Identification of the sequences responsible for nuclear targeting of the V protein of human parainfluenza virus type 2

Rinderpest virus is a morbillivirus and the causative agent of an important disease of cattle and wild bovids. The P genes of all morbilliviruses give rise to two proteins in addition to the P protein itself: use of an alternate start translation site, in a second open reading frame, gives rise to the C protein, while cotranscriptional insertion of an extra base gives rise to the V protein, a fusion of the amino-terminal half of P to a short, highly conserved, cysteine-rich zinc binding domain. Little is known about the function of either of these two proteins in the rinderpest virus life cycle. We have constructed recombinant rinderpest viruses in which the expression of these proteins has been suppressed, individually and together, and studied the replication of these viruses in tissue culture. We show that the absence of the V protein has little effect on the replication rate of the virus but does lead to an increase in synthesis of genome and antigenome RNAs and a change in cytopathic effect to a more syncytium-forming phenotype. Virus that does not express the C protein, on the other hand, is clearly defective in growth in all cell lines tested, and this defect appears to be related to a decreased transcription of mRNA from viral genes. The phenotypes of both individual mutant virus types are both expressed in the double mutant expressing neither V nor C.Rinderpest virus (RPV) belongs to the Morbillivirus genus of the family Paramyxoviridae and is thus related to Measles virus (MV), Canine and Phocid (seal) distemper viruses, the cetacean morbillivirus, and Peste des petits ruminants virus. The disease it causes (rinderpest) has for decades been one of the most widespread and economically important diseases affecting cattle in Africa, the Middle East, and the Indian subcontinent. It has been eliminated from most of these areas in recent years, largely through the efforts of the European Union-and Food and Agriculture Organization-backed EMPRES program, with the aim of global eradication by 2010 (22). Like all of the morbilliviruses, there is only one serotype of RPV, yet wide variation can be found in the pathogenicity of field isolates, from those causing essentially 100% mortality to others in which infection causes barely detectable clinical signs (58, 64). The molecular basis of this variation is unknown.All of the Paramyxoviridae have single-stranded RNA genomes of negative sense, with six viral genes, which are transcribed in order from a single promoter at the 3Ј end of the genome. From the second of those genes (the P gene), through utilization of more than one translation initiation codon and/or the introduction of one or more nontemplated residues to allow access to alternate reading frames, more than one protein is always produced, though the exact number and type of extra proteins vary both between and within genera. The expression of other proteins from overlapping reading frames was first shown in Sendai virus (SeV) (26, 55). SeV (15, 29), and Human parainfluenza virus type 1 (hPIV1) (8) expre...

Section: Rinderpest Virus (Rpv) Belongs To the Morbillivirus Genus Ofmentioning

confidence: 99%

Rinderpest Viruses Lacking the C and V Proteins Show Specific Defects in Growth and Transcription of Viral RNAs

Baron

Barrett

2000

“…Recovery of a completely V-minus rPIV2. The rubulavirus V protein is reported to be multifunctional (22,23,36,47,48). To assess the extent to which the various mutations in V affected its global functions, we attempted the recovery of a recombinant hPIV2 which completely lacks expression of V protein, to provide the other point of reference with which the mutant rPIV2s can be compared.…”

Section: Vol 79 2005 V Protein Of Paramyxoviruses 8595mentioning

confidence: 99%

“…Monoclonal antibodies (MAbs) against hPIV2 P/V protein (85A), hPIV2 V protein (53V), and hPIV2 NP protein (20A) were as described previously (26,27). MAb against hPIV2 P/V protein 85A has cross-reactivity with SV41 P/V protein (47). Anti-STAT1 MAb was purchased from BD Transduction Laboratories (Lexington, KY).…”

Section: Cells and Virusesmentioning

confidence: 99%

Identification of Paramyxovirus V Protein Residues Essential for STAT Protein Degradation and Promotion of Virus Replication

Nishio

Tsurudome

Ito

et al. 2005

Some paramyxovirus V proteins induce STAT protein degradation, and the amino acids essential for this process in the human parainfluenza virus type 2 (hPIV2) V protein have been studied. Various recombinant hPIV2s and cell lines constitutively expressing various mutant V proteins were generated. We found that V proteins with replacement of Cys residues of the Cys cluster were still able to bind STATs but were unable to induce their degradation. The hPIV2 V protein binds STATs via a W-(X) 3 -W-(X) 9 -W Trp motif located just upstream of the Cys cluster. Replacements of two or more Trp residues in this motif resulted in a failure to form a V/STAT2 complex. We have also identified two Phe residues of the hPIV2 V protein that are essential for STAT degradation, namely, Phe207, lying within the Cys cluster, and Phe143, in the P/V common region of the protein. Interestingly, infection of BHK cells with hPIV2 led to the specific degradation of STAT1 and not STAT2. Other evidence for the cell species specificity of hPIV2-induced STAT degradation is presented. Finally, a V-minus hPIV2, which can express only the P protein from its P gene, was generated and partially characterized. In contrast to V-minus viruses of other paramyxovirus genera, this V-minus rubulavirus was highly debilitated, and its growth even in Vero cells was very limited. The structural rubulavirus V proteins, as expected, are thus clearly important in promoting virus growth, independent of their anti-interferon (IFN) activity. Interestingly, many of the residues that are essential for anti-IFN activity, e.g., the Cys of this cluster and Phe207 within this cluster, as well as the Trp of this motif, are also essential for promoting virus growth.The interferon (IFN) system is the first line of host defense against virus infection. Viruses of the Paramyxovirinae, similarly to other viruses, have evolved proteins that specifically inhibit IFN-induced innate antiviral responses, at least in part through direct inhibition of cellular STAT proteins that are responsible for IFN signal transduction. The V proteins encoded by the rubulaviruses simian virus 5 (SV5), SV41, and mumps virus (MuV) and by Newcastle disease virus (NDV, an avulavirus) block IFN signaling by targeting STAT1 for degradation (1,5,6,14,20,25,33,45,46,49,51), whereas the V protein of human parainfluenza virus type 2 (hPIV2, a rubulavirus) targets STAT2 for degradation (25,32). Moreover, the V proteins of measles virus (morbillivirus) and Nipah virus and Hendra virus (henipaviruses) have been shown to inhibit IFN signaling by preventing STAT1 and STAT2 nuclear accumulation (30,37,38,41). Sendai virus (SeV) and hPIV3 also block IFN signaling, and this anti-IFN ability has been shown to be a property of these respirovirus C proteins (7,8,10,11,17,19,42). In cells that are highly IFN competent, the Sendai virus C protein also induces the intracellular loss of STAT1 (9). The rubulavirus V protein-dependent degradation of STAT proteins involves degradation complexes that contain the V protein, STA...

“…The V proteins of hPIV2, SV5, and SV41 exhibit characteristic nuclear localization patterns, as described previously (38,52). Strikingly, the hPIV4 V protein was detected exclusively in the cytoplasm (Fig.…”

Section: Resultsmentioning

confidence: 63%

Human Parainfluenza Virus Type 4 Is Incapable of Evading the Interferon-Induced Antiviral Effect

Nishio

Tsurudome

Ito

et al. 2005

The V proteins of some paramyxoviruses have developed the ability to efficiently inactivate STAT protein function as a countermeasure for evading interferon (IFN) responses. Human parainfluenza virus type 4 (hPIV4) is one of the rubulaviruses, which are members of the family Paramyxoviridae, and has a V protein with a highly conserved cysteine-rich domain that is the hallmark of paramyxovirus V proteins. In order to study the function of the hPIV4 V protein, we established HeLa cells expressing the hPIV4A V protein (HeLa/ FlagPIV4V). The hPIV4 V protein had no ability to reduce the level of STAT1 or STAT2, although it associated with STAT1, STAT2, DDB1, and Cul4A. It interfered with neither STAT1 and STAT2 tyrosine phosphorylation nor IFN-induced STAT nuclear accumulation. In addition, HeLa/FlagPIV4V cells are fully sensitive to both beta interferon (IFN-␤) and IFN-␥, indicating that the hPIV4 V protein has no ability to block IFN-induced signaling. We further established HeLa cells expressing various chimeric proteins between the hPIV2 and hPIV4A V proteins. The lack of IFN-antagonistic activity of the hPIV4 V protein is caused by both the P/V common and V-specific domains. At least two regions (amino acids [aa] 32 to 45 and aa 143 to 164) of hPIV4 V in the P/V common domain and one region (aa 200 to 212) of the C terminus are involved in the inability to evade the IFN-induced signaling. Moreover, we established HeLa cells persistently infected with hPIV4 to make sure of the inability to escape IFN and confirmed that hPIV4 is the only paramyxovirus analyzed to date that can't evade the IFN-induced antiviral responses.Human parainfluenza virus type 4 (hPIV4) is a member of the family Paramyxoviridae and is known to be an important agent of pediatric respiratory tract disease in infancy (26). hPIV4 is further divided into two antigenic subtypes, hPIV4A and hPIV4B, based mainly on in vitro hemadsorption inhibition characteristics and immunological reactivity (6). We previously identified the structural proteins of hPIV4A and 4B and sequenced all genes except the L gene (2,18,21,22,23). hPIV4 contains an mRNA-editing site, similar to other rubulaviruses. The unedited version of the "P" mRNA encodes the V protein, and addition of two G nucleotides at the editing site produces an mRNA that encodes the P protein. Therefore, the N-terminal 153 amino acids (aa) of the V and P proteins are common, and their C termini are unique (22). The C terminus of hPIV4 V protein contains seven invariant cysteine residues capable of binding two atoms of zinc, and C termini are approximately 50% identical among all paramyxovirus V proteins.It has been demonstrated recently that the Paramyxovirinae, similar to other viruses, have evolved specific proteins that inhibit interferon (IFN)-induced innate antiviral responses through direct inhibition of cellular STAT proteins. The V proteins encoded by the rubulaviruses, simian virus 5 (SV5), SV41, mumps virus (MuV), and Newcastle disease virus (NDV, an avulavirus) block IFN signaling by tar...