scholarly journals Role of Interferon in the Replication of Human Parainfluenza Virus Type 1 Wild Type and Mutant Viruses in Human Ciliated Airway Epithelium

2008 ◽  
Vol 82 (16) ◽  
pp. 8059-8070 ◽  
Author(s):  
Emmalene J. Bartlett ◽  
Margaret Hennessey ◽  
Mario H. Skiadopoulos ◽  
Alexander C. Schmidt ◽  
Peter L. Collins ◽  
...  
Keyword(s):  
Virus Type ◽  
Airway Epithelium ◽  
Parainfluenza Virus ◽  
Progeny Virus ◽  
Wild Type ◽  
Vaccine Candidates ◽  
C Proteins ◽  
Human Parainfluenza Virus

ABSTRACT Human parainfluenza virus type 1 (HPIV1) is a significant cause of pediatric respiratory disease in the upper and lower airways. An in vitro model of human ciliated airway epithelium (HAE), a useful tool for studying respiratory virus-host interactions, was used in this study to show that HPIV1 selectively infects ciliated cells within the HAE and that progeny virus is released from the apical surface with little apparent gross cytopathology. In HAE, type I interferon (IFN) is induced following infection with an HPIV1 mutant expressing defective C proteins with an F170S amino acid substitution, rHPIV1-CF170S, but not following infection with wild-type HPIV1. IFN induction coincided with a 100- to 1,000-fold reduction in virus titer, supporting the hypothesis that the HPIV1 C proteins are critical for the inhibition of the innate immune response. Two recently characterized live attenuated HPIV1 vaccine candidates expressing mutant C proteins were also evaluated in HAE. The vaccine candidates, rHPIV1-CR84G/Δ170HNT553ALY942A and rHPIV1-CR84G/Δ170HNT553ALΔ1710-11, which contain temperature-sensitive (ts) attenuating (att) and non-ts att mutations, were highly restricted in growth in HAE at permissive (32°C) and restrictive (37°C) temperatures. The viruses grew slightly better at 37°C than at 32°C, and rHPIV1-CR84G/Δ170HNT553ALY942A was less attenuated than rHPIV1-CR84G/Δ170HNT553ALΔ1710-11. The level of replication in HAE correlated with that previously observed for African green monkeys, suggesting that the HAE model has potential as a tool for the preclinical evaluation of HPIV1 vaccines, although how these in vitro data will correlate with vaccine virus replication in seronegative human subjects remains to be seen.

scholarly journals Human Parainfluenza Virus Type 1 C Proteins Are Nonessential Proteins That Inhibit the Host Interferon and Apoptotic Responses and Are Required for Efficient Replication in Nonhuman Primates

2008 ◽  
Vol 82 (18) ◽  
pp. 8965-8977 ◽  
Author(s):  
Emmalene J. Bartlett ◽  
Ann-Marie Cruz ◽  
Janice Esker ◽  
Adam Castaño ◽  
Henrick Schomacker ◽  
...  
Keyword(s):  
Virus Type ◽  
Parainfluenza Virus ◽  
Airway Epithelial ◽  
Efficient Replication ◽  
C Proteins ◽  
Induced Apoptosis ◽  
Human Parainfluenza Virus

ABSTRACT Recombinant human parainfluenza virus type 1 (rHPIV1) was modified to create rHPIV1-P(C−), a virus in which expression of the C proteins (C′, C, Y1, and Y2) was silenced without affecting the amino acid sequence of the P protein. Infectious rHPIV1-P(C−) was readily recovered from cDNA, indicating that the four C proteins were not essential for virus replication. Early during infection in vitro, rHPIV1-P(C−) replicated as efficiently as wild-type (wt) HPIV1, but its titer subsequently decreased coincident with the onset of an extensive cytopathic effect not observed with wt rHPIV1. rHPIV1-P(C−) infection, but not wt rHPIV1 infection, induced caspase 3 activation and nuclear fragmentation in LLC-MK2 cells, identifying the HPIV1 C proteins as inhibitors of apoptosis. In contrast to wt rHPIV1, rHPIV1-P(C−) and rHPIV1-CF170S, a mutant encoding an F170S substitution in C, induced interferon (IFN) and did not inhibit IFN signaling in vitro. However, only rHPIV1-P(C−) induced apoptosis. Thus, the anti-IFN and antiapoptosis activities of HPIV1 were separable: both activities are disabled in rHPIV1-P(C−), whereas only the anti-IFN activity is disabled in rHPIV1-CF170S. In African green monkeys (AGMs), rHPIV1-P(C−) was considerably more attenuated than rHPIV1-CF170S, suggesting that disabling the anti-IFN and antiapoptotic activities of HPIV1 had additive effects on attenuation in vivo. Although rHPIV1-P(C−) protected against challenge with wt HPIV1, its highly restricted replication in AGMs and in primary human airway epithelial cell cultures suggests that it might be overattenuated for use as a vaccine. Thus, the C proteins of HPIV1 are nonessential but have anti-IFN and antiapoptosis activities required for virulence in primates.

scholarly journals A novel human parainfluenza virus type 1 (HPIV1) with separated P and C genes is useful for generating C gene mutants for evaluation as live-attenuated virus vaccine candidates

Vaccine ◽  
2010 ◽  
Vol 28 (3) ◽  
pp. 767-779 ◽  
Author(s):  
Emmalene J. Bartlett ◽  
Ann-Marie Cruz ◽  
Jim Boonyaratanakornkit ◽  
Janice Esker ◽  
Adam Castaño ◽  
...  
Keyword(s):  
Virus Type ◽  
Virus Vaccine ◽  
Parainfluenza Virus ◽  
Vaccine Candidates ◽  
Attenuated Virus ◽  
Human Parainfluenza Virus

scholarly journals Attenuation and efficacy of human parainfluenza virus type 1 (HPIV1) vaccine candidates containing stabilized mutations in the P/C and L genes

2007 ◽  
Vol 4 (1) ◽  
pp. 67 ◽  
Author(s):  
Emmalene J Bartlett ◽  
Adam Castaño ◽  
Sonja R Surman ◽  
Peter L Collins ◽  
Mario H Skiadopoulos ◽  
...  
Keyword(s):  
Virus Type ◽  
Parainfluenza Virus ◽  
Vaccine Candidates ◽  
Human Parainfluenza Virus

scholarly journals Generation of Recombinant Human Parainfluenza Virus Type 1 Vaccine Candidates by Importation of Temperature-Sensitive and Attenuating Mutations from Heterologous Paramyxoviruses

2004 ◽  
Vol 78 (4) ◽  
pp. 2017-2028 ◽  
Author(s):  
Jason T. Newman ◽  
Jeffrey M. Riggs ◽  
Sonja R. Surman ◽  
Josephine M. McAuliffe ◽  
Teresa A. Mulaikal ◽  
...  
Keyword(s):  
Respiratory Tract ◽  
Virus Type ◽  
Neutralizing Antibodies ◽  
Effective Means ◽  
Parainfluenza Virus ◽  
Temperature Sensitive ◽  
C Protein ◽  
Vaccine Candidates ◽  
Human Parainfluenza Virus

ABSTRACT Human parainfluenza virus type 1 (HPIV1) is a significant cause of respiratory tract disease in infants and young children for which a vaccine is needed. In the present study, we sought to attenuate HPIV1 by the importation of one or more known attenuating point mutations from heterologous paramyxoviruses into homologous sites in HPIV1. The introduced mutations were derived from three attenuated paramyxoviruses: (i) HPIV3cp45, a live-attenuated HPIV3 vaccine candidate containing multiple attenuating mutations; (ii) the respiratory syncytial virus cpts530 with an attenuating mutation in the L polymerase protein; and (iii) a murine PIV1 (MPIV1) attenuated by a mutation in the accessory C protein. Recombinant HPIV1 (rHPIV1) mutants bearing a single imported mutation in C, any of three different mutations in L, or a pair of mutations in F exhibited a 100-fold or greater reduction in replication in the upper or lower respiratory tract of hamsters. Both temperature-sensitive (ts) (mutations in the L and F proteins) and non-ts (the mutation in the C protein) attenuating mutations were identified. rHPIV1 mutants containing a combination of mutations in L were generated that were more attenuated than viruses bearing the individual mutations, showing that the systematic accretion of mutations can yield progressive increases in attenuation. Hamsters immunized with rHPIV1 mutants bearing one or two mutations developed neutralizing antibodies and were resistant to challenge with wild-type HPIV1. Thus, importation of attenuating mutations from heterologous viruses is an effective means for rapidly identifying mutations that attenuate HPIV1 and for generating live-attenuated HPIV1 vaccine candidates.

scholarly journals Parainfluenza Virus Type 1 Induces Epithelial IL-8 Production via p38-MAPK Signalling

2014 ◽  
Vol 2014 ◽  
pp. 1-12 ◽  
Author(s):  
Miguel Ángel Galván Morales ◽  
Carlos Cabello Gutiérrez ◽  
Fidencio Mejía Nepomuceno ◽  
Leticia Valle Peralta ◽  
Elba Valencia Maqueda ◽  
...  
Keyword(s):  
Viral Replication ◽  
Cell Lines ◽  
Virus Type ◽  
Molecular Mechanisms ◽  
Parainfluenza Virus ◽  
Kinase Activation ◽  
Mitogen Activated Protein ◽  
Human Parainfluenza Virus

Human parainfluenza virus type 1 (HPIV-1) is the most common cause of croup in infants. The aim of this study was to describe molecular mechanisms associated with IL-8 production during HPIV-1 infection and the role of viral replication in MAPK synthesis and activation. Anin vitromodel of HPIV-1 infection in the HEp-2 and A549 cell lines was used; a kinetic-based ELISA for IL-8 detection was also used, phosphorylation of the mitogen-activated protein kinases (MAPKs) was identified by Western blot analysis, and specific inhibitors for each kinase were used to identify which MAPK was involved. Inactivated viruses were used to assess whether viral replication is required for IL-8 production. Results revealed a gradual increase in IL-8 production at different selected times, when phosphorylation of MAPK was detected. The secretion of IL-8 in the two cell lines infected with the HPIV-1 is related to the phosphorylation of the MAPK as well as viral replication. Inhibition of p38 suppressed the secretion of IL-8 in the HEp-2 cells. No kinase activation was observed when viruses were inactivated.

scholarly journals The Long Noncoding Region of the Human Parainfluenza Virus Type 1 F Gene Contributes to the Read-Through Transcription at the M-F Gene Junction

2002 ◽  
Vol 76 (16) ◽  
pp. 8244-8251 ◽  
Author(s):  
Tatiana Bousse ◽  
Tatyana Matrosovich ◽  
Allen Portner ◽  
Atsushi Kato ◽  
Yoshiyuki Nagai ◽  
...  
Keyword(s):  
Virus Type ◽  
Parainfluenza Virus ◽  
Noncoding Sequence ◽  
Wild Type ◽  
F Gene ◽  
Transcriptional Termination ◽  
Infected Cells ◽  
Human Parainfluenza Virus ◽  
Gene Junction

ABSTRACT Sendai virus (SV) and human parainfluenza virus type 1 (hPIV1) have genomes consisting of nonsegmented negative-sense RNA in which the six genes are separated by well-conserved intergenic (IG) sequences and transcriptional start (S) and end signals. In hPIV1-infected cells, transcriptional termination at the M-F gene junction is ineffective; a large number of M-F read-through transcripts are produced (T. Bousse, T. Takimoto, K. G. Murti, and A. Portner, Virology 232:44-52, 1997). In contrast, few M-F read-through transcripts are detected in SV-infected cells. Sequence analysis indicated that the hPIV1 IG and S sequences in the M-F junction differ from those of SV. Furthermore, the hPIV1 F gene contains an unusually long noncoding sequence. To identify the cis-acting elements that prevent transcriptional termination at the M-F junction, we rescued recombinant SV (rSVhMFjCG) in which its M-F gene junction was replaced by that of hPIV1. Cells infected with rSVhMFjCG produced an abundance of M-F read-through transcripts; this result indicated that the hPIV1 M-F junction is responsible for inefficient termination. When one or both of the IG and S sites in rSVhMFjCG were replaced by those of SV, the efficiency of transcriptional termination increased but not to the level observed in wild-type SV-infected cells. Deletion of most of the long noncoding region of the hPIV1 F gene in rSVhMFjCG in addition to the mutations in IG and S signals resulted in efficient termination that was equivalent to the level observed in wild-type virus-infected cells. Therefore, the long noncoding sequence of the hPIV1 F gene contains cis-acting element(s) that affects transcriptional termination. Our evaluation of the effect of inefficient transcriptional termination on viral replication in culture revealed that cells infected with rSVhMFjCG produced less F protein than cells infected with wild-type SV and that assembly of the recombinant SV in culture was less efficient. These phenotypes seem to be responsible for the extended survival of mice infected with rSVhMFjCG.

scholarly journals Replacement of the Ectodomains of the Hemagglutinin-Neuraminidase and Fusion Glycoproteins of Recombinant Parainfluenza Virus Type 3 (PIV3) with Their Counterparts from PIV2 Yields Attenuated PIV2 Vaccine Candidates

2000 ◽  
Vol 74 (14) ◽  
pp. 6448-6458 ◽  
Author(s):  
Tao Tao ◽  
Mario H. Skiadopoulos ◽  
Fatemeh Davoodi ◽  
Jeffrey M. Riggs ◽  
Peter L. Collins ◽  
...  
Keyword(s):  
Virus Type ◽  
Vaccine Candidate ◽  
Cytoplasmic Tail ◽  
Full Length ◽  
Parainfluenza Virus ◽  
Tail Domain ◽  
Wild Type ◽  
Vaccine Candidates

ABSTRACT We sought to develop a live attenuated parainfluenza virus type 2 (PIV2) vaccine strain for use in infants and young children, using reverse genetic techniques that previously were used to rapidly produce a live attenuated PIV1 vaccine candidate. The PIV1 vaccine candidate, designated rPIV3-1cp45, was generated by substituting the full-length HN and F proteins of PIV1 for those of PIV3 in the attenuatedcp45 PIV3 vaccine candidate (T. Tao et al., J. Virol. 72:2955–2961, 1998; M. H. Skiadopoulos et al., Vaccine 18:503–510, 1999). However, using the same strategy, we failed to recover recombinant chimeric PIV3-PIV2 isolate carrying the full-length PIV2 glycoproteins in a wild-type PIV3 backbone. Viable PIV3-PIV2 chimeras were recovered when chimeric HN and F open reading frames (ORFs) rather than complete PIV2 F and HN ORFs were used to construct the full-length cDNA. The recovered viruses, designated rPIV3-2CT, in which the PIV2 ectodomain and transmembrane domain were fused to the PIV3 cytoplasmic domain, and rPIV3-2TM, in which the PIV2 ectodomain was fused to the PIV3 transmembrane and cytoplasmic tail domain, possessed similar in vitro and in vivo phenotypes. Thus, it appeared that only the cytoplasmic tail of the HN or F glycoprotein of PIV3 was required for successful recovery of PIV3-PIV2 chimeras. Although rPIV3-2CT and rPIV3-2TM replicated efficiently in vitro, they were moderately to highly attenuated for replication in the respiratory tracts of hamsters, African green monkeys (AGMs), and chimpanzees. This unexpected finding indicated that chimerization of the HN and F proteins of PIV2 and PIV3 itself specified an attenuation phenotype in vivo. Despite this attenuation, these viruses were highly immunogenic and protective against challenge with wild-type PIV2 in hamsters and AGMs, and they represent promising candidates for clinical evaluation as a vaccine against PIV2. These chimeric viruses were further attenuated by the addition of 12 mutations of PIV3cp45 which lie outside of the HN and F genes. The attenuating effects of these mutations were additive with that of the chimerization, and thus inclusion of all or some of the cp45 mutations provides a means to further attenuate the PIV3-PIV2 chimeric vaccine candidates if necessary.

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