Necl-1/CADM3 regulates cone synapse formation in the mouse retina.

In vertebrates, retinal neural circuitry for visual perception is organized in specific layers. The outer plexiform layer is the first synaptic region in the visual pathway, where photoreceptor synaptic terminals connect with bipolar and horizontal cell processes. However, molecular mechanisms underlying cone synapse formation to mediate OFF pathways remain unknown. This study reveals that Necl-1/CADM3 is localized at S- and S/M-opsin-containing cones and dendrites of type 4 OFF cone bipolar cells (CBCs). In Necl-1-/- mouse retina, synapses between cones and type 4 OFF CBCs were dislocated, horizontal cell distribution became abnormal, and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors were dislocated. Necl-1-/- mice exhibited aberrant short-wavelength-light-elicited signal transmission from cones to OFF CBCs, which was rescued by AMPA receptor potentiator. Additionally, Necl-1-/- mice showed impaired optokinetic responses. These findings suggest that Necl-1 regulates cone synapse formation to mediate OFF cone pathways elicited by short-wavelength light in mouse retina.

NTRK2 expression in gastrointestinal stromal tumors with a special emphasis on the clinicopathological and prognostic impacts.

Gastrointestinal stromal tumors (GISTs) are typically characterized by activating mutations of the KIT proto-oncogene receptor tyrosine kinase (KIT) or platelet-derived growth factor receptor alpha (PDGFRA). Recently, the neurotrophic tyrosine receptor kinase (NTRK) fusion was reported in a small subset of wild-type GIST. We examined trk IHC and NTRK gene expressions in GIST. Pan-trk immunohistochemistry (IHC) was positive in 25 (all 16 duodenal and 9 out of 16 small intestinal GISTs) of 139 cases, and all pan-trk positive cases showed diffuse and strong expression of c-kit. Interestingly, all of these cases showed only trkB but not trkA/trkC expression. Cap analysis of gene expression (CAGE) analysis identified increased number of genes whose promoters were activated in pan-trk/trkB positive GISTs. Imbalanced expression of NTRK2, which suggests the presence of NTRK2 fusion, was not observed in any of trkB positive GISTs, despite higher mRNA expression. TrkB expression was found in duodenal GISTs and more than half of small intestinal GISTs, and this subset of cases showed poor prognosis. However, there was not clear difference in clinical outcomes according to the trkB expression status in small intestinal GISTs. These findings may provide a possible hypothesis for trkB overexpression contributing to the tumorigenesis and aggressive clinical outcome in GISTs of duodenal origin.

Regulation of lymphoid-myeloid lineage bias through regnase-1/3-mediated control of Nfkbiz.

ABSTRACT: Regulation of lineage biases in hematopoietic stem and progenitor cells (HSPCs) is pivotal for balanced hematopoietic output. However, little is known about the mechanism behind lineage choice in HSPCs. Here, we show that messenger RNA (mRNA) decay factors regnase-1 (Reg1; Zc3h12a) and regnase-3 (Reg3; Zc3h12c) are essential for determining lymphoid fate and restricting myeloid differentiation in HSPCs. Loss of Reg1 and Reg3 resulted in severe impairment of lymphopoiesis and a mild increase in myelopoiesis in the bone marrow. Single-cell RNA sequencing analysis revealed that Reg1 and Reg3 regulate lineage directions in HSPCs via the control of a set of myeloid-related genes. Reg1- and Reg3-mediated control of mRNA encoding Nfkbiz, a transcriptional and epigenetic regulator, was essential for balancing lymphoid/myeloid lineage output in HSPCs in vivo. Furthermore, single-cell assay for transposase-accessible chromatin sequencing analysis revealed that Reg1 and Reg3 control the epigenetic landscape on myeloid-related gene loci in early stage HSPCs via Nfkbiz. Consistently, an antisense oligonucleotide designed to inhibit Reg1- and Reg3-mediated Nfkbiz mRNA degradation primed hematopoietic stem cells toward myeloid lineages by enhancing Nfkbiz expression. Collectively, the collaboration between posttranscriptional control and chromatin remodeling by the Reg1/Reg3-Nfkbiz axis governs HSPC lineage biases, ultimately dictating the fate of lymphoid vs myeloid differentiation.

Morphomics via next-generation electron microscopy.

The living body is composed of innumerable fine and complex structures. Although these structures have been studied in the past, a vast amount of information pertaining to them still remains unknown. When attempting to observe these ultra-structures, the use of electron microscopy (EM) has become indispensable. However, conventional EM settings are limited to a narrow tissue area, which can bias observations. Recently, new trends in EM research have emerged that provide coverage of far broader, nano-scale fields of view for two-dimensional wide areas and three-dimensional large volumes. Moreover, cutting-edge bioimage informatics conducted via deep learning has accelerated the quantification of complex morphological bioimages. Taken together, these technological and analytical advances have led to the comprehensive acquisition and quantification of cellular morphology, which now arises as a new omics science termed 'morphomics'.

Tertiary Lymphoid Tissues Are Microenvironments with Intensive Interactions between Immune Cells and Proinflammatory Parenchymal Cells in Aged Kidneys.

SIGNIFICANCE STATEMENT: Ectopic lymphoid structures called tertiary lymphoid tissues (TLTs) develop in several kidney diseases and are associated with poor renal prognosis. However, the mechanisms underlying TLT expansion and their effect on renal regeneration remain unclear. The authors report that single-nucleus RNA sequencing and validation experiments demonstrate that TLTs potentially amplify inflammation in aged injured kidneys. Lymphocytes within TLTs promote proinflammatory phenotypes of the surrounding proximal tubules and fibroblasts within the TLTs via proinflammatory cytokine production. These proinflammatory parenchymal cells then interact with immune cells by chemokine or cytokine production. Such cell-cell interactions potentially increase inflammation, expand TLTs, and exacerbate kidney injury. These findings help illuminate renal TLT pathology and suggest potential therapeutic targets. BACKGROUND: Ectopic lymphoid structures called tertiary lymphoid tissues (TLTs) develop in several kidney diseases and are associated with poor renal prognosis. However, the mechanisms that expand TLTs and underlie exacerbation of kidney injury remain unclear. METHODS: We performed single-nucleus RNA sequencing (snRNA-seq) on aged mouse kidneys with TLTs after ischemia-reperfusion injury. The results were validated using immunostaining, in situ hybridization of murine and human kidneys, and in vitro experiments. RESULTS: Using snRNA-seq, we identified proinflammatory and profibrotic Vcam1+ injured proximal tubules (PTs) with NF κ B and IFN-inducible transcription factor activation. VCAM1 + PTs were preferentially localized around TLTs and drove inflammation and fibrosis via the production of multiple chemokines or cytokines. Lymphocytes within TLTs expressed Tnf and Ifng at high levels, which synergistically upregulated VCAM1 and chemokine expression in cultured PT cells. In addition, snRNA-seq also identified proinflammatory and profibrotic fibroblasts, which resided within and outside TLTs, respectively. Proinflammatory fibroblasts exhibited STAT1 activation and various chemokine or cytokine production, including CXCL9/CXCL10 and B cell-activating factor, contributing to lymphocyte recruitment and survival. IFN γ upregulated the expression of these molecules in cultured fibroblasts in a STAT1-dependent manner, indicating potential bidirectional interactions between IFN γ -producing CXCR3 + T cells and proinflammatory fibroblasts within TLTs. The cellular and molecular components described in this study were confirmed in human kidneys with TLTs. CONCLUSIONS: These findings suggest that TLTs potentially amplify inflammation by providing a microenvironment that allows intense interactions between renal parenchymal and immune cells. These interactions may serve as novel therapeutic targets in kidney diseases involving TLT formation.

Identification of BST2 as a conjunctival epithelial stem/progenitor cell marker.

The conjunctival epithelium consists of conjunctival epithelial cells and goblet cells derived from conjunctival epithelial stem/progenitor cells. However, the source of these cells is not well known because no specific markers for conjunctival epithelial stem/progenitor cells have been discovered. Therefore, to identify conjunctival epithelial stem/progenitor cell markers, we performed single-cell RNA sequencing of a conjunctival epithelial cell population derived from human-induced pluripotent stem cells (hiPSCs). The following conjunctival epithelial markers were identified: BST2, SLC2A3, AGR2, TMEM54, OLR1, and TRIM29. Notably, BST2 was strongly positive in the basal conjunctival epithelium, which is thought to be rich in stem/progenitor cells. Moreover, BST2 was able to sort conjunctival epithelial stem/progenitor cells from hiPSC-derived ocular surface epithelial cell populations. BST2-positive cells were highly proliferative and capable of successfully generating conjunctival epithelial sheets containing goblet cells. In conclusion, BST2 has been identified as a specific marker of conjunctival epithelial stem/progenitor cells.

Mobile element variation contributes to population-specific genome diversification, gene regulation and disease risk.

Mobile genetic elements (MEs) are heritable mutagens that recursively generate structural variants (SVs). ME variants (MEVs) are difficult to genotype and integrate in statistical genetics, obscuring their impact on genome diversification and traits. We developed a tool that accurately genotypes MEVs using short-read whole-genome sequencing (WGS) and applied it to global human populations. We find unexpected population-specific MEV differences, including an Alu insertion distribution distinguishing Japanese from other populations. Integrating MEVs with expression quantitative trait loci (eQTL) maps shows that MEV classes regulate tissue-specific gene expression by shared mechanisms, including creating or attenuating enhancers and recruiting post-transcriptional regulators, supporting class-wide interpretability. MEVs more often associate with gene expression changes than SNVs, thus plausibly impacting traits. Performing genome-wide association study (GWAS) with MEVs pinpoints potential causes of disease risk, including a LINE-1 insertion associated with keloid and fasciitis. This work implicates MEVs as drivers of human divergence and disease risk.

Comprehensive characterization of the embryonic factor LEUTX.

The paired-like homeobox transcription factor LEUTX is expressed in human preimplantation embryos between the 4- and 8-cell stages, and then silenced in somatic tissues. To characterize the function of LEUTX, we performed a multiomic characterization of LEUTX using two proteomics methods and three genome-wide sequencing approaches. Our results show that LEUTX stably interacts with the EP300 and CBP histone acetyltransferases through its 9 amino acid transactivation domain (9aaTAD), as mutation of this domain abolishes the interactions. LEUTX targets genomic cis-regulatory sequences that overlap with repetitive elements, and through these elements it is suggested to regulate the expression of its downstream genes. We find LEUTX to be a transcriptional activator, upregulating several genes linked to preimplantation development as well as 8-cell-like markers, such as DPPA3 and ZNF280A. Our results support a role for LEUTX in preimplantation development as an enhancer binding protein and as a potent transcriptional activator.

HTLV-1 bZIP factor impairs DNA mismatch repair system.

Adult T-cell leukemia (ATL) is a peripheral T-cell malignancy caused by human T-cell leukemia virus type 1 (HTLV-1). Microsatellite instability (MSI) has been observed in ATL cells. Although MSI results from impaired mismatch repair (MMR) pathway, no null mutations in the genes encoding MMR factors are detectable in ATL cells. Thus, it is unclear whether or not impairment of MMR causes the MSI in ATL cells. HTLV-1 bZIP factor (HBZ) protein interacts with numerous host transcription factors and significantly contributes to disease pathogenesis and progression. Here we investigated the effect of HBZ on MMR in normal cells. The ectopic expression of HBZ in MMR-proficient cells induced MSI, and also suppressed the expression of several MMR factors. We then hypothesized that the HBZ compromises MMR by interfering with a transcription factor, nuclear respiratory factor 1 (NRF-1), and identified the consensus NRF-1 binding site at the promoter of the gene encoding MutS homologue 2 (MSH2), an essential MMR factor. The luciferase reporter assay revealed that NRF-1 overexpression enhanced MSH2 promoter activity, while co-expression of HBZ reversed this enhancement. These results supported the idea that HBZ suppresses the transcription of MSH2 by inhibiting NRF-1. Our data demonstrate that HBZ causes impaired MMR, and may imply a novel oncogenesis driven by HTLV-1.

ATM suppresses c-Myc overexpression in the mammary epithelium in response to estrogen.

ATM gene mutation carriers are predisposed to estrogen-receptor-positive breast cancer (BC). ATM prevents BC oncogenesis by activating p53 in every cell; however, much remains unknown about tissue-specific oncogenesis after ATM loss. Here, we report that ATM controls the early transcriptional response to estrogens. This response depends on topoisomerase II (TOP2), which generates TOP2-DNA double-strand break (DSB) complexes and rejoins the breaks. When TOP2-mediated ligation fails, ATM facilitates DSB repair. After estrogen exposure, TOP2-dependent DSBs arise at the c-MYC enhancer in human BC cells, and their defective repair changes the activation profile of enhancers and induces the overexpression of many genes, including the c-MYC oncogene. CRISPR/Cas9 cleavage at the enhancer also causes c-MYC overexpression, indicating that this DSB causes c-MYC overexpression. Estrogen treatment induced c-Myc protein overexpression in mammary epithelial cells of ATM-deficient mice. In conclusion, ATM suppresses the c-Myc-driven proliferative effects of estrogens, possibly explaining such tissue-specific oncogenesis.

Prediction of the cell-type-specific transcription of non-coding RNAs from genome sequences via machine learning.

Gene transcription is regulated through complex mechanisms involving non-coding RNAs (ncRNAs). As the transcription of ncRNAs, especially of enhancer RNAs, is often low and cell type specific, how the levels of RNA transcription depend on genotype remains largely unexplored. Here we report the development and utility of a machine-learning model (MENTR) that reliably links genome sequence and ncRNA expression at the cell type level. Effects on ncRNA transcription predicted by the model were concordant with estimates from published studies in a cell-type-dependent manner, regardless of allele frequency and genetic linkage. Among 41,223 variants from genome-wide association studies, the model identified 7,775 enhancer RNAs and 3,548 long ncRNAs causally associated with complex traits across 348 major human primary cells and tissues, such as rare variants plausibly altering the transcription of enhancer RNAs to influence the risks of Crohn's disease and asthma. The model may aid the discovery of causal variants and the generation of testable hypotheses for biological mechanisms driving complex traits.

Compact laboratory-based X-ray microscope enabling nondestructive 3D structure acquisition of mouse nephron with high speed and better user accessibility.

X-ray microscopes adopting computed tomography enable nondestructive 3D visualization of biological specimens at micron-level resolution without conventional 2D serial sectioning that is a destructive/laborious method and is routinely used for analyzing renal biopsy in clinical diagnosis of kidney diseases. Here we applied a compact commercial system of laboratory-based X-ray microscope to observe a resin-embedded osmium-stained 1-mm strip of a mouse kidney piece as a model of renal biopsy, toward a more efficient diagnosis of kidney diseases. A reconstructed computed tomography image from several hours of data collection using CCD detector allowed us to unambiguously segment a single nephron connected to a renal corpuscle, which was consistent with previous reports using serial sectioning. Histogram analysis on the segmented nephron confirmed that the proximal and distal tubules were distinguishable on the basis of their X-ray opacities. A 3D rendering model of the segmented nephron visualized a convoluted structure of renal tubules neighboring the renal corpuscle and a branched structure of efferent arterioles. Furthermore, another data collection using scientific complementary metal-oxide semiconductor detector with a much shorter data acquisition time of 15 min provided similar results from the same samples. These results suggest a potential application of the compact laboratory-based X-ray microscope to analyze mouse renal biopsy.

Nucleome programming is required for the foundation of totipotency in mammalian germline development.

Germ cells are unique in engendering totipotency, yet the mechanisms underlying this capacity remain elusive. Here, we perform comprehensive and in-depth nucleome analysis of mouse germ-cell development in vitro, encompassing pluripotent precursors, primordial germ cells (PGCs) before and after epigenetic reprogramming, and spermatogonia/spermatogonial stem cells (SSCs). Although epigenetic reprogramming, including genome-wide DNA de-methylation, creates broadly open chromatin with abundant enhancer-like signatures, the augmented chromatin insulation safeguards transcriptional fidelity. These insulatory constraints are then erased en masse for spermatogonial development. Notably, despite distinguishing epigenetic programming, including global DNA re-methylation, the PGCs-to-spermatogonia/SSCs development entails further euchromatization. This accompanies substantial erasure of lamina-associated domains, generating spermatogonia/SSCs with a minimal peripheral attachment of chromatin except for pericentromeres-an architecture conserved in primates. Accordingly, faulty nucleome maturation, including persistent insulation and improper euchromatization, leads to impaired spermatogenic potential. Given that PGCs after epigenetic reprogramming serve as oogenic progenitors as well, our findings elucidate a principle for the nucleome programming that creates gametogenic progenitors in both sexes, defining a basis for nuclear totipotency.

Expression of matrix metalloproteinase-3 and -10 is up-regulated in the periodontal tissues of aged mice.

OBJECTIVE: The present study was designed to investigate the whole transcriptome of periodontal tissues of both young and aged mice to identify the characteristic up-regulation of protease genes with aging and to localize their translated protein products in the periodontal tissues. BACKGROUND: The metzincin protease superfamily is composed of matrix metalloproteinases (MMPs), a disintegrin and metalloproteinases, and a disintegrin and metalloproteinases with thrombospondin motifs. Up-regulation of these extracellular matrix-degrading proteases has been implicated in senescence of tissues and organs, including the skin. However, few studies have investigated the expression profiles of these proteases and potential involvement in aging of periodontal tissues. METHODS: Periodontal tissues with the surrounding mandibular bones were collected from 50- and 10-week-old mice. Total RNA was extracted from the periodontal tissue and analyzed by cap analysis of gene expression (CAGE) to identify differentially expressed genes encoding the metzincin proteases. Furthermore, quantitative real-time polymerase chain reaction (qRT-PCR) was performed to validate the CAGE results, and the phenotypic expression of proteases involved in aging was localized via immunohistochemical analysis. RESULTS: The CAGE results showed that the expression levels of MMP-3, -10, and -12 were up-regulated at 50 weeks. Subsequent qRT-PCR analysis showed that the gene expression levels of MMP-3 and -10 were significantly increased with age. MMP-10 immunoreactivity was localized exclusively in the cementum and alveolar bone adjacent to the periodontal ligament and was stronger and broader in aged mice than young mice. MMP-3 immunoreactivity was localized in the periodontal ligaments at both 10 and 50 weeks. CONCLUSION: In the present study, we demonstrated that the expression of MMP-3 and -10 increased with aging and identified their characteristic localizations in aged periodontal tissues.

DUX4 is a multifunctional factor priming human embryonic genome activation.

Double homeobox 4 (DUX4) is expressed at the early pre-implantation stage in human embryos. Here we show that induced human DUX4 expression substantially alters the chromatin accessibility of non-coding DNA and activates thousands of newly identified transcribed enhancer-like regions, preferentially located within ERVL-MaLR repeat elements. CRISPR activation of transcribed enhancers by C-terminal DUX4 motifs results in the increased expression of target embryonic genome activation (EGA) genes ZSCAN4 and KHDC1P1. We show that DUX4 is markedly enriched in human zygotes, followed by intense nuclear DUX4 localization preceding and coinciding with minor EGA. DUX4 knockdown in human zygotes led to changes in the EGA transcriptome but did not terminate the embryos. We also show that the DUX4 protein interacts with the Mediator complex via the C-terminal KIX binding motif. Our findings contribute to the understanding of DUX4 as a regulator of the non-coding genome.

Two ovarian candidate enhancers, identified by time series enhancer RNA analyses, harbor rare genetic variations identified in ovarian insufficiency.

The genetic regulation of ovarian development remains largely unclear. Indeed, in most cases of impaired ovarian development-such as 46,XX disorders of sex development (DSD) without SRY, and premature ovarian insufficiency (POI)-the genetic causes have not been identified, and the vast majority of disease-associated sequence variants could lie within non-coding regulatory sequences. In this study, we aimed to identify enhancers of five ovarian genes known to play key roles in early ovarian development, basing our analysis on the expression of enhancer derived transcripts (eRNAs), which are considered to characterize active enhancers. Temporal expression profile changes in mouse WT1-positive ovarian cells were obtained from cap analysis of gene expression at E13.5, E16.5 and P0. We compared the chronological expression profiles of ovarian-specific eRNA with expression profiles for each of the ovarian-specific genes, yielding two candidate sequences for enhancers of Wnt4 and Rspo1. Both sequences are conserved between mouse and human, and we confirmed their enhancer activities using transient expression assays in murine granulosa cells. Furthermore, by sequencing the region in patients with impaired ovarian development in 24 patients, such as POI, gonadal dysgenesis and 46,XX DSD, we identified rare single nucleotide variants in both sequences. Our results demonstrate that combined analysis of the temporal expression profiles of eRNA and mRNA of target genes presents a powerful tool for locating cis-element enhancers, and a means of identifying disease-associated sequence variants that lie within non-coding regulatory sequences, thus advancing an important unmet need in forward human genetics.

PAX6-positive microglia evolve locally in hiPSC-derived ocular organoids.

Microglia are the resident immune cells of the central nervous system (CNS). They govern the immunogenicity of the retina, which is considered to be part of the CNS; however, it is not known how microglia develop in the eye. Here, we studied human-induced pluripotent stem cells (hiPSCs) that had been expanded into a self-formed ectodermal autonomous multi-zone (SEAM) of cells that partially mimics human eye development. Our results indicated that microglia-like cells, which have characteristics of yolk-sac-like linage cells, naturally develop in 2D eye-like SEAM organoids, which lack any vascular components. These cells are unique in that they are paired box protein 6 (PAX6)-positive, yet they possess some characteristics of mesoderm. Collectively, the data support the notion of the existence of an isolated, locally developing immune system in the eye, which is independent of the body's vasculature and general immune system.

Cap analysis of gene expression (CAGE) and noncoding regulatory elements.

Cap analysis of gene expression (CAGE) was developed to detect the 5' end of RNA. Trapping of the RNA 5'-cap structure enables the enrichment and selective sequencing of complete transcripts. Upscaled high-throughput versions of CAGE have enabled the genome-wide identification of transcription start sites, including transcriptionally active promoters and enhancers. CAGE sequencing can be exploited to draw comprehensive maps of active genomic regulatory elements in a cell type- and activation-specific manner. The cells of the immune system are among the best candidates to be analyzed in humans, since they are easily accessible. In this review, we discuss how CAGE data are instrumental for integrative analyses with quantitative trait loci and omics data, and their usefulness in the mechanistic interpretation of the effects of genetic variations over the entire human genome. Integrating CAGE data with the currently available omics information will contribute to better understanding of the genome-wide association study variants that lie outside of annotated genes, deepening our knowledge on human diseases, and enabling the targeted design of more specific therapeutic interventions.

CD153/CD30 signaling promotes age-dependent tertiary lymphoid tissue expansion and kidney injury.

Tertiary lymphoid tissues (TLTs) facilitate local T and B cell interactions in chronically inflamed organs. However, the cells and molecular pathways that govern TLT formation are poorly defined. Here, we identified TNF superfamily CD153/CD30 signaling between 2 unique age-dependent lymphocyte subpopulations, CD153+PD-1+CD4+ senescence-associated T (SAT) cells and CD30+T-bet+ age-associated B cells (ABCs), as a driver for TLT expansion. SAT cells, which produced ABC-inducing factors IL-21 and IFN-γ, and ABCs progressively accumulated within TLTs in aged kidneys after injury. Notably, in kidney injury models, CD153 or CD30 deficiency impaired functional SAT cell induction, which resulted in reduced ABC numbers and attenuated TLT formation with improved inflammation, fibrosis, and renal function. Attenuated TLT formation after transplantation of CD153-deficient bone marrow further supported the importance of CD153 in immune cells. Clonal analysis revealed that SAT cells and ABCs in the kidneys arose from both local differentiation and recruitment from the spleen. In the synovium of aged rheumatoid arthritis patients, T peripheral helper/T follicular helper cells and ABCs also expressed CD153 and CD30, respectively. Together, our data reveal a previously unappreciated function of CD153/CD30 signaling in TLT formation and propose targeting the CD153/CD30 signaling pathway as a therapeutic target for slowing kidney disease progression.

Chromosomal-scale de novo genome assemblies of Cynomolgus Macaque and Common Marmoset.

Cynomolgus macaque (Macaca fascicularis) and common marmoset (Callithrix jacchus) have been widely used in human biomedical research. Long-standing primate genome assemblies used the human genome as a reference for ordering and orienting the assembled fragments into chromosomes. Here we performed de novo genome assembly of these two species without any human genome-based bias observed in the genome assemblies released earlier. We assembled PacBio long reads, and the resultant contigs were scaffolded with Hi-C data, which were further refined based on Hi-C contact maps and alternate de novo assemblies. The assemblies achieved scaffold N50 lengths of 149 Mb and 137 Mb for cynomolgus macaque and common marmoset, respectively. The high fidelity of our assembly is also ascertained by BAC-end concordance in common marmoset. Our assembly of cynomolgus macaque outperformed all the available assemblies of this species in terms of contiguity. The chromosome-scale genome assemblies produced in this study are valuable resources for non-human primate models and provide an important baseline in human biomedical research.