Biology subjects

Zhang, S.

Publications and source records attributed to Zhang, S..

At least 793 records · Page 44Linked to original sources

bioRxiv2022

Inferring CTCF insulators and anchored loops across human tissues and cell types

CCCTC-binding factor (CTCF) is a transcription regulator which is involved in many cellular processes. How CTCF recognizes DNA sequence to exert chromosome barrier or enhancer blocking effects remains to be fully interrogated. Despite many computational tools were developed to predict CTCF-mediated loops qualitatively or quantitatively, few could specially evaluate the regulatory potential of DNA sequence at CTCF binding sites (CBSs) and how it affects chromatin loop formation. Here, we developed a deep learning model, DeepAnchor, to precisely characterize the binding patterns for different types of CBSs. By incorporating base-wise genomic/epigenomic features, we revealed distinct chromatin and sequence features for CTCF-mediated insulation and looping at a high resolution, such as two sequence motifs flanking the core CTCF motif at loop-associated CBSs. Besides, we leveraged the predicted anchor score to optimize the loop extrusion model and achieved the best performance in predicting CTCF-anchored loops. We established a compendium of context-specific CTCF-anchored loops across 52 human tissue/cell types and found that genomic disruption of CTCF-anchored loops may represent a general causal mechanism of disease pathogenesis. These computational models, together with the established resource, could facilitate the mechanistic research on how the CTCF-mediated cis-regulatory elements (CREs) shapes context-specific gene regulation in cell development and disease progression.

bioinformatics↗

bioRxiv2022

A GPU-based computational framework that bridges Neuron simulation and Artificial Intelligence

Biophysically detailed multi-compartment models are powerful tools to explore computational principles of the brain and also serve as a theoretical framework to generate algorithms for artificial intelligence (AI) systems. However, the expensive computational cost severely limits the applications in both the neuroscience and AI fields. The major bottleneck during simulating detailed compartment models is the ability of a simulator to solve large systems of linear equations. Here, we present a novel Dendritic Hierarchical Scheduling (DHS) method to markedly accelerate such process. We theoretically prove that the DHS implementation is computationally optimal and accurate. This GPU-based method performs at 2-3 orders of magnitude higher speed than that of the classic serial Hines method in the conventional CPU platform. We build a DeepDendrite framework, which integrates the DHS method and the GPU computing engine of the NEURON simulator and demonstrate applications of DeepDendrite in neuroscience and AI tasks. We investigated how spatial patterns of spine inputs affect neuronal excitability in a detailed human pyramidal neuron model with 25,000 spines; and examined how dendrites protect morphologically detailed neural networks against adversarial attacks in typical image classification tasks.

neuroscience↗

bioRxiv2022

Basis for antibody- and hormone-mediated activation of TSHR in Graves' disease

Thyroid stimulating hormone (TSH), through activation of its G protein-coupled receptor TSHR, controls the synthesis of thyroid hormone (TH), an essential metabolic hormone. Aberrant signaling of TSHR by autoantibodies causes Graves disease and hypothyroidism that affect millions of patients worldwide. Here we report the active structures of TSHR with TSH and an activating autoantibody M22, both bound to an allosteric agonist ML-109, as well as an inactive TSHR structure with inhibitory antibody K1-70. Both TSH and M22 push the extracellular domain (ECD) of TSHR into the upright active conformation. In contrast, K1-70 blocks TSH binding and is incapable of pushing the ECD to the upright conformation. Comparisons of the active and inactive structures of TSHR with those of the luteinizing hormone-choriogonadotropin receptor (LHCGR) reveal a universal activation mechanism of glycoprotein hormone receptors, in which a conserved 10-residue fragment (P10) from the hinge C-terminal loop mediated interactions from the receptor ECD to its transmembrane domain. One surprisingly feature is that there are over 15 cholesterols surrounding TSHR, supporting its preferential location in lipid rafts. These structures also highlight a common mechanism for TSH and autoantibody M22 to activate TSHR, thus providing the molecular basis for Graves disease.

molecular biology↗

bioRxiv2022

Single-Cell RNA-Seq Uncovers Dynamic Processes Orchestrated by RNA-Binding Protein DDX43 in Chromatin Remodeling during Spermiogenesis

Advances in single-cell RNA sequencing (scRNA-seq) have allowed for elucidating biological mechanisms at cell state level. Mammalian spermatogenic process showcases dynamic switches of gene expression pattern with delicate morphological and functional alterations of germ cells, but it is unclear how such dynamics is genetically controlled. Here we demonstrate that mouse testis-enriched RNA helicase DDX43, as well as its ATP hydrolysis site, is required for spermiogenesis. Genetic mutation of Ddx43 renders spermatids heterogeneously defective in multiple steps of chromatin remodeling, resulting in incomplete substitution of transition protein by protamine and less condensed sperm nucleus. Through scRNA-seq analyses of testicular cells derived from adult wild-type and Ddx43 mutant testes in mice, we reveal that the DDX43 deficiency-elicited perturbation in the dynamic RNA regulatory processes underlies the differentiation deficiency of spermatids. Further, focused analyses on early-stage spermatids combined with enhanced CLIP sequencing (eCLIP-seq) identify Elfn2 as DDX43-targeted hub gene, whose in vivo knockdown shows similar phenotypic defects as Ddx43 mutant. Our study illustrates an essential role for DDX43 in post-meiotic chromatin remodeling and highlights the single cell-based strategy for a refined dissection of stage-specific regulation of germline differentiation.

developmental biology↗

bioRxiv2022

The cell cycle inhibitor RB is diluted in G1 and contributes to controlling cell size in the mouse liver

Every type of cell in an animal maintains a specific size, which likely contributes to its ability to perform its physiological functions. While some cell size control mechanisms are beginning to be elucidated through studies of cultured cells, it is unclear if and how such mechanisms control cell size in an animal. For example, it was recently shown that RB, the retinoblastoma protein, was diluted by cell growth in G1 to promote size-dependence of the G1/S transition. However, it remains unclear to what extent the RB-dilution mechanism controls cell size in an animal. We therefore examined the contribution of RB-dilution to cell size control in the mouse liver. The RB-dilution model has two requirements. First, manipulations changing RB concentration drive corresponding changes in cell size, and second, the endogenous RB concentration decreases with cell size in G1. We found that both these requirements were met. Genetic perturbations decreasing RB protein concentrations through inducible shRNA expression or through liverspecific Rb1 knockout reduced hepatocyte size, while perturbations increasing RB protein concentrations in an Fah-/- mouse model increased hepatocyte size. Moreover, RB concentration decreased in larger G1 hepatocytes while the concentrations of the cell cycle activators Cyclin D1 and E2f1 remained relatively constant. Lastly, we tested how Rb1 manipulations affected G1/S cell size control in primary hepatocytes using live cell imaging. Loss of Rb1 weakened cell size control, i.e., reduced the inverse correlation between how much cells grew in G1 and how large they were at birth. Taken together, our results show that an RB- dilution mechanism contributes to cell size control in the mouse liver by linking cell growth to the G1/S transition.

cell biology↗

bioRxiv2022

TE invasion fuels molecular adaptation in laboratory populations of Drosophila melanogaster

Transposable elements are mobile genetic parasites that frequently invade new host genomes through horizontal transfer. Invading TEs often exhibit a burst of transposition, followed by reduced transposition rates as repression evolves in the host. We recreated the horizontal transfer of P-element DNA transposons into a D. melanogaster host, and followed the expansion of TE copies and evolution of host repression in replicate laboratory populations reared at different temperatures. We observed that while populations maintained at high temperatures rapidly go extinct after TE invasion, those maintained at lower temperatures persist, allowing for TE spread and the evolution of host repression. We also surprisingly discovered that invaded populations experienced recurrent insertion of P-elements into a specific long non-coding RNA, lncRNA:CR43651, and that these insertion alleles are segregating at unusually high frequency in experimental populations, indicative of positive selection. We propose that, in addition to driving the evolution of repression, transpositional bursts of invading TEs can drive molecular adaptation.

evolutionary biology↗

bioRxiv2022

Development and application of an uncapped mRNA platform

A novel uncapped mRNA platform was developed. Five lipid nanoparticle (LNP)-encapsulated mRNA constructs were made to evaluate several aspects of our platform, including transfection efficiency and durability in vitro and in vivo and the activation of humoral and cellular immunity in several animal models. The constructs were eGFP-mRNA-LNP (for enhanced green fluorescence mRNA), Fluc-mRNA-LNP (for firefly luciferase mRNA), S{delta}T-mRNA-LNP (for Delta strain SARS-CoV-2 spike protein trimer mRNA), gDED-mRNA-LNP (for truncated glycoprotein D mRNA coding ectodomain from herpes simplex virus type 2 (HSV2)) and gDFR-mRNA-LNP (for truncated HSV2 glycoprotein D mRNA coding amino acids 1[~]400). Quantifiable target protein expression was achieved in vitro and in vivo with eGFP-and Fluc-mRNA-LNP. S{delta}T-mRNA-LNP, gDED-mRNA-LNP and gDFR-mRNA-LNP induced both humoral and cellular immune responses comparable to those obtained by previously reported capped mRNA-LNP constructs. Notably, S{delta}T-mRNA-LNP elicited neutralizing antibodies in hamsters against the Omicron and Delta strains. Additionally, gDED-mRNA-LNP and gDFR-mRNA-LNP induced potent neutralizing antibodies in rabbits and mice. The mRNA constructs with uridine triphosphate (UTP) outperformed those with N1-methylpseudouridine triphosphate (N1m{psi}TP) in the induction of antibodies via S{delta}T-mRNA-LNP. Our uncapped, process-simplified, and economical mRNA platform may have broad utility in vaccines and protein replacement drugs.

bioengineering↗

bioRxiv2022

Uncovering the spatial landscape of molecular interactions within the tumor microenvironment through latent spaces

Recent advances in spatial transcriptomics (ST) enable gene expression measurements from a tissue sample while retaining its spatial context. This technology enables unprecedented in situ resolution of the regulatory pathways that underlie the heterogeneity in the tumor and its microenvironment (TME). The direct characterization of cellular co-localization with spatial technologies facilities quantification of the molecular changes resulting from direct cell-cell interaction, as occurs in tumor-immune interactions. We present SpaceMarkers, a novel bioinformatics algorithm to infer molecular changes from cell-cell interaction from latent space analysis of ST data. We apply this approach to infer molecular changes from tumor-immune interactions in Visium spatial transcriptomics data of metastasis, invasive and precursor lesions, and immunotherapy treatment. Further transfer learning in matched scRNA-seq data enabled further quantification of the specific cell types in which SpaceMarkers are enriched. Altogether, SpaceMarkers can identify the location and context-specific molecular interactions within the TME from ST data.

bioinformatics↗

bioRxiv2022

Mitochondrial function determines severity but not risk of amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease. Selective vulnerability of energy-intensive motor neurons (MNs) has fostered speculation that mitochondrial function is a determinant of ALS. Previously, the position of mitochondrial function in the pathogenic cascade leading to neurotoxicity has been unclear. We separated upstream genetic determinants of mitochondrial function, including genetic variation within the mitochondrial genome or autosomes; from downstream changeable factors including mitochondrial copy number (mtCN) and MN gene expression. We discovered that functionally validated mitochondrial haplotypes are a determinant of ALS survival but not ALS risk. Loss-of-function genetic variants within, and reduced MN expression of, ACADM and DNA2 lead to shorter ALS survival; both genes impact mitochondrial function. MtCN responds dynamically to the onset of ALS independent of mitochondrial haplotype, and is also significantly correlated with disease severity. We conclude that mitochondrial function impacts ALS progression but not risk; our findings have therapeutic implications.

genetics↗

bioRxiv2022

Aridity modulates biogeographic distribution and community assembly of cyanobacterial morphotypes in drylands

The patterns of biogeographical distribution and assembly processes of microbiota are of vital importance for understanding ecological adaptation and functioning maintenance that microorganisms provide. Although the assembly of microbial communities was increasingly inspected, the role of their morphological characteristics is still poorly ascertained. Here, by integrating high-throughput sequencing and traditional trait-based classification, we investigated taxonomic and phylogenetic turnovers of different morphotypes of terrestrial cyanobacteria in biocrusts, as a model system, to evaluate the contributions of deterministic and stochastic processes across a large scale in drylands. The results showed that the bundle-forming category dominated in arid ecosystems with high tolerance against environmental fluctuations. Despite strong distance-decay relationships of {beta}-diversity in all categories, both spatial composition and phylogenetic turnover rates of bundle-forming cyanobacteria were significantly lower than unicellular/colonial, heterocystous, and other non-heterocyst filamentous cyanobacteria. Null model analysis of phylogenetic signals and abundance-based neutral model found that stochastic processes prevailed in the assembly, while aridity mediated the balance between determinism and stochasticity and prompted a shifting threshold among morphotypes. Our findings provide a unique perspective to understand the role of microbial morphology, highlighting the differentiation of biogeographic distribution, environmental response, and species preference between morphotypes with consideration of potential functional consequences, and therefore facilitated the prediction of biodiversity loss under climate change.

ecology↗

bioRxiv2022

Deciphering Hierarchical Chromatin Structures and Preference of Genomic Positions in Single Mouse Embryonic Stem Cells

The exploration of single-cell 3D genome maps reveals that chromatin domains are indeed physical structures presenting in single cells and domain boundaries vary from cell to cell. However, exhaustive analysis of regulatory factor binding or elements for preference of the formation of chromatin domains in single cells has not yet emerged. To this end, we first develop a hierarchical chromatin domain structure identification algorithm (named as HiCS) from individual single-cell Hi-C maps, with superior performance in both accuracy and efficiency. The results suggest that in addition to the known CTCF-cohesin complex, Polycomb, TrxG, pluripotent protein families and other multiple factors also contribute to shaping chromatin domain boundaries in single embryonic stem cells. Different cooperation patterns of regulatory factors decipher the preference of genomic position categories forming boundaries. And the most extensive six types of retrotransposons differentially distributed in these genomic position categories with preferential localization.

bioinformatics↗

bioRxiv2022

The SARS-CoV-2 spike protein binds and modulates estrogen receptors

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) protein binds angiotensin-converting enzyme 2 (ACE2) at the cell surface, which constitutes the primary mechanism driving SARS-CoV-2 infection. Molecular interactions between the transduced S and endogenous proteins likely occur post-infection, but such interactions are not well understood. We used an unbiased primary screen to profile the binding of full-length S against >9,000 human proteins and found significant S-host protein interactions, including one between S and human estrogen receptor alpha (ER). After confirming this interaction in a secondary assay, we used bioinformatics, supercomputing, and experimental assays to identify a highly conserved and functional nuclear receptor coregulator (NRC) LXD-like motif on the S2 subunit and an S-ER binding mode. In cultured cells, S DNA transfection increased ER cytoplasmic accumulation, and S treatment induced ER-dependent biological effects and ACE2 expression. Noninvasive multimodal PET/CT imaging in SARS-CoV-2-infected hamsters using [18F]fluoroestradiol (FES) localized lung pathology with increased ER lung levels. Postmortem experiments in lung tissues from SARS-CoV-2-infected hamsters and humans confirmed an increase in cytoplasmic ER expression and its colocalization with S protein in alveolar macrophages. These findings describe the discovery and characterization of a novel S-ER interaction, imply a role for S as an NRC, and are poised to advance knowledge of SARS-CoV-2 biology, COVID-19 pathology, and mechanisms of sex differences in the pathology of infectious disease.

immunology↗

bioRxiv2022

Comparative genomic insights into the evolution of Halobacteria-associated "Candidatus Nanohaloarchaeota"

The phylum "Candidatus Nanohaloarchaeota" is a representative halophilic lineage within DPANN superphylum. They are characterized by their nanosized cells and symbiotic lifestyle with Halobacteria. However, the development of the symbiosis remains unclear for the lack of genomes located at the transition stage. Here, we performed a comparative genomic analysis of "Ca. Nanohaloarchaeota". We propose a novel family "Candidatus Nanoanaerosalinaceae" represented by five de-replicated metagenome-assembled genomes obtained from hypersaline sediments and the enrichment cultures of soda-saline lakes. Phylogeny analysis reveals that the novel family are placed at the root of the family "Candidatus Nanosalinaceae" including the well-researched taxa. Most members of "Ca. Nanoanaerosalinaceae" contain lower proportion of putative horizontal gene transfers from Halobacteria than "Ca. Nanosalinaceae", while they maintain moderately acidic proteomes for hypersaline adaptation of "salt-in" strategy, suggesting that "Ca. Nanoanaerosalinaceae" have not established an intimate association with Halobacteria, and may descend from an intermediate stage. Functional prediction discloses that they exhibit divergent potentials in carbohydrate and organic acids metabolism, and environmental responses. Historical events reconstruction illustrates that the involved genes acquired at the putative ancestors possibly drive the evolutionary and symbiotic divergences. Globally, this research on the new family "Ca. Nanoanaerosalinaceae" enriches the taxonomic and functional diversity of "Ca. Nanohaloarchaeota", and provides insights into the evolutionary process of "Ca. Nanohaloarchaeota" and their Halobacteria-associated symbiosis. ImportanceDPANN superphylum is a group of archaea widely distributing in various habitats. They generally have small cells, and perform a symbiotic lifestyle with other archaea. The archaeal symbiotic interaction is important to understand microbial community. However, the formation and evolution of the symbiosis between the DPANN lineages and other diverse archaea remain unclear. Based on phylogeny, hypersaline adaptation, functional potentials, and historical events of "Ca. Nanohaloarchaeota", a representative phylum within the DPANN superphylum, we report a novel family descending from an intermediate stage, and we illustrate the evolutionary process of "Ca. Nanohaloarchaeota" and their Halobacteria-associated symbiosis. Furthermore, we find the acquired genes involved in carbohydrate and organic acids metabolism and environmental responses possibly drive the evolutionary and symbiotic divergences. Altogether, this research helps in understanding the evolution of the archaeal symbiosis, and provides a model for the evolution of the other DPANN lineages.

microbiology↗

bioRxiv2022

Retina Gap Junction Networks Facilitate Blind Denoising in Visual Hierarchy

Gap junctions in the retina are electrical synapses, which strength is regulated byambient light conditions. Such tunable synapses are crucial for the denoising function of the early visual system. However, it is unclear that how the plastic gap junction network processes unknown noise, specifically how this process works synergistically with the brains higher visual centers. Inspired by the electrically coupled photoreceptors, we develop a computational model of the gap junction filter (G-filter). We show that G-filter is an effective blind denoiser that converts different noise distributions into a similar form. Next, since deep convolutional neural networks (DCNNs) functionally reflect some intrinsic features of the visual cortex, we combine G-filter with DCNNs as retina and ventral visual pathways to investigate the relationship between retinal denoising processing and the brains high-level functions. In the image denoising and reconstruction task, G-filter dramatically improve the classic deep denoising convolutional neural network (DnCNN)s ability to process blind noise. Further, we find that the gap junction strength of the G-filter modulates the receptive field of DnCNNs output neurons by the Integrated Gradients method. At last, in the image classification task, G-filter strengthens the defense of state-of-the-arts DCNNs (ResNet50, VGG19 and InceptionV3) against blind noise attacks, far exceeding human performance when noise is large. Our results indicate G-filter significantly enhance DCNNs ability on various blind denoising tasks, implying an essential role for retina gap junction networks in high-level visual processing.

neuroscience↗

bioRxiv2022

RNA Methylome Reveals the m6A-Mediated Regulation of Flavor Metabolites in Tea Leaves under Solar-Withering

Epitranscriptomic mark N6-methyladenosine (m6A) is the most predominant internal modification in RNAs, which plays pivotal roles in response to diverse stresses. Multiple environmental stresses caused by withering process can greatly influence the accumulation of specialized metabolites and the formation of tea flavor. However, little is known about the effects of m6A-mediated regulatory mechanism on flavor-related metabolisms in tea leaves. Here, we explored m6A-mediated regulatory mechanism and its impacts on flavonoid and terpenoid metabolisms under solar-withering using integrated RNA methylome and transcriptome. Dynamic changes in global m6A levels of tea leaves are mainly controlled by two m6A erasers (CsALKBH4A and CsALKBH4B) under solar-withering. Differentially methylated peak (DMP)-associated genes under different shading rates of solar-withering were identified and found to be enriched in terpenoid biosynthesis and spliceosome pathways. Further analyses indicated that CsALKBH4-driven RNA demethylation can not only directly affect the accumulation of volatile terpenoids by mediating the stability and abundance of terpenoid biosynthesis-related genes, but also indirectly influence the contents of flavonoids, catechins, and theaflavins via triggering the alternative splicing (AS)-mediated regulation. Our findings underscored a novel layer of epitranscriptomic gene regulation in tea flavor-related metabolic pathways and established a compelling link between m6A-mediated regulatory mechanism and the formation of high-quality flavor in tea leaves under solar-withering.

plant biology↗

bioRxiv2022

Specific binding of Hsp27 and phosphorylated Tau mitigates abnormal Tau aggregation-induced pathology

Amyloid aggregation of phosphorylated Tau (pTau) into neurofibrillary tangles is closely associated with Alzheimers disease (AD). Several molecular chaperones have been reported to bind Tau and impede its pathological aggregation. Recent findings of elevated levels of Hsp27 in the brains of patients with AD suggested its important role in pTau pathology. However, the molecular mechanism of Hsp27 in pTau aggregation remains poorly understood. Here, we show that Hsp27 partially co-localizes with pTau tangles in the brains of patients with AD. Notably, phosphorylation of Tau by microtubule affinity regulating kinase 2 (MARK2), dramatically enhances the binding affinity of Hsp27 to Tau. Moreover, Hsp27 efficiently prevents pTau fibrillation in vitro and mitigates neuropathology of pTau aggregation in a Drosophila tauopathy model. Further mechanistic study reveals that Hsp27 employs its N-terminal domain to directly interact with multiple phosphorylation sites of pTau for specific binding. Our work provides the structural basis for the specific recognition of Hsp27 to pathogenic pTau, and highlights the important role of Hsp27 in preventing abnormal aggregation and pathology of pTau in AD.

biochemistry↗

bioRxiv2022

The peptide GOLVEN10 controls nodule and lateral root organogenesis and positioning along the longitudinal root axis

O_LIGLV/RGF peptide encoding genes can be identified in genomes of all plants that can form roots or root-like structures suggesting they were essential for transition of plants to land. C_LIO_LIIn Medicago truncatula, five of fifteen GOLVEN(GLV)/ROOT MERISTEM GROWTH FACTOR (RGF) peptide coding genes were induced during nodule organogenesis and to a varying extent under nitrogen deficiency and auxin treatment. Expression of MtGLV9 and MtGLV10 at nodule initiation sites was dependent on the transcription factor NODULE INCEPTION. C_LIO_LIOverexpression of all five nodule-induced GLV genes in M. truncatula hairy roots as well as application of the corresponding synthetic peptides resulted in a 25-50% reduction in nodule number indicating GOLVENs are negative regulators of nodule organogenesis. C_LIO_LIThe peptide GOLVEN10 shifted the position of the first formed lateral root (rhizotaxis) as well as the first formed nodule along the longitudinal primary root axis, a phenomenon we term nodulotaxis, thereby reducing the absolute length of the zone of lateral organ formation on roots. C_LIO_LIApplication of synthetic GOLVEN10 peptide caused an increase in cell number but not cell length in each root cortical cell layer causing an increase in root length and a consequent spatiotemporal delay in formation of the first lateral organ. C_LI Plain Language SummaryNodule positioning is an understudied trait, yet it determines the length of the root that can support nodule formation and consequently the total number of functional nodules formed. We identify for the first time, genetic factors called GOLVEN peptides that alter nodule and lateral root positioning on the primary root along with several other traits including nodule organ initiation and root architecture.

plant biology↗

bioRxiv2022

Deeply conserved super-enhancers maintain stem cell pluripotency in placental mammals

Despite pluripotent stem cells sharing key transcription factors, their maintenance involves distinct genetic inputs. Emerging evidence suggests that super-enhancers (SEs) can function as master regulatory hubs to control cell identity and pluripotency in humans and mice. However, whether pluripotency-associated SEs share a deep evolutionary origin in mammals remains elusive. Here, we performed comprehensive comparative epigenomic and transcription factor binding analyses among pigs, humans, and mice to identify pluripotency-associated SEs. Like typical enhancers, SEs displayed rapid evolution in mammals. We showed that BRD4 is an essential and conserved activator for mammalian pluripotency-associated SEs. Comparative motif enrichment analysis revealed 30 shared transcription factor binding motifs among the three species. The majority of the transcriptional factors that bind to identified motifs are known regulators associated with pluripotency. Further, we discovered three pluripotency-associated SEs (SE-SOX2, SE-PIM1, and SE-FGFR1) which displayed deep conservation in placental mammals and are sufficient to drive reporter gene expression in a pluripotency-dependent manner. Disruption of these conserved SEs through the CRISPR/Cas9 approach severely impaired the proliferative potential and the ability to form undifferentiated colonies. Our study provides insights into the understanding of conserved regulatory mechanisms underlying the maintenance of pluripotency as well as species-specific modulation of the pluripotency-associated regulatory networks in mammals. Significance statementSuper-enhancers (SEs) hold stronger power than regular enhancers to direct gene expression in the regulation of stem cell pluripotency. To dissect how pluripotency-associated SEs have evolved in mammals, we performed a systematic comparison of SEs among pigs, humans, and mice. Our analysis allowed the identification of three pluripotency-associated SEs (SE-SOX2, SE-PIM1, and SE-FGFR1) that are highly conserved in Placentalia (accounting for 94% of mammals) as well as many species-specific SEs. All three SEs were sufficient to direct pluripotency-dependent gene expression and disruption of each conserved SE caused the loss of stem cell pluripotency. Our work highlights a small number of highly conserved SEs essential for the maintenance of pluripotency.

cell biology↗