Biology subjects

Deng, B.

Publications and source records attributed to Deng, B..

34 records · Page 2Linked to original sources

bioRxiv2023

Genome-wide identification of silencers in human cells

Transcriptional regulation is a complex process that is controlled by a variety of factors, including enhancers and silencers. Silencers, also known as repressor elements, play a crucial role in the fine-tuning of gene expression by inhibiting or suppressing transcription in the human genome. Although significant progresses have been made, genome-wide silencer research is still in its early stages. Here, we used a genome-wide method called massively parallel reporter assays (MPRAs) to identify silencers in three human cell lines: K562, LNCap, and HEK293T. We identified 739,434, 643,484, and 491,952 silencer regions in these cell lines, respectively. We found that most of the silencers we identified had inhibitory activity and significantly enriched inhibitory motifs. These results confirm that silencers are ubiquitous in the human genome and play an important role in regulating gene expression. Therefore, our study provides a general strategy for genome-wide functional identification of silencer elements. This information could be used to better understand the mechanisms of gene regulation and to develop new therapeutic strategies for diseases that are caused by dysregulation of gene expression.

genomics↗

bioRxiv2023

Histone variant H2B.Z acetylation is necessary for maintenance of Toxoplasma gondii biological fitness

Through regulation of DNA packaging, histone proteins are fundamental to a wide array of biological processes. A variety of post-translational modifications (PTMs), including acetylation, constitute a proposed histone code that is interpreted by "reader" proteins to modulate chromatin structure. Canonical histones can be replaced with variant versions that add an additional layer of regulatory complexity. The protozoan parasite Toxoplasma gondii is unique among eukaryotes in possessing a novel variant of H2B designated H2B.Z. The combination of PTMs and the use of histone variants is important for gene regulation in T. gondii, offering new targets for drug development. In this work, T. gondii parasites were generated in which the 5 N-terminal acetylatable lysines in H2B.Z were mutated to either alanine (c-Myc-A) or arginine (c-Myc-R). c-Myc-A mutant only displayed a mild effect in its ability to kill mice. c-Myc-R mutant presented an impaired ability to grow and an increase in differentiation to latent bradyzoites. This mutant line was also more sensitive to DNA damage, displayed no virulence in mice, and provided protective immunity against future infection. While nucleosome composition was unaltered, key genes were abnormally expressed during in vitro bradyzoite differentiation. Our results show that the N-terminal positive charge patch of H2B.Z is important for these procceses. Pull down assays with acetylated N-terminal H2B.Z peptide and unacetylated one retrieved common and differential interactors. Acetylated peptide pulled down proteins associated with chromosome maintenance/segregation and cell cycle, opening the question of a possible link between H2B.Z acetylation status and mitosis.

cell biology↗

bioRxiv2022

Modulating the epigenetic state promotes the reprogramming of transformed cells to pluripotency in a line-specific manner

Somatic cell reprogramming and oncogenic transformation share surprisingly similar features, yet transformed cells are highly resistant to reprogramming. There must be barriers that block transformed cells from reprogramming, but the nature of those barriers is unclear. In this study, we generated a systematic panel of transformed mouse embryonic fibroblasts (MEFs) using a variety of oncogenic transgenes, and discovered transformed cell lines that remain compatible with reprogramming when transfected with Oct4/Sox2/Klf4/Myc. By comparing the reprogramming-capable and incapable transformed lines we identified multiple stages of failure in the reprogramming process. Some transformed lines failed very early, whilst other lines seemed to progress through a normal-looking reprogramming process. Finally, we show that MEK inhibition overcomes one critical reprogramming barrier by indirectly suppressing a hyperactive epigenetic state in some of the transformed cells. This study reveals that the barriers underlying resistance to reprogramming vary between the different transformation methods. Key findingsO_LISomatic cell reprogramming of transformed cells is context-specific C_LIO_LIInhibition of MEK converts some cell lines to reprogramming-capable C_LIO_LITransformed cell lines are characterized by a hyperactive chromatin state C_LIO_LIMEK inhibition indirectly affects chromatin to enable reprogramming C_LI

cell biology↗

bioRxiv2022

Triangulation reduces the polygon of error for the history of Transeurasian

In a recent study we used an interdisciplinary approach combining linguistics, archaeology and genetics to analyse the Transeurasian languages1. Our analysis concluded that the early dispersals of these languages were driven by agriculture. A preprint published on this server presents objections to the Transeurasian hypothesis and its association with farming dispersals2. However, close inspection of that text reveals numerous misinterpretations and inconsistencies. In the interest of furthering scientific debate over Transeurasian language and population history, we address the critiques, revising datasets and fine-tuning approaches. The linguistic critique questions the quantity and quality of our datasets. Here we show that the number of surviving cognate sets for Transeurasian is in line with that for well-established language families. In addition, we find that Tian et al.s failure to reject a core of regularly corresponding cognates in the basic vocabulary creates ground for a consensus about the genealogical relatability of the Transeurasian languages. The archaeological critique attempts a re-analysis of one Bayesian test using re-scored data only for northern China. Over half of the suggested re-scorings contain inconsistencies and it is not explained why the re-analysis retains the original data for sites outside northern China, comprising almost 60% of the total. More importantly, the sweeping claim that there is no evidence supporting the prehistoric migrations analysed in our study is not backed by any discussion of the archaeological record. With respect to genetics, the preprint claims a re-analysis showing that the data do not conclusively support the farming-driven dispersal of Turkic, Mongolian, and Tungusic, nor the two-wave spread of farming to Korea. In fact, the only genetic re-analysis presented is limited to samples from Korea and Japan and does not contradict our original conservative modelling of Neolithic individuals with Hongshan and our Bronze Age ones with Upper Xiajiadian. In sum, in bringing multiple lines of evidence together through triangulation, we gained a more balanced and richer understanding of Transeurasian dispersals than each discipline could provide individually. Our research doubtless leaves room for improvement but we remain confident that triangulation did not fail, but rather brought us a step closer to understanding the history of the Transeurasian languages.

evolutionary biology↗

bioRxiv2022

Structural mechanism of leaflet-specific phospholipid modulation of a pentameric ligand-gated ion channel

Pentameric ligand-gated ion channels (pLGICs) mediate synaptic transmission and are sensitive to their lipid environment. The mechanism of phospholipid modulation of any pLGIC is not well understood. We demonstrate that the model pLGIC, ELIC (Erwinia ligand-gated ion channel), is positively modulated by the anionic phospholipid, phosphatidylglycerol, from the outer leaflet of the membrane. To elucidate the mechanism of phosphatidylglycerol modulation, we determine a structure of ELIC in an open conformation. The structure shows a bound phospholipid in an outer leaflet site, and conformational changes in the phospholipid binding site unique to the open state. In combination with streamlined alchemical free energy perturbation calculations and functional measurements in asymmetric liposomes, the data support a mechanism by which an anionic phospholipid stabilizes the open state of a pLGIC by specific, state-dependent binding to this site.

biophysics↗

bioRxiv2022

TME induced TRAIL secretion from engineered macrophages for anti-tumor therapy

Capacity of tumor niche chemotaxis and long-term persistency making macrophage as great vehicles for anti-tumor factor delivery. Macrophages-based delivery of chemokines or cytokines have been tested for tissue homeostasis and cancer repression. TRAIL as a promising anti-tumor cytokine, made little clinical progress due to limited stability and off-target toxicity. Here we engineered macrophages with tumor micro-environment (TME)-induced trimerized CP1-TRAIL secretion under the TME specific promoter Arg1 (Tri-TRAIL-iM). The Tri-TRAIL-iM cells displayed high specific inducible activity in both cell-based co-culture assay and in tumor baring mice models. Compared to normal TRAIL over-expressed macrophages under none-inducible promoter, Tri-TRAIL-iM infiltrated to tumor sites and showed superior apoptosis induction of cancer cells and tumor growth repression as well as less systemic side effect. This inducible delivery TRAIL strategy can be effectively further applied in clinical studies and can be coupled with other engineered methods to maximize the therapeutic outcomes for solid tumors.

cancer biology↗

bioRxiv2021

Drosulfakinin signaling modulates female sexual receptivity in Drosophila

Female sexual behavior as an innate behavior is of prominent biological importance for survival and reproduction. However, molecular and circuit mechanisms underlying female sexual behavior is not well understood. Here, we identify the Cholecystokinin-like peptide Drosulfakinin (DSK) promotes female sexual behavior in Drosophila. Manipulation both Dsk and DSK neuronal activity impact female sexual receptivity. In addition, we reveal that Dsk-expressing neurons receive input signal from R71G01GAL4 neurons to promote female sexual receptivity. Based on intersectional technique, we further found the regulation of female sexual behavior relies mainly on medial DSK neurons rather than lateral DSK neurons, and medial DSK neurons modulate female sexual behavior by acting on its receptor CCKLR-17D3. Thus, we characterized DSK/CCKLR-17D3 as R71G01GAL4 neurons downstream signaling to regulate female sexual behavior.

neuroscience↗

bioRxiv2021

Bimodal Remapping of Visual Grids

Spatially modulated neurons from the rat secondary visual cortex (V2) show grid-like firing patterns during freely foraging in open-field enclosures. However, the remapping of the V2 grid cells is not well understood. Here we report two classes of V2 grid cell populations with distinct remapping properties: one regular class with invariant grid field patterns, and the other bimodal class that has remapping induced by environmental manipulations such as changes in enclosure shape, size, orientation and lighting in a familiar environment. The bimodal V2 grid cell pattern remains stable regardless of the follow-up manipulations, but restores to the original firing pattern upon animals re-entry into the familiar environment on the next day or from the novel environment. The bimodal V2 grid cells are modulated with theta frequency during the course of remapping and stabilize quickly. We also found conjunctive bistable V2 grid cells with invariant head directional tuning. Overall, our results suggest a new grid cell mechanism in V2 that is different from the medial entorhinal cortex (MEC) grid cells. HighlightsO_LIBistable V2 grid cells display bimodal or bistable remapping C_LIO_LIV2 grid cell firing patterns are not disrupted in darkness C_LIO_LIV2 grid cells preserve theta frequency modulation during remapping C_LI

neuroscience↗

bioRxiv2021

Egocentric Asymmetric Coding in Sensory Cortical Border Cells

Both egocentric and allocentric representations of space are essential to spatial navigation. Although some studies of egocentric coding have been conducted within and around the hippocampal formation, externally anchored egocentric spatial representations have not yet been fully explored. Here we record and identify two subtypes of border cell in the rat primary somatosensory cortex (S1) and secondary visual cortex (V2). Subpopulations of S1 and V2 border cells exhibit rotation-selective asymmetric firing fields in an either clockwise (CW) or counterclockwise (CCW) manner. CW- and CCW-border cells increase their firing rates when animals move unidirectionally along environmental border(s). We demonstrate that both CW- and CCW-border cells fire in an egocentric reference frame relative to environmental borders, maintain preferred directional tunings in rotated, stretched, dark as well as novel arenas, and switch their directional firings in the presence of multi-layer concentric enclosures. These findings may provide rotation-selective egocentric reference frames within a larger spatial navigation system, and point to a common computational principle of spatial coding shared by multiple sensory cortical areas. HighlightsO_LIEgocentric border cells are present in rat S1 and V2 C_LIO_LISubtypes of border cells display egocentric asymmetric coding C_LIO_LIEgocentric and allocentric streams coexist in sensory cortices C_LIO_LIRotation-selective asymmetric firing is robust with environmental manipulations C_LI

neuroscience↗

bioRxiv2021

A compact spatial map in V2 visual cortex

Vision plays a critical role in guiding spatial navigation. A traditional view of the visual cortex is to compute a world-centered map of visual space, and visual neurons exhibit diverse tunings to simple or complex visual features. The neural representation of spatio-visual map in the visual cortex is thought to be transformed from spatial modulation signals at the hippocampal-entorhinal system. Although visual thalamic and cortical neurons have been shown to be modulated by spatial signals during navigation, the exact source of spatially modulated neurons within the visual circuit has never been identified, and the neural correlate underpinning a visuospatial or spatio-visual map remains elusive. To search for direct visuospatial and visuodirectional signals, here we record in vivo extracellular spiking activity in the secondary visual cortex (V2) from freely foraging rats in a naturalistic environment. We identify that V2 neurons forms a complete spatio-visual map with a wide range of spatial tunings, which resembles the classical spatial map that includes the place, head-direction, border, grid and conjunctive cells reported in the hippocampal-entorhinal network. These spatially tuned V2 neurons display stable responses to external visual cues, and are robust with respect to non- spatial environmental changes. Spatially and directionally tuned V2 neuronal firing persists in darkness, suggesting that this spatio-visual map is not completely dependent on visual inputs. Identification of functionally distinct spatial cell types in visual cortex expands its classical role of information coding beyond a retinotopic map of the eye-centered world.

neuroscience↗

bioRxiv2020

Blocking palmitoylation of Toxoplasma gondii myosin light chain 1 disrupts glideosome composition but has little impact on parasite motility

Toxoplasma gondii is a widespread apicomplexan parasite that causes severe disease in immunocompromised individuals and the developing fetus. Like other apicomplexans, T. gondii uses an unusual form of gliding motility to invade cells of its hosts and to disseminate throughout the body during infection. It is well established that a myosin-based motor consisting of a Class XIVa heavy chain (TgMyoA) and two light chains (TgMLC1 and TgELC1/2) plays an important role in parasite motility. The ability of the motor to generate force at the parasite periphery is thought to be reliant upon its anchoring and immobilization within a peripheral membrane-bound compartment, the inner membrane complex (IMC). The motor does not insert into the IMC directly; rather, this interaction is believed to be mediated by the binding of TgMLC1 to the IMC-anchored protein, TgGAP45. The binding of TgMLC1 to TgGAP45 is therefore considered a key element in the force transduction machinery of the parasite. TgMLC1 is palmitoylated, and we show here that palmitoylation occurs on two N-terminal cysteine residues, C8 and C11. Mutations that block TgMLC1 palmitoylation disrupt the association of TgMLC1 with the membrane fraction of the parasite in phase partitioning experiments and completely block the binding of TgMLC1 to TgGAP45. Surprisingly, the loss of TgMLC1 binding to TgGAP45 in these mutant parasites has little effect on their ability to initiate or sustain movement. These results question a key tenet of the current model of apicomplexan motility and suggest that our understanding of gliding motility in this important group of human and animal pathogens is not yet complete. ImportanceGliding motility plays a central role in the life cycle of T. gondii and other apicomplexan parasites. The myosin motor thought to power motility is essential for virulence but distinctly different from the myosins found in humans. Consequently, an understanding of the mechanism(s) underlying parasite motility and the role played by this unusual myosin may reveal points of vulnerability that can be targeted for disease prevention and treatment. We show here that mutations that uncouple the motor from what is thought to be a key structural component of the motility machinery have little impact on parasite motility. This finding runs counter to predictions of the current, widely-held "linear motor" model of motility, highlighting the need for further studies to fully understand how apicomplexan parasites generate the forces necessary to move into, out of and between cells of the hosts they infect.

microbiology↗

bioRxiv2020

Transposable Element-Gene Splicing Modulates the Transcriptional Landscape of Human Pluripotent Stem Cells

Transposable elements (TEs) occupy nearly 50% of mammalian genomes and are both potential dangers to genome stability and functional genetic elements. TEs can be expressed and exonised as part of a transcript, however, their full contribution to the transcript splicing remains unresolved. Here, guided by long and short read sequencing of RNAs, we show that 26% of coding and 65% of non-coding transcripts of human pluripotent stem cells (hPSCs) contain TEs. Different TE families have unique integration patterns with diverse consequences on RNA expression and function. We identify hPSC-specific splicing of endogenous retroviruses (ERVs) as well as LINE L1 elements into protein coding genes that generate TE-derived peptides. Finally, single cell RNA-seq reveals that proliferating hPSCs are dominated by ERV-containing transcripts, and subpopulations express SINE or LINE-containing transcripts. Overall, we demonstrate that TE splicing modulates the pluripotency transcriptome by enhancing and impairing transcript expression and generating novel transcripts and peptides.

genomics↗

bioRxiv2019

The mechanosensitive Piezo1 orchestrating angiogenesis is essential in bone fracture repair

Mechanical ion channel protein Piezo1 play vital roles in angiogenesis which has been proved to be high importance in varieties of biological processes. Bone formation in the fracture repair requires oxygen and nutrients from new blood vessels generated from fractured lesion. Understanding the underlying mechanisms linking angiogenesis and bone formation must be of great value for improved fracture healing. Here we employed mice with genetically modified endothelial specific depletion of Piezo1 channels to explore the hypothesis that Piezo1 is vital to the initiation of fracture healing. In this study, we demonstrated that Piezo1 expression and wide distribution along the bone and impaired endothelial Piezo1 channels result in derangements in bone fracture repair. Intriguingly, the calcium activated proteolytic caplain activity severely disrupted during vascularization, precluded osteoblast maturation and mineralization and subsequently the phosphorylated PI3K-AKT reduction. Furthermore, Piezo1 endothelial disruption impaired Notch signaling in bone union. These data collectively suggest that Piezo1 channels serve as a basis for clinical strategies to improve bone regeneration and treat delayed or nonunion in bone fracture.

systems biology↗

bioRxiv2019

Replication gaps underlie BRCA-deficiency and therapy response

Cancers that are deficient in BRCA1 or BRCA2 are hypersensitive to genotoxic agents, including platinums and other first-line chemotherapeutics. The established models propose that these cancers are hypersensitive because the chemotherapies block or degrade DNA replication forks and thereby create DNA double strand breaks, both of which require functional BRCA proteins to prevent or resolve by mechanisms termed fork protection (FP) or homologous recombination (HR). However, recent findings challenge this dogma because genotoxic agents do not initially cause DNA double strand breaks or stall replication forks. Here, we propose a new model for genotoxic chemotherapy in which ssDNA replication gaps underlie the hypersensitivity of BRCA deficient cancer, and we propose that defects in HR or FP do not. Specifically, we observed that ssDNA gaps develop in BRCA deficient cells because DNA replication is not effectively restrained in response to genotoxic stress. Moreover, we observe gap suppression (GS) by either restored fork restraint or by gap filling, both of which conferred resistance to therapy in tissue culture and BRCA patient tumors. In contrast, restored HR and FP were not sufficient to prevent hypersensitivity if ssDNA gaps were not eliminated. Together, these data suggest that ssDNA replication gaps underlie the BRCA cancer phenotype, \"BRCAness,\" and we propose are fundamental to the mechanism of action of genotoxic chemotherapies.

cancer biology↗

bioRxiv2019

Toxoplasma F-Box Protein 1 is Required for Daughter Cell Scaffold Function During Parasite Replication

By binding to the adaptor protein SKP1 and serving as substrate receptors for the Skp1, Cullin, F-box E3 ubiquitin ligase complex, F-box proteins regulate critical cellular processes including cell cycle progression and membrane trafficking. While F-box proteins are conserved throughout eukaryotes and are well studied in yeast, plants, and animals, studies in parasitic protozoa are lagging. We have identified eighteen putative F-box proteins in the Toxoplasma genome of which four have predicted homologs in Plasmodium. Two of the conserved F-box proteins were demonstrated to be important for Toxoplasma fitness and here we focus on an F-box protein, named TgFBXO1, because it is the most highly expressed by replicative tachyzoites and was also identified in an interactome screen as a Toxoplasma SKP1 binding protein. TgFBXO1 interacts with Toxoplasma SKP1 confirming it as a bona fide F-box protein. In interphase parasites, TgFBXO1 is a component of the Inner Membrane Complex (IMC), which is an organelle that underlies the plasma membrane. Early during replication, TgFBXO1 localizes to the developing daughter cell scaffold, which is the site where the daughter cell IMC and microtubules form and extend from. TgFBXO1 localization to the daughter cell scaffold required centrosome duplication but occurred before segregation of the centrocone, which connects the spindle pole to the centrosomes. Daughter cell scaffold localization required TgFBXO1 N-myristoylation and was dependent on the small molecular weight GTPase, TgRab11b. Finally, we demonstrate that TgFBXO1 is required for parasite growth due to its function as a daughter cell scaffold effector. TgFBXO1 is the first F-box protein to be studied in apicomplexan parasites and represents the first protein demonstrated to be important for daughter cell scaffold function.\n\nAUTHOR SUMMARYToxoplasma gondii is a protozoan parasite that can cause devastating and life-threatening disease in immunocompromised patients and in fetuses. Its replication is important to study because parasite growth is responsible for the pathology that develops in toxoplasmosis patients. The parasite replicates by a unique process named endodyogeny in which two daughter parasites develop within the mother cell. Early during this process the parasite creates a structure called the Daughter Cell Scaffold whose function is to mark the site from which daughter parasites will emerge. Here, we report the identification of one of the first proteins recruited to the Daughter Cell Scaffold and its importance in executing its function.

microbiology↗

bioRxiv2018

Enabling tools for Toxoplasma glycobiology

Infection by the protozoan parasite Toxoplasma gondii is a major health risk owing to its chronic nature, ability to reactivate to cause blindness and encephalitis, and high prevalence in human populations. Like nearly all eukaryotes, Toxoplasma glycosylates many of its proteins and lipids and assembles polysaccharides. Unlike most eukaryotes, Toxoplasma divides and differentiates in vacuoles within host cells. While their glycans resemble canonical models, they exhibit species-specific variations that have inhibited deeper investigations into their roles in parasite biology and virulence. The genome of Toxoplasma encodes a suite of likely glycogenes expected to assemble a range of N-glycans, O-glycans, a C-glycan, GPI-anchors, and polysaccharides, along with their requisite precursors and membrane transporters. To facilitate genetic approaches to investigate the roles of specific glycans, we mapped probable connections between 59 glycogenes, their enzyme products, and the glycans to which they contribute. We adapted a double-CRISPR/Cas9 strategy and a mass spectrometry-based glycomics workflow to test these relationships, and conducted infection studies in fibroblast monolayers to probe cellular functions. Through the validated disruption of 17 glycogenes, we also discovered novel Glc0-2-Man6-GlcNAc2-type N-glycans, GalNAc2- and Glc-Fuc-type O-glycans, and a nuclear O-Fuc type glycan. We describe the guide sequences, disruption constructs, and mutant strains, which are freely available to practitioners for application in the context of the relational map to investigate the roles of glycans in their favorite biological processes.

microbiology↗