Biology subjects

Zhao, C.

Publications and source records attributed to Zhao, C..

At least 289 records · Page 16Linked to original sources

bioRxiv2020

Cyclin G2 inhibits oral squamous cell carcinoma growth and metastasis by binding to insulin-like growth factor binding protein 3 and regulating the FAK-SRC-STAT signaling pathway

The cell cycle protein cyclin G2 is considered a tumor suppressor. However, its regulatory effects and potential mechanisms in oral cancers are not well understood. This study aimed to investigate the effect of cyclin G2 on oral squamous cell carcinoma (OSCC). The data from 80 patients with OSCC were utilized to predict the abnormal expression of cyclin G2. The proliferation and metastasis were determined by a cell counting Kit-8 assay, flow cytometry, a wound-healing assay and a cell invasion assay. The expression of key proteins and genes associated with the cyclin G2 signaling pathways was determined by western blotting and real-time PCR, respectively. The orthotopic nude mice model was established by a mouth injection of SCC9 cells overexpressing cyclin G2. We showed that the low level of cyclin G2 in OSCC, which is negatively correlated with clinical staging, was a negative prognostic factor for the disease. We also found that cyclin G2 inhibited the proliferation, metastasis and blocked the cell cycle at G1/S of OSCC cells, suggesting that cyclin G2 has an inhibitory effect in OSCC. Mechanistically, cyclin G2 inhibited the growth and metastasis of OSCC by binding to insulin-like growth factor binding protein 3 (IGFBP3) and regulating the focal adhesion kinase (FAK) -SRC-STAT signal transduction pathway. Cyclin G2 competed with integrin to bind to IGFBP3; the binding between integrin and IGFBP3 was reduced after cyclin G2 overexpression, thereby inhibiting the phosphorylation of FAK and SRC. These results showed that cyclin G2 inhibited the progression of OSCC by interacting with IGFBP3 and that it may be a new target for OSCC treatment.

cancer biology↗

bioRxiv2020

CK2 inhibits TIMELESS nuclear export and modulates CLOCK transcriptional activity to regulate circadian rhythms

Circadian clocks orchestrate daily rhythms in organismal physiology and behavior to promote optimal performance and fitness. In Drosophila, key pacemaker proteins PERIOD (PER) and TIMELESS (TIM) are progressively phosphorylated to perform phase-specific functions. Whereas PER phosphorylation has been extensively studied, systematic analysis of site-specific TIM phosphorylation is lacking. Here, we identified phosphorylation sites of PER-bound TIM by mass spectrometry, given the importance of TIM as a modulator of PER function in the oscillator. Among the twelve TIM phosphorylation sites we identified, at least two of them are critical for circadian timekeeping as mutants expressing non-phosphorylatable mutations exhibit altered behavioral rhythms. In particular, we observed that CK2-dependent phosphorylation of TIM(S1404) promotes nuclear accumulation of PER-TIM heterodimers by inhibiting the interaction of TIM and nuclear export component, Exportin 1 (XPO1). We postulate that proper level of nuclear PER-TIM accumulation is necessary to facilitate kinase recruitment for the regulation of daily phosphorylation rhythm and phase-specific transcriptional activity of CLOCK (CLK). Our results highlight the contribution of phosphorylation-dependent nuclear export of PER-TIM heterodimers to the maintenance of circadian periodicity and identify a new mechanism by which the negative elements of the circadian oscillator (PER-TIM) regulate the positive elements (CLK-CYC). Finally, since the molecular phenotype of tim(S1404A) non-phosphorylatable mutant exhibits remarkable similarity to that of a mutation in human timeless that underlies Familial Advanced Sleep Phase Syndrome (FASPS), our results revealed an unexpected parallel between the functions of Drosophila and human TIM and may provide new insights into the molecular mechanisms underlying human FASPS. Significance StatementPhosphorylation is a central mechanism important for the regulation of circadian physiology across organisms. The molecular oscillator is composed of pacemaker proteins that undergo elaborate phosphorylation programs to regulate phase-specific functions. In Drosophila, phosphorylation of TIMELESS (TIM) has been recognized as critical for its function in the oscillator, but a systematic analysis of TIM phosphorylation is lacking. Here, we identified twelve Drosophila TIM phosphorylation sites by mass spectrometry and showed that phosphorylation at TIM(S1404) is necessary for maintaining 24-hour rhythms. Finally, since the molecular phenotype of tim(S1404A) non-phosphorylatable fly mutant exhibits remarkable similarity to that of a mutation in human timeless that underlies FASPS, our results may provide new insights into the molecular underpinnings of human FASPS.

neuroscience↗

bioRxiv2020

Ultra-rapid metagenotyping of the human gut microbiome

Sequence variation is used to quantify population structure and identify genetic determinants of phenotypes that vary within species. In the human microbiome and other environments, single nucleotide polymorphisms (SNPs) are frequently detected by aligning metagenomic sequencing reads to catalogs of genes or genomes. But this requires high-performance computing and enough read coverage to distinguish SNPs from sequencing errors. We solved these problems by developing the GenoTyper for Prokaytotes (GT-Pro), a suite of novel methods to catalog SNPs from genomes and use exact k-mer matches to perform ultra-fast reference-based SNP calling from metagenomes. Compared to read alignment, GT-Pro is more accurate and two orders of magnitude faster. We discovered 104 million SNPs in 909 human gut species, characterized their global population structure, and tracked pathogenic strains. GT-Pro democratizes strain-level microbiome analysis by making it possible to genotype hundreds of metagenomes on a personal computer. Software availabilityGT-Pro is available at https://github.com/zjshi/gt-pro.

bioinformatics↗

bioRxiv2020

Structural Basis for Negative Regulation of ABA Signaling by ROP11 GTPase

Abscisic acid (ABA) is an essential plant hormone responsible for plant development and stress responses. Recent structural and biochemical studies have identified the key components involved in ABA signaling cascade, including PYR/PYL/RCAR receptors, protein phosphatases PP2C, and protein kinases SnRK2. The plant-specific, Roh-like (ROPs) small GTPases are negative regulators of ABA signal transduction by interacting with PP2C, which can shut off "leaky" ABA signal transduction caused by constitutive activity of monomeric PYR/PYL/RCAR receptors. However, the structural basis for negative regulation of ABA signaling by ROP GTPases remains elusive. In this study, we have utilized large-scale coarse-grained (10.05 milliseconds) and allatom molecular dynamics simulations and standard protein-protein binding free energy calculations to predict the complex structure of AtROP11 and phosphatase AtABI1. In addition, we have predicted the detailed complex association pathway and identified the critical residue pairs in AtROP11 and AtABI1 for complex stability. Overall, this study has established a powerful framework of using large-scale molecular simulations to predict unknown protein complex structures and suggested the molecular mechanism of the negative regulation of ABA signal transduction by small GTPases.

plant biology↗

bioRxiv2020

Human IgG cell neutralizing monoclonal antibodies block SARS-CoV-2 infection

The coronavirus induced disease 19 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a worldwide threat to human lives, and neutralizing antibodies present a great therapeutic potential in curing affected patients. We purified more than one thousand memory B cells specific to SARS-CoV-2 S1 or RBD (receptor binding domain) antigens from 11 convalescent COVID-19 patients, and a total of 729 naturally paired heavy and light chain fragments were obtained by single B cell cloning technology. Among these, 178 recombinant monoclonal antibodies were tested positive for antigen binding, and the top 13 binders with Kd below 0.5 nM are all RBD binders. Importantly, all these 13 antibodies could block pseudoviral entry into HEK293T cells overexpressing ACE2, with the best ones showing IC50s around 2-3 nM. We further identified 8 neutralizing antibodies against authentic virus with IC50s within 10 nM. Among these, 414-1 blocked authentic viral entry at IC50 of 1.75 nM and in combination with 105-38 could achieve IC50 as low as 0.45 nM. Meanwhile, we also found that 3 antibodies could cross-react with the SARS-CoV spike protein. Altogether, our study provided a panel of potent human neutralizing antibodies for COVID19 as therapeutics candidates for further development.

immunology↗

bioRxiv2020

Mechanical Tension in Syndecan-1 is Regulated by Extracellular Mechanical Cues and Fluidic Shear Stress

The endothelium plays a central role in regulating vascular homeostasis and is key in determining the response to materials implanted in the vascular system. Endothelial cells are uniquely sensitive to biophysical cues from applied forces and their local cellular microenvironment. The glycocalyx is a layer of proteoglycans, glycoproteins and glycosaminoglycans that lines the luminal surface of the vascular endothelium, interacting directly with the components of the blood and the forces of blood flow. In this work, we examined the changes in mechanical tension of syndecan-1, a cell surface proteoglycan that is an integral part of the glycocalyx, in response to substrate stiffness and fluidic shear stress. Our studies demonstrate that syndecan-1 is mechanically responsive to extracellular mechanical cues and alters its association with cytoskeletal and adhesion-related proteins in response to substrate stiffness and physiological flow.

bioengineering↗

bioRxiv2020

Genome-wide identification and transcriptional analyses of MATE transporter genes in root tips of wild Cicer spp. under aluminium stress

Chickpea is an economically important legume crop with high nutritional value in human diets. Aluminium-toxicity poses a significant challenge for the yield improvement of this increasingly popular crop in acidic soils. The wild progenitors of chickpea may provide a more diverse gene pool for Al-tolerance in chickpea breeding. However, the genetic basis of Al-tolerance in chickpea and its wild relatives remains largely unknown. Here, we assessed the Al-tolerance of six selected wild Cicer accessions by measuring the root elongation in solution culture under control (0 {micro}M Al3+) and Al-treatment (30 {micro}M Al3+) conditions. Al-treatment significantly reduced the root elongation in all target lines compared to the control condition after 2-days growth. However, the relative reduction of root elongation in different lines varied greatly: 3 lines still retained significant root growth under Al-treatment, whilst another 2 lines displayed no root growth at all. We performed genome-wide identification of multidrug and toxic compound extrusion (MATE) encoding genes in the Cicer genome. A total of 56 annotated MATE genes were identified, which divided into 4 major phylogeny groups (G1-4). Four homologues to lupin LaMATE (> 50% aa identity; named CaMATE1-4) were clustered with previously characterised MATEs related to Al-tolerance in various other plants. qRT-PCR showed that CaMATE2 transcription in root tips was significantly up-regulated upon Al-treatment in all target lines, whilst CaMATE1 was up-regulated in all lines except Bari2_074 and Deste_064, which coincided with the lines displaying no root growth under Al-treatment. Transcriptional profiling in five Cicer tissues revealed that CaMATE1 is specifically transcribed in the root tissue, further supporting its role in Al-detoxification in roots. This first identification of MATE-encoding genes associated with Al-tolerance in Cicer paves the ways for future functional characterization of MATE genes in Cicer spp., and to facilitate future design of gene-specific markers for Al-tolerant line selection in chickpea breeding programs.

plant biology↗

bioRxiv2020

Parkinson's disease-related phosphorylation at Tyr39 rearranges α-synuclein amyloid fibril structure revealed by cryo-EM

Post-translational modifications (PTMs) of -synuclein (-syn), e.g. phosphorylation, play an important role in modulating -syn pathology in Parkinsons disease (PD) and -synucleinopathies. Accumulation of phosphorylated -syn fibrils in Lewy bodies and Lewy neurites is the histological hallmark of these diseases. However, it is unclear how phosphorylation relates to -syn pathology. Here, by combining chemical synthesis and bacterial expression, we obtained homogeneous -syn fibrils with site-specific phosphorylation at Y39, which exhibits enhanced neuronal pathology in rat primary cortical neurons. We determined the cryo-EM structure of pY39 -syn fibril, which reveals a new fold of -syn with pY39 in the center of the fibril core forming electrostatic interaction network with eight charged residues in the N-terminal region of -syn. This structure composed of residues 1-100 represents the largest -syn fibril core determined so far. This work provides structural understanding on the pathology of pY39 -syn fibril, and highlights the importance of PTMs in defining the polymorphism and pathology of amyloid fibrils in neurodegenerative diseases.

biophysics↗

bioRxiv2020

Layer-specific Loss and Compensation of Parvocellular Response in Subcortical Pathways of Adult Human Amblyopia

Abnormal visual experience in critical period causes amblyopia or lazy eye, reducing visual abilities even with corrected optics. A long-standing question is where in the human visual system does the amblyopic deficit arise. In particular, whether amblyopia induces selective deficits of the magnocellular (M) or the parvocellular (P) geniculostriate pathways, and whether the more ancient retinotectal pathway is also affected. Technical limitations to non-invasively measure layer-specific activity in human lateral geniculate nucleus (LGN) and superior colliculus (SC) hampered efforts in addressing these questions. In the current study, using lamina-resolved 3T and 7T fMRI and visual stimuli selectively activating the M and P pathways, we investigated layer-specific response properties of the LGN and the SC of amblyopia patients and normal controls. With stimuli presented to the amblyopic eye, there was a stronger response loss in the P layers than in the M layers of the LGN. Compared to normal controls, amblyopic eyes response to the P stimulus was selectively reduced in the superficial SC, while the fellow eyes response was robustly increased in the deep SC. Selective P response deficits of amblyopia were also observed in the visual pulvinar, early visual cortex, and ventral but not dorsal visual streams. These results provide strong in vivo evidence in adult amblyopic patients for selective deficits of parvocellular functions in the visual thalamus, and additionally reveal response deficits to the amblyopic eye and neural compensation to the fellow eye in the retinotectal pathway. HighlightsO_LIParvocellular response loss in the LGN P layers, visual pulvinar and ventral visual stream C_LIO_LISelective amblyopic deficits of the parvocellular pathway C_LIO_LIAmblyopic eyes response decreased in the superficial SC C_LIO_LIFellow eyes response increased in the deep SC C_LIO_LIAmblyopic deficits and neural compensation in the retinotectal pathway C_LI

neuroscience↗

bioRxiv2020

Elucidating Direct Kinase Targets of Compound Danshen Dropping Pills Employing Archived Data and Prediction Models

The research on the direct target of traditional Chinese medicine (TCM) is the key to study the mechanism and material basis of TCM, but there is still no effective technical methods at present. For Compound Danshen dropping pills (CDDP), there is no report about its direct targets. In this study, the direct targets of CDDP were studied for the first time, especially focusing on the protein kinase family, which plays causal roles in a variety of human disease. Firstly, the literature database of CDDP was constructed by literature retrieval, and the important components contained in CDDP were extracted. Secondly, the potential direct targets of important components was obtained through querying public database and predicted by Multi-voting SEA algorithm. Then, the KinomeX system was used to predict and to filter the potential kinase targets of CDDP. Finally, the experimental verification was carried out. In total, 30 active kinase targets was obtained at 25 g/ml concentration of CDDP, and 9 dose-dependent targets were obtained at 250 g/ml concentration of CDDP. This is an efficient and accurate strategy by integrating the targets recorded in several public databases and the targets calculated by two in silico modelling approaches predict potential direct targets of TCM, which can lay an important foundation for the study of the mechanism and material basis of them, promoting the modernization of TCM.

pharmacology and toxicology↗

bioRxiv2020

Aberrant pathogenic GM-CSF+ T cells and inflammatory CD14+CD16+ monocytes in severe pulmonary syndrome patients of a new coronavirus

Pathogenic human coronavirus infections, such as severe acute respiratory syndrome CoV (SARS-CoV) and Middle East respiratory syndrome CoV (MERS-CoV), cause high morbidity and mortality 1,2. Recently, a severe pneumonia-associated respiratory syndrome caused by a new coronavirus was reported at December 2019 (2019-nCoV) in the city Wuhan, Hubei province, China3-5, which was also named as pneumonia-associated respiratory syndrome (PARS)6. Up to 9th of February 2020, at least 37, 251 cases have been reported with 812 fatal cases according to the report from China CDC. However, the immune mechanism that potential orchestrated acute mortality from patients of 2019-nCoV is still unknown. Here we show that after the 2019-nCoV infection, CD4+T lymphocytes are rapidly activated to become pathogenic T helper (Th) 1 cells and generate GM-CSF etc. The cytokines environment induces inflammatory CD14+CD16+ monocytes with high expression of IL-6 and accelerates the inflammation. These aberrant and excessive immune cells may enter the pulmonary circulation in huge numbers and play an immune damaging role to causing lung functional disability and quick mortality. Our results demonstrate that excessive non-effective host immune responses by pathogenic T cells and inflammatory monocytes may associate with severe lung pathology. Therefore, we suggest that monoclonal antibody that targets the GM-CSF or interleukin 6 receptor may potentially curb immunopathology caused by 2019-nCoV and consequently win more time for virus clearance.

immunology↗

bioRxiv2020

Tracing the origin of a new organ by inferring the genetic basis of rumen evolution

The rumen is the hallmark organ of ruminants and hosts a diverse ecosystem of microorganisms that facilitates efficient digestion of plant fibers. We used 897 transcriptomes from three Cetartiodactyla lineages: ruminants, camels and cetaceans, as well as data from ruminant comparative genomics and functional assays to explore the genetic basis of rumen origin and evolution. Comparative analyses reveal that the rumen and the first-chamber stomachs of camels and cetaceans shared a common tissue origin from the esophagus. The rumen recruited genes from other tissues/organs and up-regulated many esophagus genes to aquire functional innovations involving epithelium absorption, improvement of the ketone body metabolism and regulation of microbial community. These innovations involve such genetic changes as ruminant-specific conserved elements, newly evolved genes and positively selected genes. Our in vitro experiements validate the functions of one enhancer, one positively selected gene and two newly evolved antibacterial genes. Our study provides novel insights into the origin and evolution of a complex organ.

evolutionary biology↗

bioRxiv2020

Anethole dithiolethione improves liver fatty acid metabolism in hamster fed high-fat diets

"Lipotoxicity" induced by excessive accumulation of free fatty acids (FFAs) in the liver, especially saturated FAs and their toxic metabolites, is closely related to metabolic diseases such as nonalcoholic fatty liver disease (NAFLD). Hydrogen sulfide (H2S), a novel gaseous signaling molecule, has been reported to have lipid-lowering effects, but its effect on FAs metabolism remains unclear. The purpose of this study was to investigate the effect and mechanisms of anethole dithiolethione (ADT, a sustained-release H2S donor) on hepatic FAs metabolism. ADT was administered daily for 4 weeks in male Syrian golden hamsters fed a high fat diet (HFD), and FAs profiles of liver tissues were analyzed using GC-MS. The results showed that in HFD-fed hamsters, ADT treatment significantly reduced the accumulation of saturated and monounsaturated fatty acids (C16:0, C18:0, C16:1, and C18:1n9), while increased the content of n-6 and n-3 series polyunsaturated fatty acids (C20:3n6, C20:4n6, and C22:6n3). Mechanistically, ADT obviously inhibited the overexpression of ACC1, FAS and SCD1, and up-regulated the levels of FATPs, L-FABP, CPT1, FADS1 and FADS2. Notably, ADT evidently induced Mitofusin1 to facilitate mitochondrial fusion and optimize {beta}-oxidation. These findings suggest that ADT plays a beneficial role by regulating the synthesis, desaturation, {beta}-oxidation, uptake, binding/isolation, and transport of FAs. In conclusion, ADT is effective in improving liver FAs metabolic disorders and liver injuries caused by HFD.

pharmacology and toxicology↗

bioRxiv2019

Predictive olfactory learning in Drosophila

Olfactory learning and conditioning in the fruit fly is typically modelled by correlation-based associative synaptic plasticity. It was shown that the conditioning of an odor-evoked response by a shock depends on the connections from Kenyon cells (KC) to mushroom body output neurons (MBONs). Although on the behavioral level conditioning is recognized to be predictive, it remains unclear how MBONs form predictions of aversive or appetitive values (valences) of odors on the circuit level. We present behavioral experiments that are not well explained by associative plasticity between conditioned and unconditioned stimuli, and we suggest two alternative models for how predictions can be formed. In error-driven predictive plasticity, dopaminergic neurons (DANs) represent the error between the predictive odor value and the shock strength. In target-driven predictive plasticity, the DANs represent the target for the predictive MBON activity. Predictive plasticity in KC-to-MBON synapses can also explain trace-conditioning, the valence-dependent sign switch in plasticity, and the observed novelty-familiarity representation. The model offers a framework to dissect MBON circuits and interpret DAN activity during olfactory learning.

neuroscience↗

bioRxiv2019

Parkinson's disease associated mutation E46K of α-synuclein triggers the formation of a novel fibril structure

-Synuclein (-syn) amyloid fibril, as the major component of Lewy bodies and pathological entity spreading in human brain, is closely associated with Parkinsons disease (PD) and other synucleinopathies. Several single amino-acid mutations (e.g. E46K) of -syn have been identified causative to the early onset of familial PD. Here, we determined the cryo-EM structure of a full-length -syn fibril formed by N-terminal acetylated E46K mutant -syn (Ac-E46K). The fibril structure represents a new fold of -syn, which demonstrates that the E46K mutation breaks the electrostatic interactions in the wild type (WT) -syn fibril and thus triggers the rearrangement of the overall structure. Furthermore, we show that the Ac-E46K fibril is less resistant to harsh conditions and protease cleavage, and more prone to be fragmented with a higher capability of seeding fibril formation than that of the WT fibril. Our work provides a structural view to the severe pathology of the PD familial mutation E46K of -syn and highlights the importance of electrostatic interactions in defining the fibril polymorphs.

biophysics↗

bioRxiv2019

LMSM: a modular approach for identifying lncRNA related miRNA sponge modules in breast cancer

Until now, existing methods for identifying lncRNA related miRNA sponge modules mainly rely on lncRNA related miRNA sponge interaction networks, which may not provide a full picture of miRNA sponging activities in biological conditions. Hence there is a strong need of new computational methods to identify lncRNA related miRNA sponge modules. In this work, we propose a framework, LMSM, to identify LncRNA related MiRNA Sponge Modules from heterogeneous data. To understand the miRNA sponging activities in biological conditions, LMSM uses gene expression data to evaluate the influence of the shared miRNAs on the clustered sponge lncRNAs and mRNAs. We have applied LMSM to the human breast cancer (BRCA) dataset from The Cancer Genome Atlas (TCGA). As a result, we have found that the majority of LMSM modules are significantly implicated in BRCA and most of them are BRCA subtype-specific. Most of the mediating miRNAs act as crosslinks across different LMSM modules, and all of LMSM modules are statistically significant. Multi-label classification analysis shows that the performance of LMSM modules is significantly higher than baselines performance, indicating the biological meanings of LMSM modules in classifying BRCA subtypes. The consistent results suggest that LMSM is robust in identifying lncRNA related miRNA sponge modules. Moreover, LMSM can be used to predict miRNA targets. Finally, LMSM outperforms a graph clustering-based strategy in identifying BRCA-related modules. Altogether, our study shows that LMSM is a promising method to investigate modular regulatory mechanism of sponge lncRNAs from heterogeneous data. Author summaryPrevious studies have revealed that long non-coding RNAs (lncRNAs), as microRNA (miRNA) sponges or competing endogenous RNAs (ceRNAs), can regulate the expression levels of messenger RNAs (mRNAs) by decreasing the amount of miRNAs interacting with mRNAs. In this work, we hypothesize that the "tug-of-war" between RNA transcripts for attracting miRNAs is across groups or modules. Based on the hypothesis, we propose a framework called LMSM, to identify LncRNA related MiRNA Sponge Modules. Based on the two miRNA sponge modular competition principles, significant sharing of miRNAs and high canonical correlation between the sponge lncRNAs and mRNAs, LMSM is also capable of predicting miRNA targets. LMSM not only extends the ceRNA hypothesis, but also provides a novel way to investigate the biological functions and modular mechanism of lncRNAs in breast cancer.

bioinformatics↗

bioRxiv2019

CRISPR screening of porcine sgRNA library identified host factors essential for Japanese encephalitis virus replication

Japanese encephalitis virus (JEV) is a mosquito-borne zoonotic flavivirus that causes encephalitis and reproductive disorders in mammalian species. However, key host genes involved in the JEV life cycle and cell death are poorly understood. Here, we designed 85,674 single guide RNAs (sgRNAs) targeting 17,743 protein-coding genes, 11,053 long ncRNAs (lncRNAs), and 551 microRNAs (miRNAs) in the porcine genome, and subsequently developed a porcine sgRNA library and genome-scale CRISPR/Cas9 knockout (PigGeCKO) system. These sgRNAs were delivered into porcine kidney-15 (PK-15) cells that constitutively express Cas9, positive selection screening of the resulting PigGeCKO cell collection for resistance to JEV-induced cell death led to the identification of several previously unreported genes required for JEV infection. We conducted follow-up studies to verify the dependency of JEV on these genes, and identified functional contributions for six of the many candidate JEV-related host genes, including an endoplasmic reticulum membrane protein complex subunit 3 (EMC3) and calreticulin (CALR). Additionally, we identified that four genes associated with heparan sulfate proteoglycans (HSPGs) metabolism, specifically those responsible for HSPG sulfurylation, facilitated JEV entry into PK-15 cells. Thus, beyond our development of the largest CRISPR-based functional genomic screening platform for pig research to date, this work deepens our basic understanding of flavivirus infection and identifies multiple potentially vulnerable targets for the development of medical and breeding technologies to prevent and treat diseases caused by Japanese encephalitis virus.

genetics↗

bioRxiv2019

Torsin ATPases are required to complete nuclear pore complex biogenesis in interphase

Nuclear envelope herniations (blebs) containing FG-nucleoporins and ubiquitin are the phenotypic hallmark of Torsin ATPase manipulation. Both the dynamics of blebbing and the connection to nuclear pore biogenesis remain poorly understood. We employ a proteomics-based approach to identify MLF2 as a luminal component of the bleb. Using an MLF2-based live cell imaging platform, we demonstrate that NE blebbing occurs rapidly and synchronously immediately after nuclear envelope reformation during mitosis. Bleb formation is independent of ubiquitin conjugation within the bleb, but strictly dependent on POM121, a transmembrane nucleoporin essential for interphase nuclear pore biogenesis. Nup358, a late marker for interphase nuclear pore complex (NPC) biogenesis, is underrepresented relative to FG nucleoporins in nuclear envelopes of Torsin-deficient cells. The kinetics of bleb formation, its dependence on POM121, and a reduction of mature NPCs in Torsin deficient cells lead us to conclude that the hallmark phenotype of Torsin manipulation represents the accumulation of stalled NPC assembly intermediates.

cell biology↗