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Dicoumarol Alters cellular Redox State and Inhibits Nuclear Factor Kappa B to Enhance Arsenic Trioxide-Induced Apoptosis

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Acta Biochimica et Biophysica Sinica 2004年第3期36卷 235-242页

作者： Ya-Wu JING, Jing YI, Yu-Ying CHEN, Qing-Shen HU, Gui-Ying SHI, Hue LI, and Xue-Ming TANG* Department of cell biology, Shanghai Second Medical University, Shanghai 200025, China Department of cell biology Shanghai Second Medical University

The effects of a number of cytotoxic drugs are influenced by cellular reduction/oxidation (redox) state. In the present study, we attempt to explore if dicoumarol, an inhibitor of NADPH: quinone oxidoreductase (NQO1), alters the cellular redox state and how this alteration affects the redox-related apoptosis. Flow cytometry was used to assess the reactive oxygen species (ROS) level and apoptotic rates of HeLa cells treated with arsenic trioxide (As2O3) alone or in combination with natural anthraquinone emodin and dicoumarol or plus N-acetyl-cysteine. Western blot, immunofluorescence, electrophoretic mobility shift assay and luciferase assay were used to detect Nuclear Factor kappa B (NF-κB) activation. The results showed that dicoumarol synergized with emodin to sensitize HeLa cells to As2O3-induced apoptosis through raising the ROS level. More notably, this enhanced susceptibility was associated with a ROS-mediated inhi- bition of NF-κB activation in which the combinative treatment with dicoumarol prevented NF-κB from binding to target DNA. It was suggested that dicoumarol in combination with anthraquinones might be a novel strategy to expand the chemotherapeutic spectrum of As2O3 by means of interfering the cellular redox state.

关键词： dicoumarol arsenic trioxide reduction/ oxidation state apoptosis NF-κB

SPORTS1.0: A Tool for Annotating and Profiling Non-coding RNAs Optimized for rRNA-and tRNA-derived Small RNAs

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Genomics, Proteomics & Bioinformatics 2018年第2期16卷 144-151页

作者： Junchao Shi Eun-A Ko Kenton M.Sanders Qi Chen Tong Zhou Department of Physiology and cell biology University of Nevada Reno School of Medicine

High-throughput RNAoseq has revolutionized the process of small RNA （sRNA） discovery, leading to a rapid expansion of sRNA categories. In addition to the previously wellcharacterized sRNAs such as microRNAs （miRNAs）, piwi-interacting RNAs （piRNAs）, and small nucleolar RNA （snoRNAs）, recent emerging studies have spotlighted on tRNA-derived sRNAs （tsRNAs） and rRNA-derived sRNAs （rsRNAs） as new categories of sRNAs that bear versatile functions. Since existing software and pipelines for sRNA annotation are mostly focused on analyzing miRNAs or piRNAs, here we developed the sRNA annotation pipeline _optimized for rRNA- and tRNA-derived s_RNAs （SPORTS 1 .0）. SPORTS1.0 is optimized for analyzing tsRNAs and rsRNAs from sRNA-seq data, in addition to its capacity to annotate canonical sRNAs such as miRNAs and piRNAs. Moreover, SPORTS1.0 can predict potential RNA modification sites based on nucleotide mismatches within sRNAs. SPORTS1.0 is precompiled to annotate sRNAs for a wide range of 68 species across bacteria, yeast, plant, and animal kingdoms, while additional species for analyses could be readily expanded upon end users＇ input. For demonstration, by analyzing sRNA datasets using SPORTS1.0, we reveal that distinct signatures are present in tsRNAs and rsRNAs from different mouse cell types. We also find that compared to other sRNA species, tsRNAs bear the highest mismatch rate, which is consistent with their highly modified nature. SPORTS1.0 is an opensource software and can be publically accessed at https：//***/junchaoshi/sports1.0.

关键词： Small RNA RNA-seq data analysistsRNA rsRNA Annotation pipeline

The Application of CRISPR-Cas9 Genome Editing in Caenorhabditis elegans

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Journal of Genetics and Genomics 2015年第8期42卷 413-421页

作者： Suhong Xu Division of Biological Sciences Section of Cell and Developmental Biology University of California

Genome editing using the Cas9 endonuclease of Streptococcus pyogenes has demonstrated unparalleled efficacy and facility for modifying genomes in a wide variety of organisms. Caenorhabditis elegans is one of the most convenient multicellular organisms for genetic analysis, and the application of this novel genome editing technique to this organism promises to revolutionize analysis of gene function in the future. CRISPR-Cas9 has been successfully used to generate imprecise insertions and deletions via non-homologous end-joining mechanisms and to create precise mutations by homology-directed repair from donor templates. Key variables are the methods used to deliver the Cas9 endonuclease and the efficiency of the single guide RNAs. CRISPR-Cas9-mediated editing appears to be highly specific in C. elegans, with no reported off-target effects. In this review, 1 briefly summarize recent progress in CRISPR-Cas9-based genome editing in C. elegans, highlighting technical improvements in mutagenesis and mutation detection, and discuss potential future appli- cations of this technique.

关键词： Genome editing CRISPR: Cas9 Non-homologous end-joining (NHEJ) Homology-directed repair (HDR) Somatic mutation C. elegans

Retrovirus-mediated herpes simplex virus thymidine kinase gene therapy approach for hepatocellular carcinoma

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cell Research 1999年第3期9卷 225-235页

作者： GAO DING CHENG WEI AN JIE DAI(Dcpartment of cell biology, Capital University of MedicalSciences 100054 Beijing, China) 1Department of cell biology Capital University of Medical Sciences Beijing 100054 China 1Department of cell biology Capital University of Medical Sciences Beijing 100054 China 1Department of cell biology Capital University of Medical Sciences Beijing 100054 China

The therapeutic effect of herpes simplex virus thymi-dine kinase/ganciclovir (HSV-tk/GCV) system on hepa-tocellular carcinoma was studied in this experimeflt. Thetk-containing retroviral recombinants were used to infecthepatoma cells (BEL-7402) and the cells were treated withganciclovir (0-1000 pg/ml). The results showed that HSV-tk gene could be efficielltly transferred in Vitro into hep-atoma cells and stably expressed. The growth potentialof the tk-containing cells was significantly inhibited byGCV (Pcellular carcinoma was still significantIy gener-ated. The proper mechanism of HSV-tk gene therapy onhepatic tumor referred as "bystander effect" in therapeu-tic approach has not been found in this study and requiredto be explored further.

关键词： HSV-tk retrovirus gene therapy hepatocellular carcinoma

Introduction of Foreign Genes into Silkworm Eggs by Electroporation and Its Application in Transgenic Vector Test

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Acta Biochimica et Biophysica Sinica 2004年第5期36卷 323-330页

作者： Xiu-Yang GUO#, Liang DONG#, Sheng-Peng WANG, Ting-Qing GUO, Jian-Yang WANG, and Chang-De LU* Institute of Biochemistry and cell biology, Shanghai Institutes for Biological Sciences, the Chinese Academy of Sciences, 320 Yueyang Road, Shanghai 200031, China Institute of Biochemistry and cell biology Shanghai Institutes for Biological Sciences the Chinese Academy of Sciences

Electroporation as a methodology to introduce foreign genes into silkworm eggs was system- atically analyzed. The foreign gene in both the newly hatched and 3rd instar larva DNA can be detected by PCR. The amount of foreign gene in 3rd instar larva DNA was about 1/1000 of that in newly hatched larva DNA. The ratio of foreign gene entering into silkworm eggs was voltage dependent and showed significant difference between the tested silkworm strains. When the piggyBac transposon system was applied, the effect of nuclear localization signal (NLS) peptide and the in vitro transcribed transposase mRNA on the transposi- tion rate has been measured. Results showed that the in vitro transcribed transposase mRNA facilitated trans- position to take place earlier and NLS could result in higher transposition probability and earlier transposition as well. When linearized vectors containing varied length of flanking homologous sequences around a re- porter gene were introduced into silkworm eggs by electroporation, the one with 2.6 kb total arm length gave higher G1 positive ratio than that with total arm length of 1.5 kb and 800 bp.

关键词： silkworm eggs gene introduction electroporation transposon homologous sequences

Microtubule-bundling protein Spef1 enables mammalian ciliary central apparatus formation

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Journal of Molecular cell biology 2019年第1期11卷 67-77页

作者： Jianqun Zheng Hao Liu Lei Zhu Yawen Chen Huijie Zhao Wei Zhang Fan Li Lele Xie Xiumin Yan Xueliang Zhu State Key Laboratory of cell biology CAS Center for Excellence in Molecular Cell ScienceShanghai Institute of Biochemistry and Cell BiologyChinese Academy of SciencesShanghai 200031China University of Chinese Academy of Sciences Shanghai 200031China

Cilia are cellular protrusions containing nine microtubule(MT)doublets and function to propel cell movement or extracellular liquid flow through beating or sense environmental stimuli through signal *** require the central pair(CP)apparatus,consisting of two CP MTs covered with projections of CP proteins,for planar *** the CP MTs of such‘9+2’cilia are constructed,however,remains *** we identify Spef1,an evolutionarily conserved microtubule-bundling protein,as a core CP MT regulator in mammalian ***1 was selectively expressed in mammalian cells with 9+2 cilia and specifically localized along the *** depletion in multiciliated mouse ependymal cells by RNAi completely abolished the CP MTs and markedly attenuated ciliary localizations of CP proteins such as Hydin and Spag6,resulting in rotational beat of the ependymal ***1,which binds to MTs through its N-terminal *** domain,formed homodimers through its C-terminal coiled coil region to bundle and stabilize *** of either the MT-binding or the dimerization activity abolished the ability of exogenous Spef1 to restore the structure and functions of the CP *** propose that Spefl bundles and stabilizes central MTs to enable the assembly and functions of the CP apparatus.

关键词： central pair apparatus cilium microtubule mouse ependymal cell mouse tracheal epithelial cell Spef1

cDNA cloning, functional expression and cellular localization of rat liver mitochondrial electron-transfer flavoprotein-ubiquinone oxidoreductase protein

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Science China(Life Sciences) 2005年第4期48卷 357-367页

作者： HUANG Shengbing SONG Wei LIN Qishui Key Laboratory of Molecular cell biology Institute of Biochemistry and Cell BiologyShanghai Institutes for Biological SciencesChinese Academy of SciencesShanghai 200031China

A membrane-bound protein was purified from rat liver *** being di-gested with V8 protease,two peptides containing identical 14 amino acid residue sequences were *** the 14 amino acid peptide derived DNA sequence as gene specific primer,the cDNA of correspondent gene 5′-terminal and 3′-terminal were obtained by RACE *** full-length cDNA that encoded a protein of 616 amino acids was thus cloned,which included the above mentioned peptide *** full length cDNA was highly homologous to that of human ETF-QO,indicating that it may be the cDNA of rat ***-QO is an iron sulfur protein located in mitochondria inner membrane containing two kinds of redox center:FAD and[4Fe-4S]*** comparing the sequence from the cDNA of the 616 amino acids protein with that of the mature protein of rat liver mitochondria,it was found that the N terminal 32 amino acid residues did not exist in the mature protein,indicating that the cDNA was that of *** the cDNA was expressed in Saccharomyces cerevisiae with inducible vectors,the protein product was enriched in mitochondrial fraction and exhibited electron transfer activity(NBT re-ductase activity)of *** demonstrated that the 32 amino acid peptide was a mito-chondrial targeting peptide,and both FAD and iron-sulfur cluster were inserted properly into the expressed ***-QO had a high level expression in rat heart,liver and *** fu-sion protein of GFP-ETF-QO co-localized with mitochondria in COS-7 cells.

关键词： electron transfer flavoprotein ubiquinone oxidoreductase precursor rat protein purification gene clone tissuespecificity.

Vertebrate Dynein-f depends on Wdr78 for axonemal localization and is essential for ciliary beat

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Journal of Molecular cell biology 2019年第5期11卷 383-394页

作者： Yirong Zhang Yawen Chen Jianqun Zheng Juan Wang Shichao Duan Wei Zhang Xiumin Yan Xueliang Zhu State Key Laboratory of cell biology CAS Center for Excellence in Molecular Cell Science Shanghai Institute of Biochemistry and Cell Biology University of Chinese Academy of Sciences Chinese Academy of Sciences 320 Yueyang Road Shanghai 200031 China

Motile cilia and flagella are microtubule-based organelles important for cell locomotion and extracellular liquid flow through beating. Although axonenal dyneins that drive ciliary beat have been extensively studied in unicellular Chlamydomonas, to what extent such knowledge can be applied to vertebrate is poorly known. In Chlamydomonas, Dynein-f controls flagellar waveforms but is dispensable for beating. The flagellar assembly of its heavy chains (HCs) requires its intermediate chain (IC) IC140 but not IC138. Here we show that, unlike its Chlamydomonas counte『part, vertebrate Dynein-f is essential for ciliary beat. We confirmed that Wdr78 is the vertebrate orthologue of IC3 Wdr78 associated with Dynein-f subunits such as Dnah2 (a HC) and Wdr63 (IC140 orthologue). It was expressed as a motile cilium-specific protein in mammalian cells. Depletion of Wdr78 or Dnah2 by RNAi paralyzed mouse ependymal cilia. Zebrafish Wdr78 morphants displayed ciliopathy-related phenotypes, such as curved bodies, hydrocephalus, abnormal otolith, randomized left-right asymmetry, and pronephric cysts, accompanied with paralyzed pronephric cilia. Furthermore, all the HCs and ICs of Dynein-f failed to localize in the Wdr78-depleted mouse ependymal cilia. Therefore, both the functions and subunit dependency of Dynein-fare altered in evolution, probably to comply with ciliary roles in higher organisms.

关键词： axonemal dynein Dynein-f IC138 motile cilium primary ciliary dyskinesia Wdr78 Dnah2

Nanotechnology combining photoacoustic kinetics and chemical kinetics for thrombosis diagnosis and treatment

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Chinese Chemical Letters 2021年第12期32卷 3665-3674页

作者： Hao Tian Lin Lin Zhaojing Ba Fangchao Xue Yanzhao Li Wen Zeng Department of cell biology Army Medical UniversityChongqing 400038China Department of Anatomy Army Medical UniversityChongqing 400038China departments of Neurology Southwest HospitalArmy Medical UniversityChongqing 400038China

Thrombotic disease is a major problem that endangers human health. At present, MRI and CT are commonly used clinically to diagnose thrombosis, and thrombolytic drugs are used for treatment), but the diagnosis time is lagging, the utilization of drugs is low, and the resulting systemic toxicity problems such as side effects lead to poor treatment effects. Nanotechnology combining photoacoustic dynamics and chemical dynamics has shown great application value in tumor targeting, diagnosis, detection and treatment. It has also become a new direction in the diagnosis and treatment of thrombotic diseases, and has created new applications in the field of nanomaterials. This review summarizes the new progress of this combination in the diagnosis and treatment of thrombotic diseases according to the differences in the construction of the nanotherapy system, at the same time, we put forward some new problems and prospects for the integration of thrombosis diagnosis and treatment.

关键词： Nanotechnology Photoacoustic kinetics Chemical kinetics Drug delivery and release Thrombosis disease