Biology subjects

Shen, H.

Publications and source records attributed to Shen, H..

At least 127 records · Page 7Linked to original sources

bioRxiv2020

SARS-CoV-2 Spike Protein Impairs Endothelial Function via Downregulation of ACE2

Coronavirus disease 2019 (COVID-19) includes the cardiovascular complications in addition to respiratory disease. SARS-CoV-2 infection impairs endothelial function and induces vascular inflammation, leading to endotheliitis. SARS-CoV-2 infection relies on the binding of Spike glycoprotein (S protein) to angiotensin converting enzyme 2 (ACE2) in the host cells. We show here that S protein alone can damage vascular endothelial cells (ECs) in vitro and in vivo, manifested by impaired mitochondrial function, decreased ACE2 expression and eNOS activity, and increased glycolysis. The underlying mechanism involves S protein downregulation of AMPK and upregulation of MDM2, causing ACE2 destabilization. Thus, the S protein-exerted vascular endothelial damage via ACE2 downregulation overrides the decreased virus infectivity.

pathology↗

bioRxiv2020

An improved comprehensive strategy for deep and quantitative N-glycomics based on optimization of sample preparation, isotope-based data quality control and quantification, new N-glycan libraries and new algorithms

Withdrawal statement"The authors have withdrawn their manuscript because the authors need to re-organize the data and writing, meanwhile more experimental evidence from human cellular N-glycome will be added to support their viewpoints. In this preprint, only one human cell line, Acute Promyelocytic Leukemia (APL) cells, was deeply investigated to show the N-glycome landscape. To provide the shared features of human N-glycome, more human cells should be investigated. Therefore, the authors do not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author."

biochemistry↗

bioRxiv2020

ATF6 aggravates angiogenesis-osteogenesis coupling during ankylosing spondylitis by mediating FGF2 expression in chondrocytes

Although angiogenesis-osteogenesis coupling is important in ankylosing spondylitis (AS), therapeutic agents targeting the vasculature remain elusive. Here, we identified activating transcription factor 6 (ATF6) as an important regulator of angiogenesis in AS patients. Firstly, we found that ATF6 and fibroblast growth factor 2 (FGF2) levels were higher in SKG mice and AS patient cartilage. The pro-angiogenic ability of human chondrocytes was enhanced through activated ATF6-FGF2 axis following long-term stimulation with inflammatory factors, e.g. TNF-, IFN-{gamma} or IL-17. Mechanistically, ATF6 interacted with the FGF2 promotor and promoted its transcription. Treatment with the ATF6 inhibitor Ceapin-A7 inhibited angiogenesis in vitro and angiogenesis-osteogenesis coupling in vivo. ATF6 may aggravate angiogenesis-osteogenesis coupling during AS by mediating FGF2 transcription in chondrocytes, implying that ATF6 represents a promising therapeutic target for AS.

molecular biology↗

bioRxiv2020

Prioritization of osteoporosis-associated GWAS SNPs using epigenomics and transcriptomics

Genetic risk factors for osteoporosis, a prevalent disease associated with aging, have been examined in many genome-wide association studies (GWAS). A major challenge is to prioritize transcription-regulatory GWAS-derived variants that are likely to be functional. Given the critical role of epigenetics in gene regulation, we have used an unusual epigenetics- and transcription-based approach to identify credible regulatory SNPs relevant to osteoporosis from 38 reported BMD GWAS. Using Roadmap databases, we prioritized SNPs based upon their overlap with strong enhancer or promoter chromatin preferentially in osteoblasts relative to 11 heterologous cell culture types. The selected SNPs also had to overlap open chromatin (DNaseI-hypersensitive sites) and DNA sequences predicted to bind to osteoblast-relevant transcription factors in an allele-specific manner. From >50,000 GWAS-derived SNPs, we identified 16 novel and credible regulatory SNPs (Tier-1 SNPs) for osteoporosis risk. Their associated genes, BICC1, LGR4, DAAM2, NPR3, or HMGA2, are involved in osteoblastogenesis or bone homeostasis and regulate cell signaling or enhancer function. Four of them are preferentially expressed in osteoblasts. BICC1, LGR4, and DAAM2 play important roles in canonical Wnt signaling, a pathway critical to bone formation and repair. The transcription factors that are predicted to bind to the Tier-1 SNP-containing DNA sequences also have bone-related functions. For the seven Tier-1 SNPs near the 5 end of BICC1, examination of eQTL overlap and the distribution of BMD-increasing alleles suggests that at least one SNP in each of two clusters contributes to inherited osteoporosis risk. Our study not only illustrates a method that can be used to identify novel BMD-related causal regulatory SNPs for future study, but also reveals evidence that some of the Tier-1 SNPs exert their effects on BMD risk indirectly through little-studied noncoding RNA genes, which in turn may control the nearby bone-related protein-encoding gene.

bioinformatics↗

bioRxiv2020

A systematic dissection of human primary osteoblasts in vivo at single-cell resolution

Osteoblasts are multifunctional bone cells, which play essential roles in bone formation, angiogenesis regulation, as well as maintenance of hematopoiesis. Although both in vivo and in vitro studies on mice have identified several potential osteoblast subtypes based on their different transition stages or biological responses to external stimuli, the categorization of primary osteoblast subtypes in vivo in humans has not yet been achieved. Here, we used single-cell RNA sequencing (scRNA-seq) to perform a systematic cellular taxonomy dissection of freshly isolated human osteoblasts. Based on the gene expression patterns and cell lineage reconstruction, we identified three distinct cell clusters including preosteoblasts, mature osteoblasts, and an undetermined rare osteoblast subpopulation. This novel subtype was mainly characterized by the nuclear receptor subfamily 4 group A member 1 and 2 (NR4A1 and NR4A2), and its existence was confirmed by immunofluorescence staining. Trajectory inference analysis suggested that the undetermined cluster, together with the preosteoblasts, are involved in the regulation of osteoblastogenesis and also give rise to mature osteoblasts. Investigation of the biological processes and signaling pathways enriched in each subpopulation revealed that in addition to bone formation, preosteoblasts and undetermined osteoblasts may also regulate both angiogenesis and hemopoiesis. Finally, we demonstrated that there are systematic differences between the transcriptional profiles of human osteoblasts in vivo and mouse osteoblasts both in vivo and in vitro, highlighting the necessity for studying bone physiological processes in humans rather than solely relying on mouse models. Our findings provide novel insights into the cellular heterogeneity and potential biological functions of human primary osteoblasts at the single-cell level, which is an important and necessary step to further dissect the biological roles of osteoblasts in bone metabolism under various (patho-) physiological conditions.

bioinformatics↗

bioRxiv2020

Isolating multiple formats of human monoclonal neutralizing antibodies against SARS-CoV-2 by in vitro site-directed antibody screening

Neutralizing antibody is one of the most effective interventions for acute pathogenic infection. Currently, over three million people have been identified for SARS-CoV-2 infection but SARS-CoV-2-specific vaccines and neutralizing antibodies are still lacking. SARS-CoV-2 infects host cells by interacting with angiotensin converting enzyme-2 (ACE2) via the S1 receptor-binding domain (RBD) of its surface spike glycoprotein. Therefore, blocking the interaction of SARS-CoV-2-RBD and ACE2 by antibody would cause a directly neutralizing effect against virus. In the current study, we selected the ACE2 interface of SARS-CoV-2-RBD as the targeting epitope for neutralizing antibody screening. We performed site-directed screening by phage display and finally obtained one IgG antibody (4A3) and several domain antibodies. Among them, 4A3 and three domain antibodies (4A12, 4D5, and 4A10) were identified to act as neutralizing antibodies due to their capabilities to block the interaction between SARS-CoV-2-RBD and ACE2-positive cells. The domain antibody 4A12 was predicted to have the best accessibility to all three ACE2-interfaces on the spike homotrimer. Pseudovirus and authentic SARS-CoV-2 neutralization assays showed that all four antibodies could potently protect host cells from virus infection. Overall, we isolated multiple formats of SARS-CoV-2-neutralizing antibodies via site-directed antibody screening, which could be promising candidate drugs for the prevention and treatment of COVID-19.

immunology↗

bioRxiv2020

Single-cell RNA sequencing deconvolutes the in vivo heterogeneity of human bone marrow-derived mesenchymal stem cells

Bone marrow-derived mesenchymal stem cells (BM-MSCs) are multipotent stromal cells, which have a critical role in the maintenance of skeletal tissues such as bone, cartilage, and the fat found in bone marrow. In addition to providing microenvironmental support for hematopoietic processes, BM-MSCs can differentiate into various mesodermal lineages including osteoblast/osteocyte, chondrocyte, and adipocyte cells that are crucial for bone metabolism. While BM-MSCs have high cell-to-cell heterogeneity in gene expression, the cell subtypes that contribute to this heterogeneity in vivo in humans have not been characterized. To investigate the transcriptional diversity of BM-MSCs, we applied single-cell RNA sequencing (scRNA-seq) on freshly isolated CD271+ BM-derived mononuclear cells (BM-MNCs) from two human subjects. We successfully identified LEPRhiCD45low BM-MSCs within the CD271+ BM-MNC population, and further codified the BM-MSCs into distinct subpopulations corresponding to the osteogenic, chondrogenic, and adipogenic differentiation trajectories, as well as terminal-stage quiescent cells. Biological functional annotations of transcriptomes suggest that osteoblast precursors may induce angiogenesis coupled with osteogenesis, and chondrocyte precursors may have the potential to differentiate into myocytes. We discovered transcripts for several cluster of differentiation (CD) markers that were highly expressed (e.g., CD167b, CD91, CD130 and CD118) or absent (e.g., CD74, CD217, CD148 and CD68) in BM-MSCs and could be novel markers for human BM-MSC purification. This study is the first systematic in vivo dissection of human BM-MSCs cell subtypes at the single-cell resolution, revealing insight into the extent of their cellular heterogeneity and bone homeostasis.

cell biology↗

bioRxiv2020

Glycyrrhizic acid improves cognitive levels of aging mice by regulating T/B cell proliferation

Dear Editor,Licorice is the root of Glycyrrhiza glabra L. (Leguminosae), which grows in various warm climates such as the Middle East, Asia, and Southern Europe. It is one of the oldest known medicinal herbs and is referred to as "the father of herbal medicine." Glycyrrhizic acid (GA) (Fig. 1A), a triterpenoid saponin, is a major component of licorice. It has a variety of pharmacological activities such as anti-inflammatory, antioxidant, anticancer, neuroprotective, and immune effects, among others.[1]. Previous studies indicated that GA produces robust neuroprotection via the modulation of anti-apoptotic and pro-apoptotic factors, primarily through the ERK signaling pathway and its anti-inflammatory properties against high-mobility group box 1 phosphorylation and the suppression of inflammatory cytokine induction [2-4]. These results were based on pathological models. Although numerous pathways have been implicated in the neuroprotective effects of GA, the molecular mechanisms are not yet completely understood. In this study, we aimed to investigate the effects of GA in preventing age-related immune involution and cognitive disorders, the relationship between immune involution and cognition, and the underlying molecular mechanisms. O_FIG O_LINKSMALLFIG WIDTH=141 HEIGHT=200 SRC="FIGDIR/small/008821v1_fig1.gif" ALT="Figure 1"> View larger version (43K): org.highwire.dtl.DTLVardef@5a123dorg.highwire.dtl.DTLVardef@163a085org.highwire.dtl.DTLVardef@1bfadb6org.highwire.dtl.DTLVardef@125b2ae_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure 1.C_FLOATNO Glycyrrhizic acid ameliorated cognitive declines in aged mice, improved CBF and increased T and B cell numbers. A. The chemical structure of glycyrrhizic acid (GA). B. Escape latencies from six consecutive daily tests. C. The first platform-finding latencies in the probe trial. D. The time spent in the target quadrant in the probe trial. E. The target platform crossing times in the probe trial. F. Images of CBF in C57BL/6 mice. G. Quantification of blood perfusion in the ipsilateral hemisphere. H. Bubble diagram showing GA treatment influence on hematopoietic cell lineage. I. Heatmap showing actively expressed genes related to T cells in the hematopoietic cell lineage of control and GA-treated C57BL/6 mice. J. Representative FACS plots showing CD3e+ T and CD45R/B220+ B cells in the spleens of C57BL mice; bar graphs showing the statistical analysis of T and B cells in the spleens of C57BL mice. K. Representative FACS plots showing CD3e+ T and CD45R/B220+ B cells in the blood of C57BL mice; Bar graphs showing the statistical results of T and B cell analyses in the blood of C57BL mice. GA (5mg/kg, once every other day); Data represent mean {+/-} SEM (n = 8 per group). *P < 0.05, **P < 0.01, ***P < 0.001 vs. Ctrl. C_FIG

animal behavior and cognition↗

bioRxiv2020

Accurate Diagnosis of Colorectal Cancer Based on Histopathology Images Using Artificial Intelligence

BackgroundAccurate and robust pathological image analysis for colorectal cancer (CRC) diagnosis is time-consuming and knowledge-intensive, but is essential for CRC patients treatment. The current heavy workload of pathologists in clinics/hospitals may easily lead to unconscious misdiagnosis of CRC based on their daily image analyses. MethodsBased on a state-of-the-art transfer-learned deep convolutional neural network in artificial intelligence (AI), we proposed a novel patch aggregation strategy for clinic CRC prediction/diagnosis using weakly labeled pathological whole slide image (WSI) patches. This approach was trained and validated using an unprecedented and enormously large number of 170,099 patches, >14,680 WSIs, from >9,631 subjects that covered diverse and representative clinical cases from multi-independent-sources across China, U.S., and Germany. ResultsOur innovative AI tool was consistently nearly perfectly agreed with (average Kappa-statistic 0.896) and even often better than most of the experienced expert pathologists when tested in diagnosing CRC WSIs from multi-centers. The average area under the receiver operating characteristics curve (AUC) of AI was greater than that of the pathologists (0.981 vs 0.970) and achieved the best performance among the application of other AI methods to CRC diagnosis. Our AI-generated heatmap highlights the image regions of cancer tissue/cells. ConclusionsThis first-ever generalizable AI system can handle large amounts of WSIs consistently and robustly without potential bias due to fatigue commonly experienced by clinical pathologists. Hence, it will drastically alleviate the heavy clinical burden of daily pathology diagnosis, and improve the treatment for CRC patients. This tool is generalizable to other cancer diagnosis based on image recognition.

pathology↗

bioRxiv2020

Epigenetic reprogramming towards mesenchymal-epithelial transition in ovarian cancer-associated mesenchymal stem cells drives metastasis

Ovarian cancer develops early intra-peritoneal metastasis establishing a supportive tumor microenvironment (TME) through reprogramming normal mesenchymal stem cells into carcinoma-associated mesenchymal stem cells (CA-MSCs). CA-MSCs are the stromal stem cell of the TME, supporting cancer growth, increasing desmoplasia, angiogenesis and chemotherapy resistance. We demonstrate epigenetic reprogramming drives CA-MSC formation via enhancer-enriched DNA hypermethylation, altered chromatin accessibility and differential histone modifications inducing a partial mesenchymal to epithelial transition (MET) increasing adhesion to tumor cells. Direct CA-MSC:tumor cell interactions, confirmed in patient ascites, facilitate ovarian cancer metastasis through co-migration. WT1, a developmental mediator of MET, and EZH2, mediate CA-MSC epigenetic reprogramming. WT1 overexpression induces CA-MSC conversion while WT1 knock-down, along with EZH2 inhibition, blocks CA-MSC formation. EZH2 inhibition subsequently decreases intra-abdominal metastasis. SignificanceThis work presents a new paradigm of stromal reprogramming involving a partial mesenchymal to epithelial transition. Rather than a classic tumor cell epithelial to mesenchymal transition, metastasis relies on epigenetic rewiring of a CA-MSC allowing enhanced tumor cell binding and co-migration with tumor cells to form metastasis. Indeed, CA-MSCs in complex with tumor cells are abundant in patient ascites. Reversion of CA-MSCs to normal MSCs is observed in patients achieving complete response with neoadjuvant therapy. Identification of WT1 and EZH2 as mediators of the epigenetic reprogramming of CA-MSCs present potential targets to block the formation of CA-MSCs thus disrupting the TME and limiting ovarian cancer metastasis.

cancer biology↗

bioRxiv2020

Structural insights into secretory immunoglobulin A and its interaction with a pneumococcal adhesin

Secretory Immunoglobulin A (SIgA) is the most abundant antibody at the mucosal surface. SIgA possesses two additional subunits besides IgA: the joining chain (J-chain) and secretory component (SC). SC is the ectodomain of the polymeric immunoglobulin receptor (pIgR), which functions to transport IgA to the mucosa. The underlying mechanism of how the J-chain and pIgR/SC facilitates the assembly and secretion of SIgA remains to be understood. During the infection of Streptococcus pneumoniae, a pneumococcal adhesin SpsA hijacks SIgA and unliganded pIgR/SC to evade host defense and gain entry to human cells. How SpsA specifically targets SIgA and pIgR/SC also remains unclear. Here we report a cryo-electron microscopy structure of the Fc region of human IgA1 (Fc) in complex with J-chain and SC (Fc-J-SC), which reveals the organization principle of SIgA. We also present the structure of Fc-J-SC in complex with SpsA, which uncovers the specific interaction between SpsA and human pIgR/SC. These results advance the molecular understanding of SIgA and shed light on the pathogenesis of S. pneumoniae.

immunology↗

bioRxiv2020

In vitro activities of the tetrazole SHR8008 compared to itraconazole and fluconazole against Candida and Cryptococcus species

Candida and Cryptococcus are the main pathogens of clinical fungal infection associated with high morbidity and mortality. SHR8008 (in fact, this is the only official name in China and it is called VT-1161 by FDA) is a novel tetrazole agent that selectively inhibits fungal CYP51A compared to mammalian cytochrome P450 enzymes to achieve a better antifungal effect. The in vitro activities of SHR8008 and its comparators itraconazole and fluconazole were determined in 127 Candida and 50 Cryptococcus strains isolated from Chinese patients in the last 2 years by Invasive Fungal Infection Group. The MICs of SHR8008 and other triazoles were measured by the Clinical and Laboratory Standards Institute guidelines M27-E4. For Candida spp., SHR8008 (geometric mean MIC=0.078 g/mL) was 6.5-fold and 11.2-fold more potent than itraconazole and fluconazole, respectively. There is a good correlation of MICs between SHR8008 and itraconazole/fluconazole. The MIC values of SHR8008 against Candida glabrata and Candida tropicalis were significantly lower than those of fluconazole, while for Candida albicans and Candida parapsilosis, the differences between SHR8008 and fluconazole were not statistically significant, either. For Cryptococcus spp., SHR8008 (geometric mean MIC=0.024 g/mL) was 21.7-fold and 104.5-fold more potent than itraconazole and fluconazole, respectively. Against the seven Cryptococcus neoformans isolates with elevated fluconazole MICs ([≥]8g/mL based on the MIC90 value for this azole), SHR8008 maintained potent activity, with MICs ranging between 0.031 and 0.5 g/mL. The results showed that tetrazole SHR8008 was more promising in the treatment of Candida and Cryptococcus infection than itraconazole and fluconazole.

microbiology↗

bioRxiv2019

Advanced Whole Genome Sequencing Using a Complete PCR-free Massively Parallel Sequencing (MPS) Workflow

BackgroundSystematic errors can be introduced from DNA amplification during massively parallel sequencing (MPS) library preparation and sequencing array formation. Polymerase chain reaction (PCR)-free genomic library preparation methods were previously shown to improve whole genome sequencing (WGS) quality on the Illumina platform, especially in calling insertions and deletions (InDels). We hypothesized that substantial InDel errors continue to be introduced by the remaining PCR step of DNA cluster generation. In addition to library preparation and sequencing, data analysis methods are also important for the accuracy of the output data.In recent years, several machine learning variant calling pipelines have emerged, which can correct the systematic errors from MPS and improve the data performance of variant calling. ResultsHere, PCR-free libraries were sequenced on the PCR-free DNBSEQ arrays from MGI Tech Co., Ltd. (referred to as MGI) to accomplish the first true PCR-free WGS which the whole process is truly not only PCR-free during library preparation but also PCR-free during sequencing. We demonstrated that PCR-based WGS libraries have significantly (about 5 times) more InDel errors than PCR-free libraries.Furthermore, PCR-free WGS libraries sequenced on the PCR-free DNBSEQ platform have up to 55% less InDel errors compared to the NovaSeq platform, confirming that DNA clusters contain PCR-generated errors.In addition, low coverage bias and less than 1% read duplication rate was reproducibly obtained in DNBSEQ PCR-free using either ultrasonic or enzymatic DNA fragmentation MGI kits combined with MGISEQ-2000. Meanwhile, variant calling performance (single-nucleotide polymorphisms (SNPs) F-score>99.94%, InDels F-score>99.6%) exceeded widely accepted standards using machine learning (ML) methods (DeepVariant or DNAscope). ConclusionsEnabled by the new PCR-free library preparation kits, ultra high-thoughput PCR-free sequencers and ML-based variant calling, true PCR-free DNBSEQ WGS provides a powerful solution for improving WGS accuracy while reducing cost and analysis time, thus facilitating future precision medicine, cohort studies, and large population genome projects.

genomics↗

bioRxiv2019

Osteoporosis- and obesity-risk interrelationships: An epigenetic analysis of GWAS-derived SNPs at the developmental gene TBX15

A major challenge in translating findings from genome-wide association studies (GWAS) to biological mechanisms is pinpointing functional variants because only a very small percentage of variants associated with a given trait actually impact the trait. We used an extensive epigenetics, transcriptomics, and genetics analysis of the TBX15/WARS2 neighborhood to prioritize this regions best-candidate causal variants for the genetic risk of osteoporosis (estimated bone density, eBMD) and obesity (waist-hip ratio or waist circumference adjusted for body mass index). TBX15 encodes a transcription factor that is important in bone development and adipose biology. Manual curation of 692 GWAS-derived variants gave eight strong candidates for causal SNPs that modulate TBX15 transcription in subcutaneous adipose tissue (SAT) or osteoblasts, which highly and specifically express this gene. None of these SNPs were prioritized by Bayesian fine-mapping. The eight regulatory causal SNPs were in enhancer or promoter chromatin seen preferentially in SAT or osteoblasts at TBX15 intron-1 or upstream. They overlap strongly predicted, allele-specific transcription factor binding sites. Our analysis suggests that these SNPs act independently of two missense SNPs in TBX15. Remarkably, five of the regulatory SNPs were associated with eBMD and obesity and had the same trait-increasing allele for both. We found that WARS2 obesity-related SNPs can be ascribed to high linkage disequilibrium with TBX15 intron-1 SNPs. Our findings from GWAS index, proxy, and imputed SNPs suggest that a few SNPs, including three in a 0.7-kb cluster, act as causal regulatory variants to fine-tune TBX15 expression and, thereby, affect both obesity and osteoporosis risk.

bioinformatics↗

bioRxiv2019

Binding to m6A RNA promotes YTHDF2-mediated phase separation

As the most abundant modification on mRNA in mammal, N6-Methyladenosine (m6A) has been demonstrated to play important roles in various biological processes including mRNA splicing, translation and degradation. m6A reader proteins have been shown to play central roles in these processes. One of the m6A readers, YTHDF2 is localized to the P granules, which are liquid-like droplets where RNA degradation occurs. How YTHDF2 is localized to P granules is unknown. Here we provide evidence that YTHDF2 forms liquid droplets and phase separate, mediated by its low complexity (LC) domains. Interestingly, the ability to phase separate is robustly stimulated by m6A RNAs in vitro. In vivo, YTHDF2 phase separation may in fact be dependent on m6A RNA and YTHDF2 binding to m6A RNA, since a YTHDF2 m6A-binding defective mutant or a wildtype YTHDF2 assayed in cells lacking m6A RNAs, both fail to phase separate. The ability of phase separate is not limited to YTHDF2; we find other members of the YTH-domain m6A readers can also undergo phase separation. Our findings suggest that m6A RNA induced phase separation of m6A readers may play an important role in their distributions to different phase-separated compartments in cells.

molecular biology↗

bioRxiv2019

BRD2 regulation of sigma-2 receptor expression upon cytosolic cholesterol deprivation

Traditionally a pharmacologic target for antipsychotic treatment, the sigma-2 receptor (S2R) was recently implicated in cholesterol homeostasis. Here we investigated the transcriptional regulation of S2R by the Bromo/ExtraTerminal epigenetic reader family (BETs, including BRD2, 3, 4) upon cholesterol perturbation.\n\nCytosolic cholesterol deprivation was induced using an export blocker of lysosomal cholesterol in ARPE19 cells. This condition upregulated mRNA and protein levels of S2R, and of SREBP2 but not SREBP1, transcription factors key to cholesterol/fatty acid metabolism. Silencing BRD2 but not BRD4 (though widely deemed as a master regulator) or BRD3 prevented S2R upregulation induced by cholesterol deprivation. Silencing SREBP2 but not SREBP1 diminished S2R expression. Furthermore, BRD2 co-immunoprecipitated with the SREBP2 transcription-active N-terminal domain, and chromatin immunoprecipitation-qPCR showed a BRD2 occupancy at the S2R gene promoter.\n\nIn summary, this study reveals a novel BRD2/SREBP2 cooperative regulation of S2R transcription in response to cytosolic cholesterol deprivation, thus shedding new light on epigenetic control of cholesterol biology.

biochemistry↗

bioRxiv2019

Multilayered N-glycoproteomics reveals impaired N-glycosylation promoting Alzheimer’s disease

Alzheimers disease (AD) is one of the most common neurodegenerative diseases that currently lacks clear pathogenesis and effective treatment. Protein glycosylation is ubiquitous in brain tissue and site-specific analysis of N-glycoproteome, which is technically challenging, can advance our understanding of the glycoproteins role in AD. In this study, we profiled the multilayered variations in proteins, N-glycosites, N-glycans, and in particular site-specific N-glycopeptides in the APP/PS1 and wild type mouse brain through combining pGlyco 2.0 strategy with other quantitative N-glycoproteomic strategies. The comprehensive brain N-glycoproteome landscape was constructed, and rich details of the heterogeneous site-specific protein N-glycosylations were exhibited. Quantitative analyses explored generally downregulated N-glycosylation involving proteins such as glutamate receptors, as well as fucosylated and oligo-mannose type glycans in APP/PS1 mice versus wild type mice. Moreover, our preliminary functional study revealed that N-glycosylation was crucial for the membrane localization of NCAM1 and for maintaining the excitability and viability of neuron cells. Our work offered a panoramic view of the N-glycoproteomes in Alzheimers disease and revealed that generally impaired N-glycosylation promotes Alzheimers disease progression.

systems biology↗

bioRxiv2019

Understanding the Mechanistic Contribution of Herbal Extracts in Compound Kushen Injection with Transcriptome Analysis

Herbal compatibility is the knowledge of which herbs to combine in traditional Chinese medicine (TCM) formulations. The lack of understanding of herbal compatibility is one of the key problems for the application and popularization of TCM in western society. Because of the chemical complexity of herbal medicines, it is simpler to begin to conduct compatibility research based on herbs rather than component plant secondary metabolites. We have used transcriptome analysis to explore the effects and interactions of two plant extracts (Kushen and Baituling) combined in Compound Kushen Injection (CKI). Based on shared chemical compounds and in vitro cytotoxicity comparisons, we found that both the major compounds in CKI, and the cytotoxicity effects of CKI were mainly derived from the extract of Kushen (Sophorae flavescentis). We generated and analyzed transcriptome data from MDA-MB-231 cells treated with single-herb extracts or CKI and results showed that Kushen contributed to the perturbation of the majority of cytotoxicity/cancer related pathways in CKI such as cell cycle and DNA replication. We also found that Baituling (Heterosmilax yunnanensis Gagnep) could not only enhance the cytotoxic effects of Kushen in CKI, but also activate immune-related pathways. Our analyses predicted that IL-1{beta} gene expression was upregulated by Baituling in CKI and we confirmed that IL-1{beta} protein expression was increased using an ELISA assay. Altogether, these findings help to explain the rationale for combining Kushen and Baituling in CKI, and transcriptome analysis using single herb extracts is an effective method for understanding herbal compatibility in TCM.

pharmacology and toxicology↗