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Sumoylation of Human Parainfluenza Virus Type 3 phosphoprotein Correlates with A Reduction in Viral Replication

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Virologica Sinica 2021年第3期36卷 438-448页

作者： Qi Cheng Wenjing Huai Xiaoyan Wu Mingzhou Chen State Key Laboratory of Virology Modem Virology Research CenterCollege of Life SciencesWuhan UniversityWuhan 430072China

Human parainfluenza virus type 3(HPIV3), a member of the Paramyxoviridae family, can cause lower respiratory disease in infants and young children. The phosphoprotein(P) of HPIV3 is an essential cofactor of the viral RNA-dependent RNA polymerase large protein(L). P connects nucleocapsid protein(N) with L to initiate genome transcription and replication.Sumoylation influences many important pathways of the target proteins, and many viral proteins are also themselves sumoylated. In this study, we found that the P of HPIV3 could be sumoylated, and mutation of K492 and K532 to arginine(PK492 R/K532 R) failed to be sumoylated within P, which enhances HPIV3 minigenome activity. Biochemical studies showed that PK492 R/K532 Rhad no effect on its interactions with N, formation of homo-tetramers and formation of inclusion bodies.Finally, we found that incorporation of K492 R/K532 R into a recombinant HPIV3(rHPIV3-PK492 R/K532 R) increased viral production in culture cells, suggesting that sumoylation attenuates functions of P and down-regulates viral replication.

关键词： Human parainfluenza virus type 3(HPIV3) phosphoprotein Sumoylation Replication Viral replication

Host protein ABCE1 interacts with the viral phosphoprotein and promotes rabies virus replication

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Biosafety and Health 2020年第3期2卷 157-163页

作者： Xing Liu Jiwen Zhang Fang Li Yibrah Tekle Hagoss Weldu Tesfagaber Lulu Wang Zilong Wang Dongming Zhao Zhigao Bu State Key Laboratory of Veterinary Biotechnology Harbin Veterinary Research InstituteChinese Academy of Agricultural SciencesHarbin 150069China National High Containment Laboratory for Animal Diseases Control and Prevention Harbin 150069China Jiangsu Co-Innovation Center for the Prevention and Control of Important Animal Infectious Disease and Zoonosis Yangzhou UniversityYangzhou 225009China

Rabies virus(RABV)phosphoprotein(P)plays an important role in disrupting host interferon(IFN)-mediated antiviral immune response.ABCE1 is known as an RNase L inhibitor that negatively regulates the 2′,5′-oligoadenylate(2-5A)/RNase L antiviral system related to the IFN signaling pathway.In this study,we screened the host factors associated with RABV P protein,while focusing on ABCE1 because of its role in the life cycle of several viruses.Our results showed that knockout of ABCE1 in HEK293T cells inhibited RABV replication.In contrast,the overexpression of ABCE1 in HEK293T cells enhanced RABV replication.Notably,the co-immunoprecipitation assay showed that ABCE1 interacted with RABV P protein.Since ABCE1 and RABV P proteins are related to the IFN signaling pathway,we propose that the interaction of viral P protein with ABCE1 leads to the disruption of host antiviral immune response.In summary,our research showed that ABCE1 is an important host protein that interacts with viral P protein and regulates RABV replication.Moreover,the interaction between ABCE1 and RABV P protein may facilitate the viral P protein-mediated disruption of host antiviral immune response.

关键词： Rabies virus ABCE1 Replication phosphoprotein Interaction

Identification of human cytomegalovirus phosphoprotein 65 in C57BL/6 and BXSB mice as a potential trigger of systemic lupus erythematosus related serum markers

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Asian Pacific Journal of Tropical Biomedicine 2015年第2期5卷 138-145页

作者： Yuan Zhang Ting-Ting Jia Yang Pan Wen-Li Li Yu Sun Jin-Ming Li Lu-Nan Wang Laboratory Medicine Department Peking University Third Hospital National Center for Clinical Laboratories Beijing Hospital Graduate School Peking Union Medical CollegeChinese Academy of Medical Sciences Institute for Infectious Disease and Endemic Disease Control Beijing Centers for Disease Control and Prevention

Objective:To investigate the potential role of human cytomegalovirus lower matrix phosphoprotein 65(HCMV-pp65) in murine systemic lupus erythematosus(SLE).Methods:The prokaryotic plasmid pET-28b-pp65 was constructed to express the HCMVpp65 protein.BXSB mice and C57BL/6 mice were inoculated with pp65 eukarvotic plasmid pcDNA3.0-pp65 intramuscularly 5 times at 2-week intervals,and then the blood of the mice was subsequently collected via the retro-orbital vein.Indirect ELISAs were used to evaluate the concentration of anti-pp65 immunoglobulin G,anti-double-stranded DNA and antinuclear antibodies.lnterleukin-1β and tumor necrosis factor-α were also determined by competitive ELISA.At the same time,3 major SLE-related circulating microRNAs were examined by quantitative RT-PCR.Results:The early production of autoantibodies was observed in pp65-immunized male BXSB as well as C57BL/6 mice.Overexpression of interleukin-1β and tumor necrosis factor-a were detected in pp65-immunized male BXSB mice.Quantitative RT-PCR analyses showed that three SLE related microRNAs(microRNA-126,microRNA-125 a,and microRKA-146a) were dovvnrcgulatcd in peripheral blood mononuclear cells of pp65-immunizcd mice.Conclusions:Our findings indicate that HCMV-pp65 immunization strongly triggers the development and progression of" SLE-like disease in both BXSB and C57BL/6 mice,which indicates that the immune responses induced by HCMV-pp65 may be involved in the development of SLE.

关键词： Systemic lupus erythematosus Autoantibody Human cytomegalovirus Lower matrix phosphoprotein 65 Cytokine MicroRNA

Reproducible large-scale synthesis of surface silanized nanoparticles as an enabling nanoproteomics platform: Enrichment of the human heart phosphoproteome

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Nano Research 2019年第6期12卷 1473-1481页

作者： David S.Roberts Bifan Chen Timothy N.Tiambeng Zhijie Wu Ying Ge Song Jin Department of Chemistry University of Wisconsin-MadisonMadisonWisconsin 53706USA Department of Cell and Regenerative Biology University of Wisconsin-MadisonMadisonWisconsin 53705USA

A reproducible synthetic strategy was developed for facile large-scale (200 mg) synthesis of surface silanized magnetite (Fe3O4) nanoparticles (NPs) for biological applications.After further coupling a phosphate-specific affinity ligand,these functionalized magnetic NPs were used for the highly specific enrichment of phosphoproteins from a complex biological mixture.Moreover,correlating the surface silane density of the silanized magnetite NPs to their resultant enrichment performance established a simple and reliable quality assurance control to ensure reproducible synthesis of these NPs routinely in large scale and optimal phosphoprotein enrichment performance from batch-to-batch.Furthermore,by successful exploitation of a top-down phosphoproteomics strategy that integrates this high throughput nanoproteomics platform with online liquid chromatography (LC) and tandem mass spectrometry (MS/MS),we were able to specifically enrich,identify,and characterize endogenous phosphoproteins from highly complex human cardiac tissue homogenate.This nanoproteomics platform possesses a unique combination of scalability,specificity,reproducibility,and efficiency for the capture and enrichment of low abundance proteins in general,thereby enabling downstream proteomics applications.

关键词： nanoparticles nanoproteomics surface functionalization large-scale phosphoprotein enrichment top-down proteomics mass spectrometry

Magnetic Fe3O4@mTiO2-AIPA Microspheres for Separation of phosphoproteins and Non-phosphoproteins

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Journal of Wuhan University of Technology(Materials Science) 2019年第3期34卷 752-759页

作者：汤秋菡 ZHAO Rui 陆琦卿光焱 School of Chemistry Chemical Engineering and LifeScienceWuhan University of TechnologyWuhan430070China State Key Laboratory of Advanced Technology for Materials Synthesis and Processing Wuhan University of TechnologyWuhan430070China Research &Development Center Jushi Group Co.UdTongxiang314500China

A novel phosphoprotein separation material was developed, which is constructed by a magnetic mesoporous Fe3O4@TiO2 (Fe3O4@mTiO2) microsphere and a 5-aminoisophthalic acid (AIPA) monolayer that provides additional binding sites toward phosphate groups. The results of characteristic experiments demonstrated that Fe3O4@mTiO2-AIPA had good dispersability, high magnetic susceptibility, and satisfactory grafting ratio of AIPA, ascribed to the large specific surface area of the inorganic substrate. Taking advantages of these features, Fe3O4@mTiO2-AIPA was successfully utilized to separate a-casein (a typical phosphoprotein) and bovine serum albumin (BSA, a typical non-phosphoprotein) from their mixtures (molar ratio = 1:2). Through adjusting pH and polarity of solutions, the BSA and a-casein were respectively enriched in washing fraction and elution fraction. This result displays the good potential of Fe3O4@mTiO2-AIPA for application in phosphoprotein enrichment.

关键词： magnetic microsphere phosphoprotein separation a-casein

Proteomic analysis of hippocampal phosphoprotein reveals up-regulation of phosphorylated 14-3-3 theta in F344 rats exposed to 1-bromopropane

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Proteomic analysis of hippocampal phosphoprotein reveals up-...

Detection of Sugar-Regulated Gene Expression and Signaling in Suspension-Cultured Rice Cells

中国毒理学会第六届全国毒理学大会

作者： HUANG Zhen-lie Gaku ICHIHARA Sahoko ICHIHARA CHANG Jie ZHANG Ling-yi Guangdong Prevention and Treatment Center for Occupational Diseases Nagoya University Graduate School of Medicine Mie University Graduate School of Regional Innovation Studies

OBJECTIVE 1-Bromopropane(1-BP) has been introduced as an alternative toozone-depleting solvents and exhibits neurotoxicity both in experimental animals and human.To obtain further information on the underlying mechanisms of 1-BP-induced neurotoxicity in the central nerve system (CNS),a proteomic analysis was conducted to identify and characterize 1-BP-induced modification of phosphoprotein expression in the hippocampus of rats.METHODS Male F344 rats were exposed to 1-BP at 0,400,or 1000 ppm for 8 h·d for 1 week and 4 weeks by inhalation(n =9 rats of each group).Hippocampal protein extracts(n=3 rats of each group) from control and 1000 ppm in 4-week exposure were resolved by two-dimensional electrophoresis(2DE),Pro-Q Diamond phosphoprotein Gel Staining and SYPRO gel staining for qualitation of phosphoproteins.Two-dimensional difference in gel electrophoresis(2D-DIGE) was conducted to quantitatively analysis of phosphoprotein(n=6 rats in each group) and matrix-assisted laser-desorption ionization time-of-flight(MALDI-TOF) mass spectrometry (MS) was used for protein identification.Selected protein was further confirmed by Phos-tag SDS-PAGE. RESULTS 2D-DIGE revealed 28 protein spots in 1000 ppm group of 4-week 1-BP exposure with statistically significant difference(Pphosphoproteins by Pro-Q Diamond gel staining combined with SYPRO staining.MALDI-TOF-TOF/MS successfully identified 19 up-regulated spots and 7 down-regulated spots.These identified proteins participate in the biological processes of metabolic, immune,response to stimulus,transport and cellular communication,and involved in pathways including Parkinson’s disease,apoptosis signaling,FGF and EGF receptor signaling pathway,etc.Expression of phosphorylated 14-3-3 theta protein was further confirmed by Phos-tag SDS-PAGE.CONCLUSION These results help us understand the pro-apoptosis mechanism of 1 -BP-induced hippocampal response and establish pote

关键词： 1-bromopropane proteomic analysis phosphoprotein

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American Journal of Plant Sciences 2018年第6期9卷 1124-1142页

作者： Shin-Lon Ho Department of Agronomy National Chiayi University Chiayi Taiwan

To better understand the mechanism of sugar signaling in rice cell, the suspension-cultured rice cells were transferred from sucrose-containing (+S) to sucrose-free (-S) of MS culture medium, we found that ribosomal RNAs (rRNAs) were degraded progressively. This suggests that carbon, nitrogen, and phosphate were recycled in this process and the reduction in cellular rRNAs might lead to decreased translation to save energy in response to sugar starvation. Differential screening revealed that two groups of genes, sugar-starvation-repressed (SSR) and sugar-starvation-activated (SSA) genes, were regulated by sugar in an opposing manner. Northern-blot analysis showed that two major hybridization signals of 0.8 and 1.9 kb were induced strongly under sugar starvation. The two populations of genes corresponded with homologs of α-amylases (1.9 kb) and the glycine-rich proteins (GRPs) gene family (0.8 kb), and all were SSA genes. Expression of GRP genes was strongly induced in sugar-starved cells, which suggests that GRPs may help to protect cells against nutritional stress. Treatment of +S and -S cells with the protein kinase (PK) inhibitor staurosporine (St) and the serine/theronine phosphoprotein phosphatases 1 (PP1) and 2A (PP2A) inhibitor okadaic acid (OA) revealed that PP1 and PP2A (PPs) might be involved in increasing SSR gene expression in +S cells, and that activation of the majority of the SSA genes in -S cells might be due to PKs activity. These results suggested that PKs and PPs might be involved in the sugar regulation of SSR and SSA gene expression. An in-gel PK activity assay demonstrated that the activity of two classes of PKs (50 and 66 kDa) may be induced rapidly after transfer of +S cells to -S medium. Following transfer of -S cells to +S medium, a novel class of 38 kDa PK was induced rapidly and showed high activity. The 38 kDa PK might play a role in sugar sensing, and the 50 and 66 kDa PKs might play roles in signal sensing under sugar starvation in rice cells. These results provide valuable informati

关键词： Suspension-Cultured Rice Cells Glycine-Rich Proteins Sugar-Starvation Repressed Sugar-Starvation Activated Protein Kinases phosphoprotein Phosphatases