scholarly journals Characterization of the Nuclear Localization Signal of the Borna Disease Virus Polymerase

2002 ◽  
Vol 76 (16) ◽  
pp. 8460-8467 ◽  
Author(s):  
Michelle Portlance Walker ◽  
W. Ian Lipkin
Keyword(s):  
Disease Virus ◽  
Nuclear Localization ◽  
Nuclear Localization Signal ◽  
Rna Virus ◽  
Flag Epitope ◽  
Borna Disease Virus ◽  
Localization Signal ◽  
Infected Cells ◽  
Pass Through ◽  
Borna Disease

ABSTRACT Borna disease virus (BDV) is a nonsegmented negative-strand RNA virus that replicates and transcribes its genome in the nucleus of infected cells. BDV proteins involved in replication and transcription must pass through the nuclear envelope to associate with the genomic viral RNA. The RNA-dependent RNA polymerase (L) of BDV is postulated to be the catalytic enzyme of replication and transcription. We demonstrated previously that BDV L localizes to the nucleus of BDV-infected cells and L-transfected cells. Nuclear localization of the protein presupposes the presence of a nuclear localization signal (NLS) within its primary amino acid sequence or cotransport to the nucleus with another karyophilic protein. Because L localized to the nucleus in the absence of other viral proteins, we investigated the possibility that L contains an NLS. The minimal sequence required for nuclear localization of L was identified by analyzing the subcellular distribution of deletion mutants of L fused to a flag epitope tag or β-galactosidase. Although the majority of the L fusion proteins localized to the cytoplasm of transfected BSR-T7 cells, a strong NLS (844RVVKLRIAP852) with basic and proline residues was identified. Mutation of this sequence resulted in cytoplasmic distribution of L, confirming that this sequence was necessary and sufficient to drive the nuclear localization of L.

scholarly journals Authentic Borna disease virus transcripts are spliced less efficiently than cDNA-derived viral RNAs

2000 ◽  
Vol 81 (8) ◽  
pp. 1947-1954 ◽  
Author(s):  
Christian Jehle ◽  
W. Ian Lipkin ◽  
Peter Staeheli ◽  
Rosa M. Marion ◽  
Martin Schwemmle
Keyword(s):  
Disease Virus ◽  
Rna Virus ◽  
Borna Disease Virus ◽  
Viral Rnas ◽  
Negative Strand ◽  
Intron 1 ◽  
Splicing Pattern ◽  
Alternatively Spliced ◽  
Infected Cells ◽  
Borna Disease

Borna disease virus (BDV) is a non-segmented, negative-strand RNA virus that replicates and transcribes its genome in the nucleus of infected cells. It uses the cellular splicing machinery to generate a set of alternatively spliced mRNAs from the 2·8 and 7·1 kb primary transcripts, each harbouring two introns. To determine whether splicing of these transcripts is regulated by viral factors, the extent of splicing was studied in infected cells and COS-7 cells transiently transfected with plasmids encoding the 2·8 kb RNA of BDV. Unspliced RNA was found to be the most abundant RNA species in infected cells, whereas viral transcripts lacking both introns were only found in minute amounts. In sharp contrast, plasmid-derived 2·8 kb RNA was predominantly intron 1-spliced and double-spliced. Co-expression of the BDV proteins P, N and X did not influence splicing of plasmid-expressed 2·8 kb RNA. Furthermore, the splicing pattern did not change when the 2·8 kb RNA was expressed in BDV-infected cells. Based on these results we speculate that splicing of authentic BDV transcripts is tightly linked to transcription by the viral polymerase.

scholarly journals Characterization of the P Protein-Binding Domain on the 10-Kilodalton Protein of Borna Disease Virus

2000 ◽  
Vol 74 (7) ◽  
pp. 3413-3417 ◽  
Author(s):  
Tahir H. Malik ◽  
Masahiko Kishi ◽  
Patrick K. Lai
Keyword(s):  
Disease Virus ◽  
Nuclear Localization ◽  
Cell Nucleus ◽  
Binding Domain ◽  
Borna Disease Virus ◽  
Nuclear Localization Signals ◽  
P Protein ◽  
Infected Cells ◽  
Borna Disease

ABSTRACT The Borna disease virus (BDV) is the prototype member of the Bornaviridae, and it replicates in the cell nucleus. The BDV p24P and p40N proteins carry nuclear localization signals (NLS) and are found in the nuclei of infected cells. The BDV p10 protein does not have an NLS, but it binds with P and/or N and is translocated to the nucleus. Hence, p10 may play a role in the replication of BDV in the cell nucleus. Here, we show that the P-binding domain is located in the N terminus of p10 and that S3 and L16 are important for the interaction.

scholarly journals Modulation of Borna Disease Virus Phosphoprotein Nuclear Localization by the Viral Protein X Encoded in the Overlapping Open Reading Frame

2003 ◽  
Vol 77 (14) ◽  
pp. 8099-8107 ◽  
Author(s):  
Takeshi Kobayashi ◽  
Guoqi Zhang ◽  
Byeong-Jae Lee ◽  
Satoko Baba ◽  
Makiko Yamashita ◽  
...  
Keyword(s):  
Subcellular Localization ◽  
Disease Virus ◽  
Viral Protein ◽  
Rna Virus ◽  
Viral Proteins ◽  
Borna Disease Virus ◽  
Negative Strand ◽  
Intracellular Movement ◽  
Infected Cells ◽  
Borna Disease

ABSTRACT Borna disease virus (BDV) is a nonsegmented, negative-strand RNA virus that belongs to the Mononegavirales order. Unlike other animal viruses in this order, BDV replicates and transcribes in the nucleus of infected cells. Therefore, regulation of the intracellular movement of virus components must be critical for accomplishing the BDV life cycle in mammalian cells. Previous studies have demonstrated that BDV proteins are prone to accumulate in the nucleus of cells transiently transfected with each expression plasmid of the viral proteins. In BDV infection, however, cytoplasmic distribution of the viral proteins is frequently found in cultured cells and animal brains. In this study, to understand the modulation of subcellular localization of BDV proteins, we investigated the intracellular localization of the viral phosphoprotein (P). Transient-transfection analysis with a cDNA clone corresponding to a bicistronic transcript that expresses both viral X and P revealed that P efficiently localizes in the cytoplasm only when BDV X is expressed in the cells. Furthermore, our analysis revealed that the direct binding between X and P is necessary for the cytoplasmic localization of the P. Interestingly, we showed that X is not detectably expressed in the BDV-infected cells in which P is predominantly found in the nucleus, with little or no signal in the cytoplasm. These observations suggested that BDV P can modulate their subcellular localization through binding to X and that BDV may regulate the expression ratio of each viral product in infected cells to control the intracellular movement of the viral protein complexes. The results presented here provide a new insight into the regulation of the intracellular movement of viral proteins of a unique, nonsegmented, negative-strand RNA virus.

scholarly journals Borna disease virus, a negative-strand RNA virus, transcribes in the nucleus of infected cells.

1992 ◽  
Vol 89 (23) ◽  
pp. 11486-11489 ◽  
Author(s):  
T. Briese ◽  
J. C. de la Torre ◽  
A. Lewis ◽  
H. Ludwig ◽  
W. I. Lipkin
Keyword(s):  
Disease Virus ◽  
Rna Virus ◽  
Borna Disease Virus ◽  
Negative Strand ◽  
Infected Cells ◽  
Borna Disease

scholarly journals ADAR2 Is Involved in Self and Nonself Recognition of Borna Disease Virus Genomic RNA in the Nucleus

2019 ◽  
Vol 94 (6) ◽  
Author(s):  
Mako Yanai ◽  
Shohei Kojima ◽  
Madoka Sakai ◽  
Ryo Komorizono ◽  
Keizo Tomonaga ◽  
...  
Keyword(s):  
Immune Responses ◽  
Disease Virus ◽  
Persistent Infection ◽  
Rna Virus ◽  
Innate Immune ◽  
Genomic Rna ◽  
Borna Disease Virus ◽  
Infected Cells ◽  
Nonself Recognition ◽  
Borna Disease

ABSTRACT Cells sense pathogen-derived double-stranded RNA (dsRNA) as nonself. To avoid autoimmune activation by self dsRNA, cells utilize A-to-I editing by adenosine deaminase acting on RNA 1 (ADAR1) to disrupt dsRNA structures. Considering that viruses have evolved to exploit host machinery, A-to-I editing could benefit innate immune evasion by viruses. Borna disease virus (BoDV), a nuclear-replicating RNA virus, may require escape from nonself RNA-sensing and immune responses to establish persistent infection in the nucleus; however, the strategy by which BoDV evades nonself recognition is unclear. Here, we evaluated the involvement of ADARs in BoDV infection. The infection efficiency of BoDV was markedly decreased in both ADAR1 and ADAR2 knockdown cells at the early phase of infection. Microarray analysis using ADAR2 knockdown cells revealed that ADAR2 reduces immune responses even in the absence of infection. Knockdown of ADAR2 but not ADAR1 significantly reduced the spread and titer of BoDV in infected cells. Furthermore, ADAR2 knockout decreased the infection efficiency of BoDV, and overexpression of ADAR2 rescued the reduced infectivity in ADAR2 knockdown cells. However, the growth of influenza A virus, which causes acute infection in the nucleus, was not affected by ADAR2 knockdown. Moreover, ADAR2 bound to BoDV genomic RNA and induced A-to-G mutations in the genomes of persistently infected cells. We finally demonstrated that BoDV produced in ADAR2 knockdown cells induces stronger innate immune responses than those produced in wild-type cells. Taken together, our results suggest that BoDV utilizes ADAR2 to edit its genome to appear as “self” RNA in order to maintain persistent infection in the nucleus. IMPORTANCE Cells use the editing activity of adenosine deaminase acting on RNA proteins (ADARs) to prevent autoimmune responses induced by self dsRNA, but viruses can exploit this process to their advantage. Borna disease virus (BoDV), a nuclear-replicating RNA virus, must escape nonself RNA sensing by the host to establish persistent infection in the nucleus. We evaluated whether BoDV utilizes ADARs to prevent innate immune induction. ADAR2 plays a key role throughout the BoDV life cycle. ADAR2 knockdown reduced A-to-I editing of BoDV genomic RNA, leading to the induction of a strong innate immune response. These data suggest that BoDV exploits ADAR2 to edit nonself genomic RNA to appear as self RNA for innate immune evasion and establishment of persistent infection.

scholarly journals Mutations in the nuclear localization signal of nsP2 influencing RNA synthesis, protein expression and cytotoxicity of Semliki Forest virus

2008 ◽  
Vol 89 (3) ◽  
pp. 676-686 ◽  
Author(s):  
Kristi Tamm ◽  
Andres Merits ◽  
Inga Sarand
Keyword(s):  
Nuclear Localization ◽  
Nuclear Localization Signal ◽  
Rna Synthesis ◽  
Semliki Forest Virus ◽  
Temperature Sensitive ◽  
Structural Protein ◽  
Arginine Residues ◽  
Localization Signal ◽  
Infected Cells ◽  
Replicase Complex

The cytotoxicity of Semliki Forest virus (SFV) infection is caused partly by the non-structural protein nsP2, an essential component of the SFV replicase complex. Due to the presence of a nuclear localization signal (NLS), nsP2 also localizes in the nucleus of infected cells. The present study analysed recombinant SFV replicons and genomes with various deletions or substitutions in the NLS, or with a proline-to-glycine mutation at position 718 of nsP2 (P718G). Deletion of one or two arginine residues from the NLS or substitution of two of the arginines with aspartic acid resulted in a virus with a temperature-sensitive phenotype, and substitution of all three arginines was lethal. Thus, most of the introduced mutations severely affected nsP2 functioning in viral replication; in addition, they inhibited the ability of SFV to induce translational shut-off and kill infected cells. SFV replicons with a P718G mutation or replacement of the NLS residues 648RRR650 with RDD were found to be the least cytotoxic. Corresponding replicons expressed non-structural proteins at normal levels, but had severely reduced genomic RNA synthesis and were virtually unable to replicate and transcribe co-electroporated helper RNA. The non-cytotoxic phenotype was maintained in SFV full-length genomes harbouring the corresponding mutations; however, during a single cycle of cell culture, these were converted to a cytotoxic phenotype, probably due to the accumulation of compensatory mutations.

scholarly journals Borna Disease Virus Phosphoprotein Represses p53-Mediated Transcriptional Activity by Interference with HMGB1

2003 ◽  
Vol 77 (22) ◽  
pp. 12243-12251 ◽  
Author(s):  
Guoqi Zhang ◽  
Takeshi Kobayashi ◽  
Wataru Kamitani ◽  
Satoshi Komoto ◽  
Makiko Yamashita ◽  
...  
Keyword(s):  
Disease Virus ◽  
Transcriptional Activation ◽  
Transcriptional Activity ◽  
Rna Virus ◽  
Broad Host Range ◽  
Neural Cells ◽  
Cellular Functions ◽  
Borna Disease Virus ◽  
Multifunctional Protein ◽  
Borna Disease

ABSTRACT Borna disease virus (BDV) is a noncytolytic, neurotropic RNA virus that has a broad host range in warm-blooded animals, probably including humans. Recently, it was demonstrated that a 24-kDa phosphoprotein (P) of BDV directly binds to a multifunctional protein, amphoterin-HMGB1, and inhibits its function in cultured neural cells (W. Kamitani, Y. Shoya, T. Kobayashi, M. Watanabe, B. J. Lee, G. Zhang, K. Tomonaga, and K. Ikuta, J. Virol. 75:8742-8751, 2001). This observation suggested that expression of BDV P may cause deleterious effects in cellular functions by interference with HMGB1. In this study, we further investigated the significance of the binding between P and HMGB1. We demonstrated that P directly binds to the A-box domain on HMGB1, which is also responsible for interaction with a tumor suppression factor, p53. Recent works have demonstrated that binding between HMGB1 and p53 enhances p53-mediated transcriptional activity. Thus, we examined whether BDV P affects the transcriptional activity of p53 by interference with HMGB1. Mammalian two-hybrid analysis revealed that p53 and P competitively interfere with the binding of each protein to HMGB1 in a p53-deficient cell line, NCI-H1299. In addition, P was able to significantly decrease p53-mediated transcriptional activation of the cyclin G promoter. Furthermore, we showed that activation of p21waf1 expression was repressed in cyclosporine-treated BDV-infected cells, as well as p53-transduced NCI-H1299 cells. These results suggested that BDV P may be a unique inhibitor of p53 activity via binding to HMGB1.

scholarly journals 1-β-d-Arabinofuranosylcytosine Inhibits Borna Disease Virus Replication and Spread

2002 ◽  
Vol 76 (12) ◽  
pp. 6268-6276 ◽  
Author(s):  
Jeffrey J. Bajramovic ◽  
Sylvie Syan ◽  
Michel Brahic ◽  
Juan Carlos de la Torre ◽  
Daniel Gonzalez-Dunia
Keyword(s):  
Disease Virus ◽  
Measles Virus ◽  
Rna Virus ◽  
Neurological Diseases ◽  
Borna Disease Virus ◽  
Negative Strand ◽  
Neuropsychiatric Diseases ◽  
Borna Disease

ABSTRACT Borna disease virus (BDV) is a nonsegmented, negative-strand RNA virus that causes neurological diseases in a variety of warm-blooded animal species. There is general consensus that BDV can also infect humans, being a possible zoonosis. Although the clinical consequences of human BDV infection are still controversial, experimental BDV infection is a well-described model for human neuropsychiatric diseases. To date, there is no effective treatment against BDV. In this paper, we demonstrate that the nucleoside analog 1-β-d-arabinofuranosylcytosine (Ara-C), a known inhibitor of DNA polymerases, inhibits BDV replication. Ara-C treatment inhibited BDV RNA and protein synthesis and prevented BDV cell-to-cell spread in vitro. Replication of other negative-strand RNA viruses such as influenza virus or measles virus was not inhibited by Ara-C, underscoring the particularity of the replication machinery of BDV. Strikingly, Ara-C treatment induced nuclear retention of viral ribonucleoparticles. These findings could not be attributed to known effects of Ara-C on the host cell, suggesting that Ara-C directly inhibits the BDV polymerase. Finally, we show that Ara-C inhibits BDV replication in vivo in the brain of infected rats, preventing persistent infection of the central nervous system as well as the development of clinical disease. These findings open the way to the development of effective antiviral therapy against BDV.

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