Vast emerging evidences are linking the base modifications and determination of stem cell fate such as proliferation and differentiation. Among the base modification markers extensively studied, 5-methylcytosine (5-mC) and its oxidative derivatives (5-hydroxymethylcytosine (5-hmC), 5-formylcytosine (5-fC), and 5-carboxylcytosine (5-caC)) dynamically occur in DNA and RNA and have been acknowledged as important epigenetic markers involved in regulation of cellular biological processes. N6-Methyladenosine modification in DNA (m6dA), mRNA (m6A), tRNA, and other noncoding RNAs has been defined as another important epigenetic and epitranscriptomic marker in eukaryotes in recent years. The mRNA m6A modification has been characterized biochemically, molecularly, and phenotypically, including elucidation of its methyltransferase complexes (m6A writer), demethylases (m6A eraser), and direct interaction proteins (readers), while limited information on the DNA m6dA is available. The levels and the landscapes of m6A in the epitranscriptomes and epigenomes are precisely and dynamically regulated by the fine-tuned coordination of the writers and erasers in accordance with stages of the growth, development, and reproduction as naturally programmed during the lifespan. Additionally, progress has been made in appreciation of the link between aberrant m6A modification in stem cells and diseases, like cancers and neurodegenerative disorders. These achievements are inspiring scientists to further uncover the epigenetic mechanisms for stem cell development and to dissect pathogenesis of the multiple diseases conferred by development aberration of the stem cells. This review article will highlight the research advances in the role of m6A methylation modifications of DNA and RNA in the regulation of stem cell and genesis of the closely related disorders. Additionally, this article will also address the research directions in the future.

GGC repeat expansions within NOTCH2NLC have been identified as the genetic cause of neuronal intranuclear inclusion disease (NIID). To understand the molecular pathogenesis of NIID, here, we established both a transgenic mouse model and a human neural progenitor cells (hNPCs) model. Expression of the NOTCH2NLC with expanded GGC repeats produced widespread intranuclear and perinuclear polyglycine (polyG), polyalanine (polyA), and polyarginine (polyR) inclusions, leading to behavioral deficits and severe neurodegeneration, which faithfully mimicked the clinical and pathological features associated with NIID. Furthermore, conserved alternative splicing events were identified between the NIID mouse and hNPC models, among which was the enrichment of the binding motifs of hnRNPM, an RNA binding protein known as alternative splicing regulator. Expanded NOTCH2NLC-polyG and NOTCH2NLC-polyA could interact with and sequester hnRNPM, while overexpression of hnRNPM could ameliorate the cellular toxicity. These results together suggested that dysfunction of hnRNPM could play an important role in the molecular pathogenesis of NIID.

Parkinson’s disease (PD) is the second most common neurodegenerative disorder, and aging and genetic and environmental exposure can contribute to its pathogenesis. DNA methylation has been suggested to play a pivotal role in neurodevelopment and neurodegenerative diseases. 5-hydroxymethylcytosine (5hmC) is generated through 5-methylcytosine (5mC) oxidization by ten-eleven translocation proteins and is particularly enriched in the brain. Although 5hmC has been linked to multiple neurological disorders, little is known about 5hmC alterations in the substantia nigra of patients with PD. To determine the specific alterations in DNA methylation and hydroxymethylation in PD brain samples, we examined the genome-wide profiles of 5mC and 5hmC in the substantia nigra of patients with PD and Alzheimer’s disease (ad). We identified 4119 differentially hydroxymethylated regions (DhMRs) and no differentially methylated regions (DMRs) in the postmortem brains of patients with PD compared with those of controls. These DhMRs were PD-specific when compared with the results of AD. Gene ontology analysis revealed that several signaling pathways, such as neurogenesis and neuronal differentiation, were significantly enriched in PD DhMRs. KEGG enrichment analysis revealed substantial alterations in multiple signaling pathways, including phospholipase D (PLD), cAMP and Rap1. In addition, using a PD Drosophila model, we found that one of the 5hmC-modulated genes, PLD1, modulated α-synuclein toxicity. Our analysis suggested that 5hmC may act as an independent epigenetic marker and contribute to the pathogenesis of PD.

The epigenetic memory stored in the dynamic modifications, such as base modifications of cytosine (C) in DNA, including methylation/hydroxymethylation/demethylation, causes heritable phenotypes via regulating gene expression without alteration of DNA sequence. The process from cytosine modification to the epigenetic effect is orchestrated by complicated machinery consisting of writers, erasers, readers, and other factors. The two major forms of cytosine modification include methylcytosine (5-mC) and hydroxymethylcytosine (5-hmC). DNA methyltransferases (DNMTs) including DNMT1, DNMT3A, and DNMT3B function as writers for 5-mC. The ten-eleven translocation proteins (TET) including TET1, TET2, and TET3 in the mammalian genome are responsible for hydroxymethylation of 5-mC to generate 5-hmC, 5-formylcytosine (5-fC), and 5-carboxylcytosine (5-caC). The 5-mC and 5-hmC have become the two most extensively investigated epigenetic markers, and the dynamic balance of these two markers shape the landscape of the epigenome, functioning as a platform to regulate gene expression epigenetically. The landscape of the 5-hmC in epigenome is precisely and tightly regulated during the development. Aberrant alterations of the epigenetic regulation may cause severe consequences such as phenotype change as well as initiation of disease. Progressively, significant achievements have been made in characterization of writers, erasers, and readers of 5-mC and 5-hmC, as well as the contribution of aberrant alteration of 5-hmC/5-mC landscape to the pathogenesis of human diseases, such as cancers and neurological disorders. This article will highlight the research advances in the distinct contribution of TET proteins as suppressors or promoters to the pathogenesis of tumorigenesis and progression. Furthermore, this article also discusses the challenges and the directions for research in the future.

5-hydroxymethylcytosine, also known as the sixth DNA base of the genome, plays an important role in brain aging and neurological disorders such as Alzheimer’s disease. However, little is known about its genome-wide distribution and its association with Alzheimer’s disease pathology. Here, we report a genome-wide profiling of 5-hydroxymethylcytosine in 1079 autopsied brains (dorsolateral prefrontal cortex) of older individuals and assess its association with multiple measures of Alzheimer’s disease pathologies, including pathological diagnosis of Alzheimer’s disease, amyloid-β load, and PHFtau tangle density. Of 197,765 5-hydroxymethylcytosine regions detected, we identified 2821 differentially hydroxymethylated regions associated with Alzheimer’s disease neuropathology after controlling for multiple testing and covariates. Many differentially hydroxymethylated regions are located within known Alzheimer’s disease loci, such as RIN3, PLCG2, ITGA2B, and USP6NL. Integrative multi-omics analyses support a potential mechanistic role of 5-hydroxymethylcytosine alterations in Alzheimer’s disease. Our study presents a large-scale genome-wide atlas of 5-hydroxymethylcytosine in Alzheimer’s brain and offers insight into the mechanism underlying Alzheimer’s disease pathogenesis.

Clinical trials serve as the fundamental prerequisite for clinical therapy of human disease, which is primarily based on biomedical studies in animal models. Undoubtedly, animal models have made a significant contribution to gaining insight into the developmental and pathophysiological understanding of human diseases. However, none of the existing animal models could efficiently simulate the development of human organs and systems due to a lack of spatial information; the discrepancy in genetic, anatomic, and physiological basis between animals and humans limits detailed investigation. Therefore, the translational efficiency of the research outcomes in clinical applications was significantly weakened, especially for some complex, chronic, and intractable diseases. For example, the clinical trials for human fragile X syndrome (FXS) solely based on animal models have failed such as mGluR5 antagonists. To mimic the development of human organs more faithfully and efficiently translate in vitro biomedical studies to clinical trials, extensive attention to organoids derived from stem cells contributes to a deeper understanding of this research. The organoids are a miniaturized version of an organ generated in vitro, partially recapitulating key features of human organ development. As such, the organoids open a novel avenue for in vitro models of human disease, advantageous over the existing animal models. The invention of organoids has brought an innovative breakthrough in regeneration medicine. The organoid-derived human tissues or organs could potentially function as invaluable platforms for biomedical studies, pathological investigation of human diseases, and drug screening. Importantly, the study of regeneration medicine and the development of therapeutic strategies for human diseases could be conducted in a dish, facilitating in vitro analysis and experimentation. Thus far, the pilot breakthrough has been made in the generation of numerous types of organoids representing different human organs. Most of these human organoids have been employed for in vitro biomedical study and drug screening. However, the efficiency and quality of the organoids in recapitulating the development of human organs have been hindered by engineering and conceptual challenges. The efficiency and quality of the organoids are essential for downstream applications. In this article, we highlight the application in the modeling of human neurodegenerative diseases (NDDs) such as FXS, Alzheimer’s disease (AD), Parkinson’s disease (PD), and autistic spectrum disorders (ASD), and organoid-based drug screening. Additionally, challenges and weaknesses especially for limits of the brain organoid models in modeling late onset NDDs such as AD and PD., and future perspectives regarding human brain organoids are addressed.