Bluetongue virus (BTV) causes Bluetongue (BT) of ruminants vectored by culicoides midges. It is also a classic model for studying the release mechanism of non-enveloped virus. This review begins with the infection and assembly of BTV, then summarizes the advances of different ways of releasing BTV. This includes BTV-induced autophagy and the release as extracellular vesicles via multivesicular bodies, BTV-induced apoptosis and the lytic release, as well as different pathways of release through budding via plasma membrane. The regulatory mechanisms of NS3 which is a key non-structural protein during the release of BTV are also discussed, providing a basis for further understanding the molecular mechanisms underpinning the infection, proliferation and release of BTV.
Animals ; Bluetongue ; Bluetongue virus ; Ceratopogonidae ; Sheep ; Viral Nonstructural Proteins

Influenza B virus ; genetics ; pathogenicity ; Phylogeny ; RNA, Viral ; genetics ; Viral Nonstructural Proteins ; genetics

Escherichia coli ; genetics ; Humans ; Recombinant Fusion Proteins ; immunology ; Viral Nonstructural Proteins ; genetics ; immunology

OBJECTIVETo investigate the biological role of auto-induced expression of hepatitis C virus (HCV) core protein (protein C) using a recombinant protein in an in vitro cell-based system.

METHODSThe PCR-amplified full-length HCV protein C gene (573 bp) was inserted into the pET28a prokaryotic expression vector. The recombinant plasmid was transformed into BL21(DE3)pLysS E. coli to achieve high-concentration expression of the recombinant C protein by auto-induction. The recombinant protein C was purified by Ni-NTA affinity chromatography, and tested in a protein binding assay for its ability to bind the HCV NS3 protein.

RESULTSThe transformed E. coli produced a large amount of recombinant protein C, as detected in the sonicated supernatant of the bacteria culture. The antigenic reactivity of the recombinant protein C was confirmed by western blotting. However, the recombinant protein C could not be purified by Ni-NTA affinity chromatography, but co-precipitated with the HCV NS3 protein.

CONCLUSIONSoluble recombinant protein C was successfully expressed by auto-induction, and shown to interact with the HCV NS3 protein, which provides a novel insight into the putative biological activity of this factor in HCV-related molecular processes. Future studies of this recombinant HCV protein C's crystal structure and antigenicity may provide further clues to its biological function(s) and potential for clinical applications.

Escherichia coli ; metabolism ; Genetic Vectors ; Hepacivirus ; Recombinant Proteins ; genetics ; metabolism ; Viral Core Proteins ; biosynthesis ; genetics ; metabolism ; Viral Nonstructural Proteins ; metabolism

OBJECTIVETo investigate biological functions of hepatitis C virus (HCV) non-structural protein 4A (NS4A).

METHODSYeast-two hybrid technique was performed to seek proteins in hepatocytes interacting with HCV NS4A. HCV NS4A bait plasmid was constructed by ligating the NS4A gene with carrier plasmid pGBKT7, then it was transformed into yeast AH109 (alpha type). The transformed yeast cells were amplified and mated with yeast cells Y187 (alpha type) containing liver cDNA library plasmid pACT2 in 2 x YPDA medium. Diploid yeast cells were plated on synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) and synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) containing X-alpha-gal for selection two times. After extracting plasmid from blue colonies, plasmid DNA was transformed into competent E.coli and analyzed by DNA sequencing and bioinformatics methods.

RESULTSAmong twenty-two positive colonies there were eleven positive for metallothionein 2A, three for eukaryotic translation elongation factor 1 alpha 1, two for albumin, two for RNA binding motif protein 21, two for myomesin, one for cytochrome C oxidase II, and one for ATPase.

CONCLUSIONSGenes of HCV NS4A interacting proteins in hepatocytes were successfully cloned and the results pave the way for studying the biological functions of NS4A and associated proteins.

Carrier Proteins ; genetics ; Cloning, Molecular ; Hepacivirus ; genetics ; Hepatocytes ; metabolism ; Humans ; Two-Hybrid System Techniques ; Viral Nonstructural Proteins ; Viral Proteins ; genetics

Animals ; Arteritis Virus, Equine ; chemistry ; genetics ; Horses ; Open Reading Frames ; Viral Nonstructural Proteins ; chemistry ; physiology ; Viral Structural Proteins ; chemistry ; physiology

Apoptosis ; Cell Line ; Hepacivirus ; genetics ; Humans ; Serum ; metabolism ; Viral Nonstructural Proteins ; genetics

OBJECTIVETo confirm the activity of non structural protein 1 (NSP1) of Rotavirus (RV) as E3 ubiquitin ligase by experiments and to provide some clues for NSP1 on the pathogenic mechanisms and replication of RV.

METHODSThe whole gene and RING deleted mutation gene of NSP1 were coloned into pEGFPC1 expression plasmid, and transfected into human embryonic kidney (HEK) 293 FT cells with pBlue-Script-HA-Ubiquitin. The expression of proteins were proved by using con-focal microscope and western blotting. The ubiquination of proteins were detected by co-immunoprecite.

RESULTSThe cellular proteins of HEK293FT are ubiquinated by NSP1 protein and NSP1 protein was self-ubiquinated also.

CONCLUSIONSIt revealed that RV NSP1 had the activity of E3 ubiquitin ligase and it may play a role on the modulate mechanisms of ubiquination.

HEK293 Cells ; Humans ; Rotavirus ; enzymology ; genetics ; Ubiquitin-Protein Ligases ; metabolism ; Viral Nonstructural Proteins ; genetics ; metabolism

Until now, more than seventeen parvovirus have been reported which can infect mammals and poultries. The infected cells appeared different properties of apoptosis and death, present a typical cytopathic effect. NS1 is a major nonstructural protein of parvovirus, with a conservative structure and function, which plays an important role in the viral life cycle. In addition to the influence on viral replication, the NS1 also participates in apoptosis induced by viruses. Parvovirus induced apoptosis which is mainly mediated by mitochondrial pathway, this review summarized the latest research progresses of parvovirus induced apoptosis.
Animals ; Apoptosis ; Humans ; Parvoviridae Infections ; physiopathology ; veterinary ; virology ; Parvovirus ; genetics ; metabolism ; Viral Nonstructural Proteins ; genetics ; metabolism

Human bocavirus 1 (HBoV1) non-structural protein NS1 is a multifunctional protein important for virus replication and induction of apoptosis in host cell. To better understand the function of the NS1 protein, it is urgent to address reducing the toxicity of NS1 to host cells. In the present study, we established a stable cell line that regulates expression of NS1 of HBoV1. The recombinant lentivirus plasmid containing a regulatable promoter fused with ns1 gene was constructed and transfected into HEK 293T cells using transfection reagent. The HEK 293T cell lines stably expressing NS1-100 and NS1-70 proteins were established by screening resistant cells with puromycin and inducing NS1 expression with doxycycline. The expression of NS1 protein was determined by fluorescent labeling protein and Western blotting. HBoV1 promoter was transfected into stably expressing NS1 cell line and its trans-transcriptional activity was analyzed. The results showed that NS1 protein was expressed stably in the established cell lines and had a strong activation activity on the HBoV1 promoter driving luciferase gene. Taken together, this study provides a solid basis for further research on the function of NS1 and the pathogenesis of human bocavirus.
Human bocavirus ; Promoter Regions, Genetic ; Transcriptional Activation ; Viral Nonstructural Proteins ; Virus Replication