Apoptosis in Cultured Cells Infected with Feline Calicivirus

Roberts, Lisa O.; Al-Molawi, Naema; Carter, Michael J.; Kass, George E.N.

doi:10.1196/annals.1299.110

Cited by 16 publications

(9 citation statements)

References 4 publications

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“…(Fig. 5; Video S1 can be viewed to observe fluorescence over a longer time course), although cells began to collapse and bleb approximately 5-6 hours p.i., a morphology consistent with reports of virus-induced apoptosis (Natoni et al, 2006; Roberts et al, 2003; Sosnovtsev et al, 2003). Supplementary Video 2 (which extended beyond the first 9.5 hrs shown in Fig.…”

Section: Resultssupporting

confidence: 91%

Visualization of feline calicivirus replication in real-time with recombinant viruses engineered to express fluorescent reporter proteins

et al. 2010

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Caliciviruses are non-enveloped, icosahedral viruses with a single-stranded, positive sense RNA genome. Transposon-mediated insertional mutagenesis was used to insert a transprimer sequence into random sites of an infectious full-length cDNA clone of the feline calicivirus (FCV) genome. A site in the LC gene (encoding the capsid leader protein) of the FCV genome was identified that could tolerate foreign insertions, and two viable recombinant FCV variants expressing LC fused either to AcGFP, or DsRedFP were recovered. The effects of the insertions on LC processing, RNA replication, and stability of the viral genome were analyzed, and the progression of a calicivirus single infection and co-infection were captured by real-time imaging fluorescent microscopy. The ability to engineer viable recombinant caliciviruses expressing foreign markers enables new approaches to investigate virus and host cell interactions, as well as studies of viral recombination, one of the driving forces of calicivirus evolution.

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Section: Resultssupporting

confidence: 91%

Visualization of feline calicivirus replication in real-time with recombinant viruses engineered to express fluorescent reporter proteins

et al. 2010

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“…Apoptosis of hepatocytes was observed in naturally infected (3). Apoptosis has also been described for feline calicivirus (FCV) infection of cell cultures (2,37,44). Of interest, induction of apoptosis by FCV requires active viral replication (44).…”

Section: Discussionmentioning

confidence: 93%

Pathogenesis of a Genogroup II Human Norovirus in Gnotobiotic Pigs

Cheetham

Souza

Meulia

et al. 2006

J Virol

256

264

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Human noroviruses (HuNoVs) are a major cause of foodborne gastroenteritis worldwide. Because they do not grow in cell culture and there is no animal model for HuNoVs, pathogenesis studies have been hampered. Thus, little is known about their replication strategies or induction of neutralizing antibodies.The limited information on their pathogenesis is from human volunteer studies of HuNoV infections in which villus atrophy in duodenal biopsies and presence of malabsorptive diarrhea were described (1,7,8). No information is available on lesions in other portions of the intestines of these volunteers (9). Intestinal transplant pediatric patients that were diagnosed with HuNoV infection developed secretory or osmotic diarrhea (19,20,31). These patients had prolonged diarrhea (17 to 326 days) due to immunosuppressive therapy. The detection of HuNoV RNA and the clinical symptoms remitted after reduction of the immunosuppressive therapy. Usually in exposed individuals, histologic lesions correlate with diarrhea, but in one report, lesions in volunteers who did not show clinical symptoms were described (42). There are also numerous reports of asymptomatic individuals who were infected with HuNoVs and shed virus in the feces (11, 28).Most past attempts to study these viruses in an animal model may have failed because (i) the human strains that were used were not closely related to the host animal NoV strains, (ii) sensitive detection techniques were lacking, and finally (iii) the role of histo-blood group antigen (HBGA) phenotypes in differential susceptibility of the host was unrecognized. Our goal was to adapt a HuNoV strain to replicate in the gnotobiotic (Gn) pig to develop an animal model for the study of HuNoV pathogenesis. Gnotobiotic pigs are good models for human enteric diseases (40) because pigs resemble humans in their gastrointestinal anatomy, physiology, and immune responses. The Gn pigs are immunocompetent at birth, but they lack maternal antibodies and previous or ongoing exposure to microbial agents, including caliciviruses.Recently, viral RNA genetically similar to that of human NoV GII (65 to 71% amino acid sequence identity in the capsid gene) was detected in pigs in Japan (46, 47) and Europe (22,48). In U.S. swine, our laboratory detected both viral RNA and virus particles similar to GII HuNoV (70% sequence identity in the capsid region) which were infectious for Gn pigs (50). Our approach to infect Gn pigs with a HuNoV was to use a GII strain that is closely related genetically to the identified GII porcine NoVs and that has a broad HBGA binding pattern because little information or reagents are available for pig HBGA. Additionally, we used sensitive assays and reagents including reverse transcription (RT)-PCR to detect fecal shedding, virus-like particles (VLPs) for serological assays, and antisera to these VLPs for antigen enzyme-linked immunosor-

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“…5C), indicating that overexpression of the LC was toxic to CRFK cells and caused cell death. To determine if the cell death caused by transient LC expression was associated with apoptosis, which is induced in FCV-infected cells (31,34,35), a cell-permeable fluorogenic peptide marker (FITC-VAD-FMK) was employed to detect activated caspases in LCmKate-expressing cells. The majority of cells positive for LCmKate were positive also for activated caspases (Fig.…”

Section: Resultsmentioning

confidence: 99%

The Feline Calicivirus Leader of the Capsid Protein Is Associated with Cytopathic Effect

Abente

Sosnovtsev

Sandoval-Jaime

et al. 2013

J Virol

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Open reading frame 2 (ORF2) of the feline calicivirus (FCV) genome encodes a capsid precursor that is posttranslationally processed to release the mature capsid protein (VP1) and a small protein of 124 amino acids, designated the leader of the capsid (LC). To investigate the role of the LC protein in the virus life cycle, mutations and deletions were introduced into the LC coding region of an infectious FCV cDNA clone. Three cysteine residues that are conserved among all vesivirus LC sequences were found to be critical for the recovery of FCV with a characteristic cytopathic effect in feline kidney cells. A cell-rounding phenotype associated with the transient expression of wild-type and mutagenized forms of the LC correlated with the cytopathic and growth properties of the corresponding engineered viruses. The host cellular protein annexin A2 was identified as a binding partner of the LC protein, consistent with a role for the LC in mediating host cell interactions that alter the integrity of the cell and enable virus spread.M embers of the family Caliciviridae are small nonenveloped viruses that contain a positive-sense single-stranded RNA genome. Feline calicivirus (FCV) is in the genus Vesivirus of the family and has been an important model for studying calicivirus replication because it grows efficiently in cell culture and has a reverse genetics system (1-5). The RNA genomes of caliciviruses range in size from ϳ6.7 to 8.5 kb and typically encode 8 or 9 viral proteins from two (Sapovirus, Nebovirus, and Lagovirus) or three (Norovirus and Vesivirus) open reading frames (ORFs). A unique ORF4 in the murine norovirus genome that encodes a protein reported to downregulate the innate immune response has been identified (6). The 5= end of the viral RNA genome is covalently linked to a VPg protein; the 3= end of the genome is polyadenylated (2, 7-9). The infectious virion is composed of 180 copies of the major capsid protein VP1 and 1 to 10 copies of the minor structural protein VP2, which together form a capsid around the VPg-linked genome (10-15). The genomic RNA serves as a template for translation of the ORF1 polyprotein that is proteolytically cleaved by the viral protease to release the nonstructural proteins (16,17), and an ϳ2.2-to 2.6-kb subgenomic bicistronic RNA template is used for the translation of VP1 and VP2 (7, 18). A genetic feature unique to members of the genus Vesivirus is expression of the major capsid protein from ORF2 as a precursor protein (5,(18)(19)(20). This precursor is processed in trans by the viral protease to release two proteins: the leader of the capsid (LC) and the mature capsid protein (VP1) (5,19,21,22). The function of the LC protein is not clear, but cleavage of the precursor to release LC and VP1 is essential for the recovery of infectious virions (5). Transient expression of the LC was reported to enhance replication of a human norovirus RNA replicon (23), and an increase in the level of mRNA for the low-density lipoprotein receptor (LDLR) was observed (24). We showed previo...

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Apoptosis in Cultured Cells Infected with Feline Calicivirus

Cited by 16 publications

References 4 publications

Visualization of feline calicivirus replication in real-time with recombinant viruses engineered to express fluorescent reporter proteins

Visualization of feline calicivirus replication in real-time with recombinant viruses engineered to express fluorescent reporter proteins

Pathogenesis of a Genogroup II Human Norovirus in Gnotobiotic Pigs

The Feline Calicivirus Leader of the Capsid Protein Is Associated with Cytopathic Effect

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