a central hallmark of aging1-3, but its impact on specific cells is still largely unknown. Fibroblasts are present in all tissues and contribute to wound healing4-6. They are also the cell type that is mostly used for induced pluripotent stem cell (iPSC) reprogramming7 - a process that has implications for regenerative medicine and rejuvenation strategies8-17. Here we show that primary fibroblasts from old mice secrete inflammatory cytokines and that there is an increased variability in reprogramming efficiency between fibroblast cultures from old individuals. Individual-to-individual variability is emerging as a key feature of old age18-21, which could reflect distinct aging trajectories, but the underlying causes remain unknown. To identify drivers of this variability, we perform a multi-omic assessment of young and old fibroblast cultures with different reprogramming efficiency. This approach, coupled with single cell transcriptomics, reveals that old fibroblast cultures are heterogeneous and show a greater proportion of activated fibroblasts that secrete inflammatory cytokines, which correlates with reprogramming efficiency. We experimentally validate that activated fibroblasts express inflammatory cytokines in vivo and that their presence is linked to enhanced reprogramming efficiency in culture. Conditioned-media swapping experiments show that extrinsic factors secreted by activated fibroblasts are more critical than intrinsic factors for the individual-to-individual variability in reprogramming efficiency, and we identify TNF as a key inflammatory cytokine underlying this variability. Interestingly, old mice also exhibit variability in wound healing efficiency in vivo and old wounds show an increased subpopulation of activated fibroblasts with a unique TNF signature. Our study shows that a switch in fibroblast composition, and the ratio of inflammatory cytokines they secrete, drives variability in reprogramming in vitro and may influence wound healing in vivo. These findings could help identify personalized strategies to improve iPSC generation and wound healing in older individuals.

cell biology↗

bioRxiv2018

The novel lncRNA lnc-NR2F1 is pro-neurogenic and mutated in human neurodevelopmental disorders

Long noncoding RNAs (lncRNAs) have been shown to act as important cell biological regulators including cell fate decisions but are often ignored in human genetics. Combining differential lncRNA expression during neuronal lineage induction with copy number variation morbidity maps of a cohort of children with autism spectrum disorder/intellectual disability versus healthy controls revealed focal genomic mutations affecting several lncRNA candidate loci. Here we find that a t(5:12) chromosomal translocation in a family manifesting neurodevelopmental symptoms disrupts specifically lnc-NR2F1. We further show that lnc-NR2F1 is an evolutionarily conserved lncRNA functionally enhances induced neuronal cell maturation and directly occupies and regulates transcription of neuronal genes including autism-associated genes. Thus, integrating human genetics and functional testing in neuronal lineage induction is a promising approach for discovering candidate lncRNAs involved in neurodevelopmental diseases.

neuroscience↗