Eukaryotic genomes are organised into complex higher-order structures within the nucleus, and the three-dimensional arrangement of chromosomes is functionally important for global gene regulation. The existence of supernumerary chromosome 21 in Down syndrome may perturb the nuclear architecture at different levels, which is normally optimised to maintain the physiological balance of gene expression. However, it has not been clearly elucidated whether and how aberrant configuration of chromosomes affects gene activities. To investigate the effects of trisomy 21 on nuclear organisation and gene expression, we performed three-dimensional fluorescent imaging analysis of chromosome-edited human induced pluripotent stem cells (iPSCs), which enabled identification of the parental origin of the three copies of chromosome 21. We found that two copies of maternal chromosomes resulting from meiotic nondisjunction had a higher tendency to form an adjacent pair and were located relatively distant from the nuclear membrane, suggesting the conserved interaction between these homologous chromosomes. Transcriptional profiling of parental-origin-specific corrected disomy 21 iPSC lines indicated upregulated expression of the maternal alleles for a group of genes, which was accompanied by a fluctuating expression pattern. These results suggest the unique effects of a pair of maternal chromosomes in trisomy 21, which may contribute to the pathological phenotype.

Chromosome abnormalities induces profound alterations in gene expression, leading to various disease phenotypes. Recent studies on yeast and mammalian cells have demonstrated that aneuploidy exerts detrimental effects on organismal growth and development, regardless of the karyotype, suggesting that aneuploidy-associated stress plays an important role in disease pathogenesis. However, whether and how this effect alters cellular homeostasis and long-term features of human disease are not fully understood. Here, we aimed to investigate cellular stress responses in human trisomy syndromes, using fibroblasts and induced pluripotent stem cells (iPSCs). Dermal fibroblasts derived from patients with trisomy 21, 18 and 13 showed a severe impairment of cell proliferation and enhanced premature senescence. These phenomena were accompanied by perturbation of protein homeostasis, leading to the accumulation of protein aggregates. We found that treatment with sodium 4-phenylbutyrate (4-PBA), a chemical chaperone, decreased the protein aggregates in trisomy fibroblasts. Notably, 4-PBA treatment successfully prevented the progression of premature senescence in secondary fibroblasts derived from trisomy 21 iPSCs. Our study reveals aneuploidy-associated stress as a potential therapeutic target for human trisomies, including Down syndrome.

Individuals with Down syndrome (DS) commonly show unique pathological phenotypes throughout their life span. Besides the specific effects of dosage-sensitive genes on chromosome 21, recent studies have demonstrated that the gain of a chromosome exerts an adverse impact on cell physiology, regardless of the karyotype. Although dysregulated transcription and perturbed protein homeostasis are observed in common in human fibroblasts with trisomy 21, 18, and 13, whether and how this aneuploidy-associated stress acts on other cell lineages and affects the pathophysiology are unknown. Here, we investigated cellular stress responses in human trisomy 21 and 13 neurons differentiated from patient-derived induced pluripotent stem cells. Neurons of both trisomies showed increased vulnerability to apoptotic cell death, accompanied by dysregulated protein homeostasis and upregulation of the endoplasmic reticulum stress pathway. In addition, misfolded protein aggregates, comprising various types of neurodegenerative disease-related proteins, were abnormally accumulated in trisomic neurons. Intriguingly, treatment with sodium 4-phenylbutyrate, a chemical chaperone, successfully decreased the formation of protein aggregates and prevented the progression of cell apoptosis in trisomic neurons. These results suggest that aneuploidy-associated stress might be a therapeutic target for the neurodegenerative phenotypes in DS. Down syndrome (DS) is the most common genetic chromosomal disorder, affecting approximately one in 700-800 newborns 1. All individuals with DS exhibit cognitive and learning deficits, with broad variation in phenotypes and severities 2. Studies using in vitro and in vivo experiments with postmortem brains, neural stem/ progenitor cells (NPCs), or induced pluripotent stem cells (iPSCs) derived from DS patients and DS mouse models have shown various levels of neuropathology, including reduced neurogenesis, impaired neuronal maturation, and accelerated neural cell death. These features arise from the extra copy of human chromosome 21 (Hsa21), and specific DS phenotypes are generally thought to be derived from the increased expression of a specific subset of dosage-sensitive genes on Hsa21 3. Based on this 'gene dosage imbalance' hypothesis, multiple studies have been conducted to identify the genes responsible for impaired neural development 4-6. Human neural progenitors isolated from foetal brains with DS show vulnerability to oxidative stress and preference for gliogenesis, which is accompanied by elevated expression levels of S100B and APP genes 7. The DYRK1A gene is reported to affect the differentiation of mouse cortical NPCs and play a critical role in brain morphogenesis 8. The administration

Astrocytes exert adverse effects on the brains of individuals with Down syndrome (DS). Although a neurogenic-to-gliogenic shift in the fate-specification step has been reported, the mechanisms and key regulators underlying the accelerated proliferation of astrocyte precursor cells (APCs) in DS remain elusive. Here, we established a human isogenic cell line panel based on DS-specific induced pluripotent stem cells, the XIST-mediated transcriptional silencing system in trisomic chromosome 21, and genome/chromosome-editing technologies to eliminate phenotypic fluctuations caused by genetic variation. The transcriptional responses of genes observed upon XIST induction and/or downregulation are not uniform, and only a small subset of genes show a characteristic expression pattern, which is consistent with the proliferative phenotypes of DS APCs. Comparative analysis and experimental verification using gene modification reveal dose-dependent proliferation-promoting activity of DYRK1A and PIGP on DS APCs. Our collection of human isogenic cell lines provides a comprehensive set of cellular models for further DS investigations.

Cerebrovasculature is critical in maintaining brain homeostasis; its dysregulation often leads to vascular cognitive impairment and dementia (VCID) during aging. VCID is the second most prevalent cause of dementia in the elderly, after Alzheimer’s disease (AD), with frequent cooccurrence of VCID and AD. While multiple factors are involved in the pathogenesis of AD and VCID, APOE4 increases the risk for both diseases. A major apolipoprotein E (apoE) receptor, the low-density lipoprotein receptor-related protein 1 (LRP1), is abundantly expressed in vascular mural cells (pericytes and smooth muscle cells). Here, we investigated how deficiency of vascular mural cell LRP1 affects the cerebrovascular system and cognitive performance using vascular mural cell–specific Lrp1 -KO mice ( smLrp1 –/– ) in a human APOE3 or APOE4 background. We found that spatial memory was impaired in the 13- to 16-month-old APOE4 smLrp1 –/– mice but not in the APOE3 smLrp1 –/– mice, compared with their respective littermate control mice. These disruptions in the APOE4 smLrp1 –/– mice were accompanied with excess paravascular glial activation and reduced cerebrovascular collagen IV. In addition, blood-brain barrier (BBB) integrity was disrupted in the APOE4 smLrp1 –/– mice. Together, our results suggest that vascular mural cell LRP1 modulates cerebrovasculature integrity and function in an APOE genotype–dependent manner.