scholarly journals Replication and transcription machinery for ranaviruses: components, correlation, and functional architecture

2022 ◽  
Vol 12 (1) ◽  
Author(s):  
Fei Ke ◽  
Xue-Dong Yu ◽  
Zi-Hao Wang ◽  
Jian-Fang Gui ◽  
Qi-Ya Zhang
Keyword(s):  
Mammalian Cells ◽  
Core Protein ◽  
Nuclear Antigen ◽  
Progeny Virus ◽  
Efficient Production ◽  
Host Proteins ◽  
The Core ◽  
Transcription Machinery ◽  
Nucleocytoplasmic Large Dna Viruses ◽  
Fish Cells

Abstract Background Ranaviruses (family Iridoviridae) are promiscuous pathogens that can infect across species barriers in poikilotherms and can replicate in amphibian and fish cells and even in cultured mammalian cells. However, as nucleocytoplasmic large DNA viruses (NCLDVs), their replication and transcription mechanisms remain largely unknown. Here, we screened and uncovered the replication and transcription machinery of two ranaviruses, Andrias davidianus ranavirus (ADRV) and Rana grylio virus (RGV), by a combination of methods, including the isolation of proteins on nascent DNA, recombinant virus-based affinity, and NanoLuc complementation assay. Results The ranavirus replication and transcription machinery was deeply dissected and identified as a complicated apparatus containing at least 30 viral and 6 host proteins. The viral proteins ADRV-47L/RGV-63R (DNA polymerase, vDPOL), ADRV-23L/RGV-91R (proliferating cell nuclear antigen, vPCNA), ADRV-85L/RGV-27R (single-stranded DNA binding protein, vSSB), ADRV-88L/RGV-24R (vhelicase/primase), etc., constitute the core replisome. Specifically, the core of the transcription complex, the viral RNA polymerase, contain the host RNAPII subunits Rpb3, Rpb6, and Rpb11, which was a first report in NCLDVs. Furthermore, correlations and interactions among these factors in the machinery were described. Significantly, the replisome core protein vDPOL (ADRV-47L) can interact with numerous viral and host proteins and could act as a linker and regulation center in viral DNA replication and transcription. Thus, these results depicted an architecture for ranavirus replication and transcription. Conclusions Up to 36 components from ranavirus and their host were found to form viral replisomes and transcription complexes using a series of precise methods, which further constructed an architecture for ranavirus replication and transcription in which vDPOL was a key central factor and various components correlated and cooperated. Therefore, it provides a cornerstone for further understanding the mechanisms of the replication and transcription of ranaviruses which can ensure the efficient production of progeny virus and adaptation to cross-species infection.

scholarly journals Nuclear Localization of Japanese Encephalitis Virus Core Protein Enhances Viral Replication

2005 ◽  
Vol 79 (6) ◽  
pp. 3448-3458 ◽  
Author(s):  
Yoshio Mori ◽  
Tamaki Okabayashi ◽  
Tetsuo Yamashita ◽  
Zijiang Zhao ◽  
Takaji Wakita ◽  
...  
Keyword(s):  
Japanese Encephalitis Virus ◽  
Cell Lines ◽  
Japanese Encephalitis ◽  
Nuclear Localization ◽  
Mammalian Cells ◽  
Core Protein ◽  
Encephalitis Virus ◽  
Wild Type ◽  
The Core

ABSTRACT Japanese encephalitis virus (JEV) core protein was detected in both the nucleoli and cytoplasm of mammalian and insect cell lines infected with JEV or transfected with the expression plasmid of the core protein. Mutation analysis revealed that Gly42 and Pro43 in the core protein are essential for the nuclear and nucleolar localization. A mutant M4243 virus in which both Gly42 and Pro43 were replaced by Ala was recovered by plasmid-based reverse genetics. In C6/36 mosquito cells, the M4243 virus exhibited RNA replication and protein synthesis comparable to wild-type JEV, whereas propagation in Vero cells was impaired. The mutant core protein was detected in the cytoplasm but not in the nucleus of either C6/36 or Vero cell lines infected with the M4243 virus. The impaired propagation of M4243 in mammalian cells was recovered by the expression of wild-type core protein in trans but not by that of the mutant core protein. Although M4243 mutant virus exhibited a high level of neurovirulence comparable to wild-type JEV in spite of the approximately 100-fold-lower viral propagation after intracerebral inoculation to 3-week-old mice of strain Jcl:ICR, no virus was recovered from the brain after intraperitoneal inoculation of the mutant. These results indicate that nuclear localization of JEV core protein plays crucial roles not only in the replication in mammalian cells in vitro but also in the pathogenesis of encephalitis induced by JEV in vivo.

scholarly journals Characterization of Essential Domains and Plasticity of the Classical Swine Fever Virus Core Protein

2010 ◽  
Vol 84 (21) ◽  
pp. 11523-11531 ◽  
Author(s):  
Christiane Riedel ◽  
Benjamin Lamp ◽  
Manuela Heimann ◽  
Till Rümenapf
Keyword(s):  
Amino Acids ◽  
Classical Swine Fever Virus ◽  
Core Protein ◽  
Yellow Fluorescent Protein ◽  
Classical Swine Fever ◽  
Fever Virus ◽  
Progeny Virus ◽  
Special Importance ◽  
The Core ◽  
Rna Interaction

ABSTRACT Pestiviruses are pathogens of cloven-hoofed animals, belonging to the Flaviviridae. The pestiviral particle consists of a lipid membrane containing the three envelope glycoproteins Erns, E1, and E2 and a nucleocapsid of unknown symmetry, which is composed of the Core protein and the viral positive-sense RNA genome. The positively charged pestiviral Core protein consists of 86 to 89 amino acids. To analyze the organization of essential domains, N- and C-terminal truncations, as well as internal deletions, were introduced into the Core coding sequence in the context of an infectious cDNA clone of classical swine fever virus strain Alfort. Amino acids 179 to 180, 194 to 198, and 208 to 212 proved to be of special importance for the generation of progeny virus. The results of transcomplementation of a series of C-terminally truncated Core molecules indicate the importance of Ala255 at the C terminus. The plasticity of Core protein was examined by the construction of concatemeric arrays of Core coding regions and the insertion of up to three yellow fluorescent protein (YFP) genes between two Core genes. Even a Core fusion protein with more than 10-fold-increased molecular mass was integrated into the viral particle and supported the production of infectious progeny virus. The unexpected plasticity of Core protein brings into question the formation of a regular icosahedric particle and supports the idea of a histone-like protein-RNA interaction. All viruses with a duplicated Core gene were unstable and reverted to the wild-type sequence. Interestingly, a nonviable YFP-Core construct was rescued by a mutation within the C-terminal domain of the nonstructural protein NS3.

scholarly journals Phosphorylation-dependent Binding of Hepatitis B Virus Core Particles to the Nuclear Pore Complex

1999 ◽  
Vol 145 (1) ◽  
pp. 45-55 ◽  
Author(s):  
Michael Kann ◽  
Beate Sodeik ◽  
Angelika Vlachou ◽  
Wolfram H. Gerlich ◽  
Ari Helenius
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Mammalian Cells ◽  
Core Protein ◽  
Nuclear Pore ◽  
Nuclear Localization Signals ◽  
The Core ◽  
Core Proteins ◽  
B Virus ◽  
Core Particles

Although many viruses replicate in the nucleus, little is known about the processes involved in the nuclear import of viral genomes. We show here that in vitro generated core particles of human hepatitis B virus bind to nuclear pore complexes (NPCs) in digitonin-permeabilized mammalian cells. This only occurred if the cores contained phosphorylated core proteins. Binding was inhibited by wheat germ agglutinin, by antinuclear pore complex antibodies, and by peptides corresponding either to classical nuclear localization signals (NLS) or to COOH-terminal sequences of the core protein. Binding was dependent on the nuclear transport factors importins (karyopherins) α and β. The results suggested that phosphorylation induces exposure of NLS in the COOH-terminal portion of the core protein that allows core binding to the NPCs by the importin- (karyopherin-) mediated pathway. Thus, phosphorylation of the core protein emerged as an important step in the viral replication cycle necessary for transport of the viral genome to the nucleus.

scholarly journals RGD Tripeptide of Bluetongue Virus VP7 Protein Is Responsible for Core Attachment to Culicoides Cells

2001 ◽  
Vol 75 (8) ◽  
pp. 3937-3947 ◽  
Author(s):  
Boon-Huan Tan ◽  
Emma Nason ◽  
Norbert Staeuber ◽  
Wenrong Jiang ◽  
Katherine Monastryrskaya ◽  
...  
Keyword(s):  
Bluetongue Virus ◽  
Mammalian Cells ◽  
Inner Core ◽  
Core Protein ◽  
Binding Activity ◽  
Cell Binding ◽  
Rgd Motif ◽  
The Core ◽  
Vp7 Protein

ABSTRACT Bluetongue virus (BTV) is an arthropod-borne virus transmitted byCulicoides species to vertebrate hosts. The double-capsid virion is infectious for Culicoides vector and mammalian cells, while the inner core is infectious for onlyCulicoides-derived cells. The recently determined crystal structure of the BTV core has revealed an accessible RGD motif between amino acids 168 to 170 of the outer core protein VP7, whose structure and position would be consistent with a role in cell entry. To delineate the biological role of the RGD sequence within VP7, we have introduced point mutations in the RGD tripeptide and generated three recombinant baculoviruses, each expressing a mutant derivative of VP7 (VP7-AGD, VP7-ADL, and VP7-AGQ). Each expressed mutant protein was purified, and the oligomeric nature and secondary structure of each was compared with those of the wild-type (wt) VP7 molecule. Each mutant VP7 protein was used to generate empty core-like particles (CLPs) and were shown to be biochemically and morphologically identical to those of wt CLPs. However, when mutant CLPs were used in an in vitro cell binding assay, each showed reduced binding to Culicoides cells compared to wt CLPs. Twelve monoclonal antibodies (MAbs) was generated using purified VP7 or CLPs as a source of antigen and were utilized for epitope mapping with available chimeric VP7 molecules and the RGD mutants. Several MAbs bound to the RGD motif on the core, as shown by immunogold labeling and cryoelectron microscopy. RGD-specific MAb H1.5, but not those directed to other regions of the core, inhibited the binding activity of CLPs to the Culicoides cell surface. Together, these data indicate that the RGD motif present on BTV VP7 is responsible for Culicoides cell binding activity.

Early signalling events of autophagy

2013 ◽  
Vol 55 ◽  
pp. 1-15 ◽  
Author(s):  
Laura E. Gallagher ◽  
Edmond Y.W. Chan
Keyword(s):  
Protein Kinase ◽  
Focal Adhesion Kinase ◽  
Focal Adhesion ◽  
Mammalian Cells ◽  
Mammalian Target Of Rapamycin ◽  
Interacting Protein ◽  
The Core ◽  
Autophagy Gene ◽  
Autophagy Induction ◽  
Cellular Pathways

Autophagy is a conserved cellular degradative process important for cellular homoeostasis and survival. An early committal step during the initiation of autophagy requires the actions of a protein kinase called ATG1 (autophagy gene 1). In mammalian cells, ATG1 is represented by ULK1 (uncoordinated-51-like kinase 1), which relies on its essential regulatory cofactors mATG13, FIP200 (focal adhesion kinase family-interacting protein 200 kDa) and ATG101. Much evidence indicates that mTORC1 [mechanistic (also known as mammalian) target of rapamycin complex 1] signals downstream to the ULK1 complex to negatively regulate autophagy. In this chapter, we discuss our understanding on how the mTORC1–ULK1 signalling axis drives the initial steps of autophagy induction. We conclude with a summary of our growing appreciation of the additional cellular pathways that interconnect with the core mTORC1–ULK1 signalling module.

scholarly journals Plasma Treatment of Fish Cells: The Importance of Defining Cell Culture Conditions in Comparative Studies

2021 ◽  
Vol 11 (6) ◽  
pp. 2534
Author(s):  
Henrike Rebl ◽  
Claudia Bergemann ◽  
Sebastian Rakers ◽  
Barbara Nebe ◽  
Alexander Rebl
Keyword(s):  
Plasma Treatment ◽  
Cell Lines ◽  
Mammalian Cells ◽  
Atmospheric Pressure Plasma ◽  
Fish Farming ◽  
Skin Lesions ◽  
Atmospheric Plasma ◽  
Fish Cell ◽  
Farmed Fish ◽  
Fish Cells

The present study provides the fundamental results for the treatment of marine organisms with cold atmospheric pressure plasma. In farmed fish, skin lesions may occur as a result of intensive fish farming. Cold atmospheric plasma offers promising medical potential in wound healing processes. Since the underlying plasma-mediated mechanisms at the physical and cellular level are yet to be fully understood, we investigated the sensitivity of three fish cell lines to plasma treatment in comparison with mammalian cells. We varied (I) cell density, (II) culture medium, and (III) pyruvate concentration in the medium as experimental parameters. Depending on the experimental setup, the plasma treatment affected the viability of the different cell lines to varying degrees. We conclude that it is mandatory to use similar cell densities and an identical medium, or at least a medium with identical antioxidant capacity, when studying plasma effects on different cell lines. Altogether, fish cells showed a higher sensitivity towards plasma treatment than mammalian cells in most of our setups. These results should increase the understanding of the future treatment of fish.

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