Biology subjects

Qin, T.

Publications and source records attributed to Qin, T..

48 records · Page 3Linked to original sources

bioRxiv2022

Zinc Finger MYND-Type Containing 8 (ZMYND8) is epigenetically regulated in mutant Isocitrate Dehydrogenase 1 (IDH1) glioma to promote radioresistance

Mutant isocitrate dehydrogenase 1 (mIDH1) alters the epigenetic regulation of chromatin, leading to a hypermethylation phenotype in adult glioma. Establishment of glioma-specific methylation patterns by mIDH1 reprogramming drives oncogenic features of cancer metabolism, stemness and therapeutic resistance. This work focuses on identifying gene targets epigenetically dysregulated by mIDH1. Treatment of glioma cells with mIDH1 specific inhibitor AGI-5198, upregulated gene networks involved in replication stress. Specifically, we found that the expression of ZMYND8, a regulator of DNA damage response was decreased in three patient-derived glioma cell cultures (GCC) after treatment with AGI-5198. ZMYND8 functions as a chromatin reader to modulate enhancer activity and recruit DNA repair machinery. Knockdown of ZMYND8 expression sensitized mIDH1 GCCs to radiotherapy marked by decreased cellular viability. Following IR, mIDH1 glioma cells with ZMYND8 knockout (KO) exhibit significant phosphorylation of ATM and sustained {gamma}H2AX activation. ZMYND8 KO mIDH1 GCCs were further responsive to IR when treated with either BRD4 or HDAC inhibitors. The accumulation of DNA damage in ZMYND8 KO mIDH1 GCCs promoted the phosphorylation of cell cycle checkpoint proteins Chk1 and Chk2. In addition, the recruitment of ZMYND8 to sites of DNA damage has been shown to be PARP- dependent. PARP inhibition further enhanced the efficacy of radiotherapy in ZMYND8 KO mIDH1 glioma cells. These findings indicate the impact of ZMYND8 in the maintenance of genomic integrity and repair of IR-induced DNA damage in mIDH1 glioma. Translational RelevanceOur understanding of radioresistance mechanisms in patient-derived glioma cell cultures (GCC) that endogenously express mIDH1-R132H are limited. We have uncovered a novel gene target Zinc Finger MYND-Type Containing 8 (ZMYND8) that is downregulated following treatment of human mIDH1 GCCs with mIDH1 specific inhibitors. We demonstrate that suppression of ZMYND8 expression by shRNA knockdown or genetic knockout reduces the cellular viability of mIDH1 GCCs to ionizing radiation (IR). Our findings reveal an epigenetic vulnerability of mIDH1 GCCs to ZMYND8 knockout (KO) which results in impaired resolution of IR-induced DNA damage and induction of cell cycle arrest. Additionally, ZMYND8 KO mIDH1 GCCs display increased radiosensitivity to inhibition of epigenetic regulators BRD4, HDAC, and PARP which could be mediated by enhanced replicative stress.

cancer biology↗

bioRxiv2021

Functional Anatomy of the TDP-43 Redox Sensor

TAR binding protein 43 (TDP-43) is an RNA binding protein that assists in the maturation, export and sub-cellular localization of mRNA. The carboxyl terminal 153 residues of TDP-43 are of low sequence complexity and allow for self-association of the protein in a manner leading to its phase separation from an aqueous environment. These interactions assist TDP-43 in forming cytoplasmic RNA granules involved in the transport of mRNA for localized translation. Self-association of the TDP-43 low complexity (LC) domain is facilitated by a region of twenty five residues that are of extreme evolutionary conservation. The molecular basis for self-adherence of the protein through this region has been illuminated by a combination of structural and biochemical studies, allowing definition of a morphologically specific cross-{beta} structure predicted to be weakly assembled by main chain hydrogen bonds. In this study we have investigated the importance of individual, Pauling hydrogen bonds hypothesized to facilitate self-adherence of the TDP-43 LC domain.

biochemistry↗

bioRxiv2021

Characterization of the Endogenous DAF-12 Ligand and Its Use as an Anthelmintic Agent in Strongyloides stercoralis

A prevalent feature of Strongyloides stercoralis is a life-long and potentially lethal infection that is due to the nematode parasites ability to autoinfect and, thereby, self-replicate within its host. Here, we investigated the role of the parasites nuclear receptor, Ss-DAF-12, in governing infection. We identified {Delta}7-DA as the endogenous Ss-DAF-12 ligand and elucidated the hormones biosynthetic pathway. Genetic loss of function of the ligands rate-limiting enzyme demonstrated that {Delta}7-DA synthesis is necessary for parasite reproduction, whereas its absence is required for development of infectious larvae. Availability of the ligand permits Ss-DAF-12 to function as an on/off switch governing autoinfection, making it vulnerable to therapeutic intervention. In a preclinical model of hyperinfection, pharmacologic activation of DAF-12 suppressed autoinfection and markedly reduced lethality. Moreover, when {Delta}7-DA was administered with ivermectin, the current but limited drug of choice for treating strongyloidiasis, the combinatorial effects of the two drugs resulted in a near cure of the disease.

pharmacology and toxicology↗

bioRxiv2021

Therapeutic reversal of prenatal pontine ID1 signaling in DIPG

Diffuse intrinsic pontine glioma (DIPG) is a highly aggressive brain tumor with rare survival beyond two years. This poor prognosis is largely due to the tumors highly infiltrative and invasive nature. Previous reports demonstrate upregulation of the transcription factor ID1 with H3K27M and ACVR1 mutations, but this has not been confirmed in human tumors or therapeutically targeted. We developed an in utero electroporation (IUE) murine H3K27M-driven tumor model, which demonstrates increased ID1 expression in H3K27M- and ACVR1-mutated tumor cells. In human tumors, elevated ID1 expression is associated with H3K27M/ACVR1-mutation, brainstem location, and reduced survival. The ID1 promoter demonstrates a similar active epigenetic state in H3K27M tumor cells and murine prenatal hindbrain cells. In the developing human brain, ID1 is expressed highest in oligo/astrocyte-precursor cells (OAPCs). These ID1+/SPARCL1+ cells share a transcriptional program with astrocyte-like (AC-like) DIPG cells, and demonstrate upregulation of gene sets involved with regulation of cell migration. Both genetic and pharmacologic [cannabidiol (CBD)] suppression of ID1 results in decreased DIPG cell invasion/migration in vitro and invasion/tumor growth in multiple in vivo models. CBD reduces proliferation through reactive oxygen species (ROS) production at low micromolar concentrations, which we found to be achievable in the murine brainstem. Further, pediatric high-grade glioma patients treated off-trial with CBD (n=15) demonstrate tumor ID1 reduction and improved overall survival compared to historical controls. Our study identifies that ID1 is upregulated in DIPG through reactivation of a developmental OAPC transcriptional state, and ID1-driven invasiveness of DIPG is therapeutically targetable with CBD. One Sentence SummaryThe transcription factor ID1 is upregulated in a subset of DIPG tumor cells, and ID1-driven invasiveness is therapeutically targetable with CBD.

cancer biology↗

bioRxiv2021

Analysis of Spleen Histopathology, Splenocyte composition and Hematological Parameters in Mice Infected with Plasmodium berghei K173

Malaria is a fatal disease that presents clinically as a continuum of symptoms and severity, which are determined by complex host-parasite interactions. Clearance of infection is believed to be accomplished by the spleen and mononuclear phagocytic system (MPS), both in the presence and absence of artemisinin treatment. The spleen filters infected RBCs from circulation through immune-mediated recognition of the infected RBCs followed by phagocytosis. Using different strains of mice infected with P. berghei K173 (PbK173), the mechanisms leading to splenomegaly, histopathology, splenocyte activation and proliferation, and their relationship to control of parasitemia and host mortality were examined. Survival time of mice infected with PbK173 varied, although the infection was uniformly lethal. Mice of the C57BL/6 strain were the most resistant, while mice of the strain ICR were the most susceptible. BALB/c and KM mice were intermediate. In the course of PbK173 infection, both strains of mice experienced significant splenomegaly. Parasites were observed in the red pulp at 3 days post infection in all animals. All spleens retained late trophozoite stages as well as a fraction of earlier ring-stage parasites. The percentages of macrophages in infected C57BL/6 and KM mice were higher than uninfected mice on 8 dpi. Spleens of infected ICR and KM mice exhibited structural disorganization and remodeling. Furthermore, parasitemia was significantly higher in KM versus C57BL/6 mice at 8 dpi. The percentages of macrophages in ICR infected mice were lower than uninfected mice, and the parasitemia was higher than other strains. The results presented here demonstrate the rate of splenic mechanical filtration and the splenic macrophages likely contribute to an individuals total parasite burden. This in turn can influence the pathogenesis of malaria. Finally, different genetic backgrounds of mice have different splenic mechanisms for controlling malaria infection.

microbiology↗

bioRxiv2020

Comprehensive enhancer-target gene assignments improve gene set level interpretation of genome-wide regulatory data

Revealing the gene targets of distal regulatory elements is challenging yet critical for interpreting regulome data. Experiment-derived enhancer-gene links are restricted to a small set of enhancers and/or cell types, while the accuracy of genome-wide approaches remains elusive due to the lack of a systematic evaluation. We combined multiple spatial and in silico approaches for defining enhancer locations and linking them to their target genes aggregated across >500 cell types, generating 1,860 human genome-wide distal Enhancer to Target gene Definitions (EnTDefs). To evaluate performance, we used gene set enrichment testing on 87 independent ENCODE ChIP-seq datasets of 34 transcription factors (TFs) and assessed concordance of results with known TF Gene Ontology (GO) annotations., assuming that greater concordance with TF-GO annotation signifies better enrichment results and thus more accurate enhancer-to-gene assignments. Notably, the top ranked 741 (40%) EnTDefs significantly outperformed the common, naive approach of linking distal regions to the nearest genes (FDR < 0.05), and the top 10 ranked EnTDefs performed well when applied to ChIP-seq data of other cell types. These general EnTDefs also showed comparable performance to EnTDefs generated using cell-type-specific data. Our findings illustrate the power of our approach to provide genome-wide interpretation regardless of cell type.

bioinformatics↗

bioRxiv2020

High-resolution architecture of human epiphysis formation

Human limb skeletal system consists of both bone and cartilage which originated from fetal cartilage. However, the roadmap of chondrocyte divergent differentiation to bone and articular cartilage has yet to be established. Epiphysis possesses articular cartilage, growth plate and the secondary ossification center (SOC), making it an ideal model to uncover the trajectory of chondrocyte divergent differentiation. Here, we mapped differentiation trajectory of human chondrocyte during postnatal finger epiphysis development by using single-cell RNA sequencing. Our results uncovered that chondroprogenitors have two differentiation pathways to hypertrophic chondrocytes during ossification, and one pathway to articular chondrocytes for formation of cartilages. Interestingly, we found that, as an addition to the known typical endochondral ossification path from resting, proliferative to hypertrophic chondrocytes, there was a bypass by which chondroprogenitors differentiate into hypertrophic chondrocytes without proliferative stage. Furthermore, our results revealed two new chondrocyte subpopulations (bypass chondrocytes as it appeared in the ossification bypass, and ID1+ chondroblasts in articular chondrocyte path) during postnatal epiphysis development in addition to six well-known subpopulations. Overall, our study provides a comprehensive roadmap of chondrocyte differentiation in human epiphysis thereby expanding the knowledge of bone and articular cartilage, which could be utilized to design biotherapeutics for bone and articular cartilage regeneration.

developmental biology↗

bioRxiv2019

Subarachnoid hemorrhage leads to early and persistent functional connectivity and behavioral changes in mice

Aneurysmal subarachnoid hemorrhage (SAH) leads to significant long-term cognitive deficits. Studies in survivors of SAH show an association between persistent cognitive deficits and alterations in resting state functional connectivity (RSFC). However, modalities commonly used to assess RSFC in humans, such as fMRI, have practical limitations in small animals. Therefore, we used non-invasive functional optical intrinsic signal imaging to determine the effect of SAH on measures of RSFC in mice at early (day 4), intermediate (1 month), and late (3 months) time points after prechiasmatic arterial blood injection. We assessed Morris water maze, open field test, Y-maze, and rotarod performance from approximately 2 weeks to 3 months after SAH induction. We found qualitative and quantitative differences in seed-based connectivity maps between sham and SAH mice. SAH reduced motor, retrosplenial and visual seed-based connectivity indices, which persisted in retrosplenial and visual cortex seeds at 3 months. Seed-to-seed connectivity analysis confirmed attenuation of correlation coefficients in SAH mice, which persisted in predominantly posterior network connections at later time points. Seed-independent global and interhemispheric indices of connectivity revealed decreased correlations following SAH for at least 1 month. SAH led to Morris water maze hidden platform and open field deficits at 2 weeks, and Y-maze deficits for at least 3 months, without altering rotarod performance. In conclusion, experimental SAH leads to early and persistent alterations both in hemodynamically-derived measures of RSFC and in cognitive performance.

neuroscience↗

bioRxiv2019

Single-Cell RNA-Seq Reveals Novel Mitochondria-related Musculoskeletal Cell Populations during Adult Axolotl Limb Regeneration Process

While the capacity to regenerate tissues or limbs is limited in mammals including humans, unlike us, axolotls are able to regrow entire limbs and major organs. The wound blastema have been extensively studied in limb regeneration. However, due to the inadequate characterization and coordination of cell subpopulations involved in the regeneration process, it hinders the discovery of the key clue for human limb regeneration. In this study, we applied unbiased large-scale single-cell RNA sequencing to classify cells throughout the adult axolotl limb regeneration process. We computationally identified 7 clusters in regenerating limbs, including the novel regeneration-specific mitochondria-related cluster supporting regeneration through energy providing and the COL2+ cluster contributing to regeneration through cell-cell interactions signals. We also discovered the dedifferentiation and re-differentiation of the COL1+/COL2+ cellular subpopulation and uncovered a COL2-mitochondria sub-cluster supporting the musculoskeletal system regeneration. On the basis of these findings, we reconstructed the dynamic single-cell transcriptome atlas of adult axolotl limb regenerative process, and identified the novel regenerative mitochondria-related musculoskeletal populations, which yielded deeper insights into the crucial interactions between cell clusters within the regenerative microenvironment.

genomics↗

bioRxiv2019

Degradation of miR-466d-3p by JEV NS3 facilitates viral replication and IL-1β expression

Previous studies revealed that Japanese encephalitis virus (JEV) infection alters the expression of miRNA in central nervous system (CNS). However, the mechanism of JEV infection contributes to the regulation of miRNAs in CNS remain obscure. Here, we found that a global degradation of mature miRNA in mouse brain and neuroblastoma cells after JEV infection. In additional, the integrative analysis of miRNAs and mRNAs suggests that those down-regulated miRNAs are primarily targeted inflammation genes and the miR-466d-3p target the IL-1{beta} which in the middle of those inflammation genes. Transfection of miR-466d-3p decreased the IL-1{beta} expression and inhibited the JEV replication in NA cells. Interestingly, the miR-466d-3p level increased after JEV infection in the presence of cycloheximide, which indicated that viral protein expression reduces miR-466d-3p. Therefore, we generated all the JEV coding protein and demonstrated that NS3 is a potent miRNA suppressor. Furthermore, the NS3 of ZIKA virus, WNV, DENV1 and DENV2 also decreased the expression of miR-466d-3p. The in vitro unwinding assay demonstrated that the NS3 could unwind the pre-miR-466d and induce the disfunction of miRNA. Using computational models and RNA immunoprecipitation assay, we found that arginine-rich domains of NS3 are critical for pre-miRNA binding and the degradation of host miRNAs. Importantly, site-directed mutagenesis of conserved residues revealed that R226G and R202W significantly reduced the binding affinity and degradation of pre-miR-466d. Together, these results extend the helicase of Flavivirus function beyond unwinding duplex RNA to the decay of pre-miRNAs, which provides a new mechanism of NS3 in regulating miRNA pathways and promoting the neuroinflammation.\n\nAuthor SummaryHost miRNAs had been reported to regulate JEV induced inflammation in central nervous system. We found that the NS3 of JEV can reduce most of host miRNA expression. The helicase region of the NS3 specifically binds to precursors of miRNA and lead to incorrect unwinding of precursors of miRNAs which inhibits the function of miRNAs. This observation leads to two major findings. First, we identified the miR-466d-3p targets to the host IL-1{beta} and E protein of JEV, and NS3 degrades the miR-466d-3p to promote the brain inflammation and viral replication. Second, we proved that the arginine on the helicase of NS3 is the main miRNA binding sites, and the miRNA degradation by NS3 was abolished when the R226 and R202 were mutated on the NS3. These findings were also confirmed with NS3 of ZIKA virus, WNV and DENV which could decrease the expression level of miR-466d-3p to enhance the inflammation. Our study provides new insights into the molecular mechanism of encephalitis caused by JEV, and reveals several amino acid sites to further attenuate the JEV vaccine.

microbiology↗

bioRxiv2018

Poly-Enrich: Count-based Methods for Gene Set Enrichment Testing with Genomic Regions and Updates to ChIP-Enrich

Gene set enrichment (GSE) testing enhances the biological interpretation of ChIP-seq data and other large sets of genomic regions. Our group has previously introduced two GSE methods for genomic regions: ChIP-Enrich for narrow regions and Broad-Enrich for broad genomic regions, such as histone modifications. Here, we introduce new methods and extensions that more appropriately analyze sets of genomic regions with vastly different properties. First, we introduce Poly-Enrich, which models the number of peaks assigned to a gene using a generalized additive model with a negative binomial family to determine gene set enrichment, while adjusting for gene locus length (#bps associated with each gene). This is the first method that controls for locus length while accounting for the number of peaks per gene and variability among genes. We also introduce a flexible weighting approach to incorporate region scores, a hybrid enrichment approach, and support for new gene set databases and reference genomes/species. As opposed to ChIP-Enrich, Poly-Enrich works well even when nearly all genes have a peak. To illustrate this, we used Poly-Enrich to characterize the pathways and types of genic regions (introns, promoters, etc) enriched with different families of repetitive elements. By comparing ChIP-Enrich and Poly-Enrich results from ENCODE ChIP-seq data, we found that the optimal test depends more on the pathway being regulated than on the transcription factor or other properties of the dataset. Using known transcription factor functions, we discovered clusters of related biological processes consistently better modeled with either the binary score method (ChIP-Enrich) or count based method (Poly-Enrich). This suggests that the regulation of certain processes is more often modified by multiple binding events (count-based), while others tend to require only one (binary). Our new hybrid method handles this by automatically choosing the optimal method, with correct FDR-adjustment. Author SummaryAlthough every cell in our body contains the same DNA, our cells perform vastly different functions due to differences in how our genes are regulated. Certain regions of the genome are bound by DNA binding proteins (transcription factors), which regulate the expression of nearby genes. After an experiment to identify a large set of these regions, we can then model the association of these regions with various cellular pathways and biological processes. This analysis helps understand the overall biological effect that the binding events have on the cells. For example, if genes relating to apoptosis tend to have the transcription factor, Bcl-2, bind more often nearby, then Bcl-2 is likely to have a vital role in regulating apoptosis. The specifics of how to best perform this analysis is still being researched and depends on properties of the set of genomic regions. Here, we introduce a new, more flexible method that counts the number of occurrences per gene and models that in a sophisticated statistical test, and compare it to a previous method. We show that the optimal method depends on multiple factors, and the new method, Poly-Enrich, allows interesting findings in scenarios where the previous method failed.

bioinformatics↗

bioRxiv2018

IDH1R132H acts as a tumor suppressor in glioma via epigenetic upregulation of the DNA damage response

One sentence summaryMutant IDH1 acts as a tumor suppressor when co-expressed together with TP53 and ATRX inactivating mutations in glioma, inducing genomic stability, DNA repair and resistance to genotoxic therapies.\n\nAbstractGlioma patients whose tumors carry a mutation in the Isocitrate Dehydrogenase 1 (IDH1R132H) gene are younger at the time of diagnosis and survive longer. The molecular glioma subtype which we modelled, harbors IDH1R132H, tumor protein 53 (TP53) and alpha-thalassemia/mental retardation syndrome X-linked (ATRX) loss. The impact of IDH1R132H on genomic stability, DNA damage response (DDR) and DNA repair in this molecular glioma subtype is unknown. We discovered that IDH1R132H expression in the genetic context of ATRX and TP53 inactivation: (i) increases median survival (MS), (ii) enhances DDR activity via epigenetic upregulation of Ataxia-telangiectasia mutated (ATM) signaling, and (iii) elicits tumor radioresistance. Pharmacological inhibition of ATM or checkpoint kinase 1 and 2 (CHK1/2), two essential kinases in the DDR pathways, restored tumors radiosensitivity. Translation of these findings for mlDH1 glioma patients could significantly improve the therapeutic efficacy of radiotherapy, and thus have a major impact on patient survival.

cancer biology↗