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Suresh K. Tikoo - One of the best experts on this subject based on the ideXlab platform.

  • Bovine Adenovirus type 3 virions cannot be rescued in vivo after full length viral genome transfection in the absence of detectable polypeptide ix
    Journal of Veterinary Science, 2017
    Co-Authors: Peng Zhang, Qinghong Xue, Jingjing Ren, Shuili Xia, Lei Zhang, Wenbin Wang, Suresh K. Tikoo
    Abstract:

    Bovine Adenovirus type 3 (BAdV3) is being used in the development of potential vehicles for gene therapy and vectored vaccine. To that end, a more comprehensive description of BAdV3 biology is essential. In this study, we focused on the role of pIX in BAdV3 virion rescue after full-length BAdV3 genome transfection. Initially, pIX deletion or initiation codon mutation abolished the production of progeny virions, which suggested that pIX was essential for the rescue of BAdV3 containing a full-length genome. Moreover, through transfection of a panel of pIX mutant BAdV3 genomes, we observed that the conserved N-terminus and the putative leucine zipper element (PLZP) were essential for virion rescue, whereas the C-terminus following the coiled-coil domain was non-essential. In addition, swap of the PLZP element and its following region of BAdV3 pIX to corresponding domains of human Adenovirus type 5 (HAdV5) did not affect virion production, whereas swap of the entire pIX abolished production of progeny virions. We suggest that failure of the full-length BAdV3 pIX swap might be due to species specificity of its N-terminus region before the PLZP element.

  • Leucine residues in conserved region of 33K protein of Bovine Adenovirus – 3 are important for binding to major late promoter and activation of late gene expression
    Virology, 2015
    Co-Authors: Vikas Kulshreshtha, Lisanework E. Ayalew, Azharul Islam, Suresh K. Tikoo
    Abstract:

    Abstract The L6 region of Bovine Adenovirus 3 (BAdV-3) encode 33K (spliced) and 22K (unspliced) proteins. Earlier, anti-33K serum detected five major and three minor proteins in BAdV-3 infected cells. Here, we demonstrate that anti-sera raised against L6-22K protein detected two proteins of 42 and 37 kDa in BAdV-3 infected cells and one protein of 42 kDa in transfected cells expressing splice-site variant 22K protein (pC.22K containing substituted splice acceptor/donor sequence). Unlike 22K, 33K stimulated the transcription from the major late promoter (MLP) by binding to the downstream sequence elements (DE). Analysis of the variant proteins demonstrated that amino acids 201–240 of the conserved C-terminus of 33K containing the potential leucine zipper and RS repeat are required for the activation of MLP. Furthermore, amino acid substitution analysis demonstrated that unlike arginine residues of RS repeat, the leucine residues (217, 224, 232 and 240) of the conserved leucine zipper appear required for the binding of 33K to the MLP.

  • Conserved Arginines of Bovine Adenovirus-3 33K Protein Are Important for Transportin-3 Mediated Transport and Virus Replication
    2014
    Co-Authors: Vikas Kulshreshtha, Lisanework E. Ayalew, Azharul Islam, Suresh K. Tikoo
    Abstract:

    The L6 region of Bovine Adenovirus (BAdV)-3 encodes a spliced protein designated 33K. The 33K specific sera detected five major proteins and three minor proteins in transfected or virus infected cells, which could arise by internal initiation of translation and alternative splicing. The 33K protein is predominantly localized to the nucleus of BAdV-3 infected cells. The 33K nuclear transport utilizes both classical importin-a/-b and importin-b dependent nuclear import pathways and preferentially binds to importin-a5 and transportin-3 receptors, respectively. Analysis of mutant 33K proteins demonstrated that amino acids 201–240 of the conserved C-terminus of 33K containing RS repeat are required for nuclear localization and, binding to both importin-a5 and transportin-3 receptors. Interestingly, the arginine residues of conserved RS repeat are required for binding to transportin-3 receptor but not to importin-a5 receptor. Moreover, mutation of arginines residues of RS repeat proved lethal for production of progeny virus. Our results suggest that arginines of RS repeat are required for efficient nuclear transport of 33K mediated by transportin-3, which appears to be essential for replication and production of infectious virion

  • Bovine Adenovirus type 3 e1b small protein is essential for growth in Bovine fibroblast cells
    Virology, 2001
    Co-Authors: Yan Zhou, Lorne A. Babiuk, P S Reddy, Suresh K. Tikoo
    Abstract:

    In order to study the function of Bovine Adenovirus type 3 (BAV-3) E1A and E1B(small) proteins, we constructed two mutants: (a) BAV102A carries an in-frame deletion in the coding region for the E1A protein (nt 831-1080); (b) BAV102B carries an insertion of triple stop codons in the E1B region (nt 1654, 178 bp downstream of the E1B(small) start codon), which stops the translation of the E1B(small) gene. BAV102A virus could grow to the wild-type BAV-3 titer in transformed cell line VIDO R2 (HAV-5 E1 transformed) cells, but no progeny virus could be found in fetal Bovine retina cells (FBRC). RT-PCR and Western blot analysis showed that neither mRNA transcripts nor protein expression of early genes [E1B(small) and DNA binding protein (DBP)] could be detected in BAV102A infected FBRC. The BAV102B grew 1.5 log less than wild-type BAV-3 in FBRC; however, no BAV102B progeny virus could be observed in Bovine fibroblast (BFB) cells. No appreciable difference was observed in DBP transcript synthesis between wild-type BAV-3- or BAV102B-infected FBRC. However, compared to wild-type BAV-3, BAV102B viral DNA synthesis and fiber gene expression were found to be slightly reduced in FBRC. In contrast, compared to wild-type BAV-3, DBP transcripts and viral DNA synthesis were drastically reduced in BAV102B-infected BFB cells. In addition, no fiber gene expression could be detected in BAV102B-infected BFB cells. These results suggest that BAV-3 E1A is essential for virus replication and is required for activating the transcription of other BAV-3 early genes. However, the requirement for E1B(small) protein for BAV-3 replication appears to be cell type-dependent.

  • optimization of Bovine coronavirus hemagglutinin estrase glycoprotein expression in e3 deleted Bovine Adenovirus 3
    Virus Research, 2000
    Co-Authors: Seshidhar P Reddy, Lorne A. Babiuk, Majid Mehtali, Neeraja Idamakanti, Lexander N Zakhartchouk, Suresh K. Tikoo
    Abstract:

    Adenoviral vectors expressing foreign genes have many desirable properties in applications such as vaccination. Recently, we have generated replication-competent (E3 deleted) Bovine Adenovirus-3 (BAV-3) recombinants expressing significant amounts of glycoprotein D (gD) of Bovine herpesvirus-1 (a DNA virus). However, attempts to express the RNA virus genes using the same strategy were not successful. In an effort to optimize the expression, we have constructed several BAV-3 recombinants carrying the hemagglutinin esterase (HE) gene of Bovine coronavirus (BCV) in the E3 region with or without exogenous transcription control elements. The expression studies suggest that the introduction of a 137 bp chimeric intron upstream of the HE cDNA is able to increase the level of HE gene expression. The introduction of a SV40 early promoter or human cytomegalovirus (HCMV) immediate early (IE) promoter into the expression cassette changed the kinetics of the HE expression. However, the recombinant BAV-3 containing HE under the HCMV IE promoter replicated less efficiently than the wild-type BAV-3. These studies should prove useful in expression of other RNA viral genes in the E3 region of BAV-3 expression system.

Suresh K. Mittal - One of the best experts on this subject based on the ideXlab platform.

  • Efficiency of Airborne Sample Analysis Platform (ASAP) Bioaerosol Sampler for Pathogen Detection
    Frontiers Media S.A., 2015
    Co-Authors: Anurag Esharma, Elizabeth Eclark, James D Mcglothlin, Suresh K. Mittal
    Abstract:

    The threat of bioterrorism and pandemics has highlighted the urgency for rapid and reliable bioaerosol detection in different environments. Safeguarding against such threats requires continuous sampling of the ambient air for pathogen detection. In this study we investigated the efficacy of the Airborne Sample Analysis Platform (ASAP) 2800 bioaerosol sampler to collect representative samples of air and identify specific viruses suspended as bioaerosols. To test this concept, we aerosolized an innocuous replication-defective Bovine Adenovirus serotype 3 (BAdV3) in a controlled laboratory environment. The ASAP efficiently trapped the surrogate virus at 5×10E3 plaque-forming units (p.f.u.) [2×10E5 genome copy equivalent] concentrations or more resulting in the successful detection of the virus using quantitative PCR. These results support the further development of ASAP for bioaerosol pathogen detection

  • Bovine Adenovirus serotype 3 utilizes sialic acid as a cellular receptor for virus entry.
    Virology, 2009
    Co-Authors: Dinesh S Bangari, Anurag Sharma, Suresh K. Mittal
    Abstract:

    Bovine Adenovirus serotype 3 (BAd3) and porcine Adenovirus serotype 3 (PAd3) entry into the host cells is independent of Coxsackievirus Adenovirus receptor and integrins. The role of sialic acid in BAd3 and PAd3 entry was investigated. Removal of sialic acid by neuraminidase, or blocking sialic acid by wheat germ agglutinin lectin significantly inhibited BAd3, but not PAd3, transduction of Madin-Darby Bovine kidney cells. Maackia amurensis agglutinin or Sambucus nigra (elder) agglutinin treatment efficiently blocked BAd3 transduction suggesting that BAd3 utilized alpha(2,3)-linked and alpha(2,6)-linked sialic acid as a cell receptor. BAd3 transduction of MDBK cells was sensitive to sodium periodate, bromelain, or trypsin treatment indicating that the receptor sialoconjugate was a glycoprotein rather than a ganglioside. To determine sialic acid-containing cell membrane proteins that bind to BAd3, virus overlay protein binding assay (VOPBA) was performed and showed that sialylated cell membrane proteins in size of approximately 97 and 34 kDa bind to BAd3. The results suggest that sialic acid serves as a primary receptor for BAd3.

  • Comparative analysis of vector biodistribution, persistence and gene expression following intravenous delivery of Bovine, porcine and human adenoviral vectors in a mouse model.
    Virology, 2009
    Co-Authors: Anurag Sharma, Dinesh S Bangari, Harm Hogenesch, Manish Tandon, Aseem Pandey, Suresh K. Mittal
    Abstract:

    Nonhuman Adenoviruses including Bovine Adenovirus serotype 3 (BAd3) and porcine Adenovirus serotype 3 (PAd3) can circumvent pre-existing immunity against human Adenovirus serotype 5 (HAd5) and are being developed as alternative vectors for gene delivery. To assess the usefulness of these vectors for in vivo gene delivery, we compared biodistribution, persistence, state of vector genome, and transgene and vector gene expression by replication-defective BAd3 and PAd3 vectors with those of HAd5 vector in a FVB/n mouse model following intravenous inoculation. BAd3 vector efficiently transduced the heart, kidney and lung in addition to the liver and spleen and persisted for a longer duration compared to PAd3 or HAd5 vectors. Biodistribution of PAd3 vector was comparable to that of HAd5 vector but showed more rapid vector clearance. Only linear episomal forms of BAd3, PAd3, and HAd5 vector genomes were detected. All three vectors efficiently expressed the green fluorescent protein (GFP) transgene proportionate to the vector genome copy number in various tissues. Furthermore, leaky expression of vector genes, both the early (E4) and the late (hexon) was observed in all three vectors and gradually declined with time. These results suggest that BAd3 and PAd3 vectors could serve as an alternative or supplement to HAd5 for gene delivery applications.

  • Bovine Adenovirus type 3 internalization is independent of primary receptors of human Adenovirus type 5 and porcine Adenovirus type 3.
    Biochemical and biophysical research communications, 2005
    Co-Authors: Dinesh S Bangari, Anurag Sharma, Suresh K. Mittal
    Abstract:

    Abstract Usefulness of adenoviral vectors derived from human Adenovirus (HAd) type 5 (HAd5) is mainly limited by wide prevalence of preexisting anti-HAd5 immunity as well as non-specific tissue tropism of these vectors. As an alternative, non-human adenoviral vectors including Bovine Adenovirus type 3 (BAd3) are currently being investigated. Non-prevalence of BAd3 in humans and its ability to evade preexisting HAd immunity are some of the features that make BAd3 a promising vector for human gene delivery. BAd3 appears to have a tissue tropism distinct from that of HAd5 and also the repertoire of cells efficiently transduced by BAd3 is different. We performed antibody-mediated receptor blocking experiments to show that BAd3 internalization was independent of coxsackievirus–Adenovirus receptor, the primary determinant of HAd5 tropism, or integrin ανβ3, a secondary molecule involved in HAd5 entry. Using homologous and heterologous knob-mediated competition assays with recombinant knobs of HAd5, porcine Adenovirus type 3 (PAd3), or BAd3, we observed that BAd3 internalization was independent of the primary receptors of HAd5 and PAd3. These results provide support for further exploration of BAd3 vectors for designing targeted vectors for human gene therapy.

  • a 72 bp internal deletion in the left inverted terminal repeat of the Bovine Adenovirus type 3 genome does not affect virus replication
    Intervirology, 2002
    Co-Authors: Alberto Van Olphen, Suresh K. Mittal
    Abstract:

    The genome of Bovine Adenovirus type 3 (BAV3) is flanked by 195-base pair (bp) inverted terminal repeats (ITR). We isolated a BAV3 mutant (BAV3c29) having an internal deletion within the left ITR. The deletion eliminated 72 bp between nucleotides (nt) 89 and 162, including most of the GC-rich sequences located close to the end of the ITR sequences. This deletion did not seem to have any affect on the virus plaque size or morphology and the kinetics of viral replication compared to wild-type (wt) BAV3. The nt sequence of the right ITR of BAV3c29 remained identical to the right or left ITR of wt BAV3. These results indicate that the cis-acting sequences present within the 72 bp between nt 89 and 162 of the left ITR are not essential for BAV3 DNA replication in cultured cells.

Lorne A. Babiuk - One of the best experts on this subject based on the ideXlab platform.

  • Bovine Adenovirus type 3 e1b small protein is essential for growth in Bovine fibroblast cells
    Virology, 2001
    Co-Authors: Yan Zhou, Lorne A. Babiuk, P S Reddy, Suresh K. Tikoo
    Abstract:

    In order to study the function of Bovine Adenovirus type 3 (BAV-3) E1A and E1B(small) proteins, we constructed two mutants: (a) BAV102A carries an in-frame deletion in the coding region for the E1A protein (nt 831-1080); (b) BAV102B carries an insertion of triple stop codons in the E1B region (nt 1654, 178 bp downstream of the E1B(small) start codon), which stops the translation of the E1B(small) gene. BAV102A virus could grow to the wild-type BAV-3 titer in transformed cell line VIDO R2 (HAV-5 E1 transformed) cells, but no progeny virus could be found in fetal Bovine retina cells (FBRC). RT-PCR and Western blot analysis showed that neither mRNA transcripts nor protein expression of early genes [E1B(small) and DNA binding protein (DBP)] could be detected in BAV102A infected FBRC. The BAV102B grew 1.5 log less than wild-type BAV-3 in FBRC; however, no BAV102B progeny virus could be observed in Bovine fibroblast (BFB) cells. No appreciable difference was observed in DBP transcript synthesis between wild-type BAV-3- or BAV102B-infected FBRC. However, compared to wild-type BAV-3, BAV102B viral DNA synthesis and fiber gene expression were found to be slightly reduced in FBRC. In contrast, compared to wild-type BAV-3, DBP transcripts and viral DNA synthesis were drastically reduced in BAV102B-infected BFB cells. In addition, no fiber gene expression could be detected in BAV102B-infected BFB cells. These results suggest that BAV-3 E1A is essential for virus replication and is required for activating the transcription of other BAV-3 early genes. However, the requirement for E1B(small) protein for BAV-3 replication appears to be cell type-dependent.

  • optimization of Bovine coronavirus hemagglutinin estrase glycoprotein expression in e3 deleted Bovine Adenovirus 3
    Virus Research, 2000
    Co-Authors: Seshidhar P Reddy, Lorne A. Babiuk, Majid Mehtali, Neeraja Idamakanti, Lexander N Zakhartchouk, Suresh K. Tikoo
    Abstract:

    Adenoviral vectors expressing foreign genes have many desirable properties in applications such as vaccination. Recently, we have generated replication-competent (E3 deleted) Bovine Adenovirus-3 (BAV-3) recombinants expressing significant amounts of glycoprotein D (gD) of Bovine herpesvirus-1 (a DNA virus). However, attempts to express the RNA virus genes using the same strategy were not successful. In an effort to optimize the expression, we have constructed several BAV-3 recombinants carrying the hemagglutinin esterase (HE) gene of Bovine coronavirus (BCV) in the E3 region with or without exogenous transcription control elements. The expression studies suggest that the introduction of a 137 bp chimeric intron upstream of the HE cDNA is able to increase the level of HE gene expression. The introduction of a SV40 early promoter or human cytomegalovirus (HCMV) immediate early (IE) promoter into the expression cassette changed the kinetics of the HE expression. However, the recombinant BAV-3 containing HE under the HCMV IE promoter replicated less efficiently than the wild-type BAV-3. These studies should prove useful in expression of other RNA viral genes in the E3 region of BAV-3 expression system.

  • Transcription map and expression of Bovine herpesvirus-1 glycoprotein D in early region 4 of Bovine Adenovirus-3.
    Virology, 1999
    Co-Authors: Mohit K. Baxi, Lorne A. Babiuk, Majid Mehtali, Suresh K. Tikoo
    Abstract:

    Abstract Early region 4 (E4) of Bovine Adenovirus type 3 (BAV-3) was analyzed by Northern blotting, RT–PCR analysis, cDNA sequencing, and S1 nuclease protection assays. The transcriptional map of the E4 region of BAV-3 has marked dissimilarities from those of mouse Adenovirus-1, ovine Adenovirus-287, and human Adenovirus-2, for which the transcriptional maps have been constructed. The E4 region of BAV-3, located between 98.6 and 89.8 MU transcribes seven distinct classes of Bovine Adenovirus type 3 mRNA. The seven mRNA species formed by the removal of one to three introns share both the 3′ end and a short 5′ leader (25 nucleotides). The E4 mRNAs can encode at least five unique polypeptides, namely, 143R1, 69R, 143R2, 268R, and 219R. Isolation of a replication-competent recombinant “BAV404” containing 1.9-kb insertion [glycoprotein (gD) of Bovine herpesvirus 1, under the control of a SV40 early promoter and poly(A)] in the region between E4 and the right ITR suggested that this region is nonessential for BAV-3 replication. Expression of gD by BAV404 recombinant virus was confirmed by immunoprecipitation with gD-specific monoclonal antibodies. Analysis of the kinetics of protein expression indicated that gD is expressed at both early and late times postinfection. These results suggest that: (a) E4 produces seven 5′–3′ coterminal mRNAs and (b) the right terminal region of BAV-3 can be used for the expression of vaccine antigens.

  • Characterization of early region 1 and pIX of Bovine Adenovirus-3.
    Virology, 1999
    Co-Authors: P. Seshidhar Reddy, Lorne A. Babiuk, Neeraja Idamakanti, Caron Pyne, Yan Chen, Suresh K. Tikoo
    Abstract:

    The mRNAs from early region 1 (E1) and pIX of Bovine Adenovirus type 3 (BAV-3) have been studied by Northern blot, S1 nuclease, and cDNA analysis and transcriptional maps for the regions were constructed. The transcriptional map for the E1 region of BAV-3 is different from those of mouse and human Adenoviruses for which transcriptional maps for the regions have been constructed. The E1A region of BAV-3 is located between 0.8 and 10.5 map units and several different transcripts are produced from the region using alternative splice donor sites. The transcripts from the E1A region overlap with those of E1B and pIX. In BAV-3, the E1B region maps between 4.2 and 10.5 map units and encodes two major mRNA species. The mRNAs of E1B region differ from each other in that the smaller mRNA coding for the 157R protein has a large intron removed from a region corresponding to the coding region of E1B 420R protein. As in HAVs, the E1B 420R protein of BAV-3 could be translated only by internal initiation from the larger bicistronic mRNA as there are no transcripts produced exclusively for the production of 420R protein. The transcriptional unit of pIX is transcribed from an independent promoter and encodes a structural component of the Adenovirus capsid. To identify and characterize the proteins produced from the region, antibodies were raised in rabbits that recognized specific proteins in Western blot and immunoprecipitation assays.

  • nucleotide sequence genome organization and transcription map of Bovine Adenovirus type 3
    Journal of Virology, 1998
    Co-Authors: Seshidhar P Reddy, Lorne A. Babiuk, Mohit K. Baxi, Neeraja Idamakanti, Alexandre N Zakhartchouk, Joong Bok Lee, Caron Pyne, Suresh K. Tikoo
    Abstract:

    The complete DNA sequence of Bovine Adenovirus type 3 is reported here. The size of the genome is 34,446 bp in length with a G+C content of 54%. All the genes of the early and late regions are present in the expected locations of the genome. However, the late-region genes are organized into seven families, instead of five as they are in human Adenovirus type 2. The deduced amino acid sequences of open reading frames (ORFs) in the late regions and early region 2 (E2) and for IVa2 show higher degrees of homology, whereas the predicted amino acid sequences of ORFs in the E1, E3, and E4 regions and the pIX, fiber, and 33,000-molecular-weight nonstructural proteins show little or no homology with the corresponding proteins of other Adenoviruses. In addition, the penton base protein lacks the integrin binding motif, RGD, but has an LDV motif instead of an MDV motif. Interestingly, as in other animal Adenoviruses, the virus-associated RNA genes appear to be absent from their usual location. Sequence analysis of cDNA clones representing the early- and late-region genes identified splice acceptor and splice donor sites, polyadenylation signals and polyadenylation sites, and tripartite leader sequences.

Ludvik Prevec - One of the best experts on this subject based on the ideXlab platform.

  • Transcription units of E1a, E1b and pIX regions of Bovine Adenovirus type 3
    Journal of General Virology, 1999
    Co-Authors: B J Zheng, Frank L Graham, Ludvik Prevec
    Abstract:

    The major mRNA species in the E1 region of the genome of Bovine Adenovirus type 3 (BAV3) have been defined by using a combination of PCR, 5' RACE, Northern analysis and DNA sequencing. Independent transcription initiation sites were identified for each of the E1a, E1b and protein IX (pIX) transcription units, but all mRNA species terminated at the same poly(A) addition site immediately downstream of the pIX open reading frame. Thus, the BAV3 E1 region, which consists of the E1a and E1b genes together with that for pIX, functions as a nested overlapping transcription unit. One major mRNA species encoding the E1a protein was found and two mRNAs encoding E1b species, the smaller of which encodes the E1b 17K protein alone and the larger encodes both 17K and 47K E1b proteins, were identified. One mRNA species encodes pIX. The E1a transcript, encoding the predicted 214 residue E1a protein, has four exons. The smaller E1b mRNA has two exons, the second of which corresponds to the last exon of E1a. No introns were detected in the larger E1b mRNA that encodes both the E1b 17K and 47K proteins nor in the mRNA encoding pIX. The relative times of appearance of the mRNAs from the E1-pIX gene region following infection of Bovine cells with BAV3 was determined.

  • pathology and immunogenicity in the cotton rat sigmodon hispidus model after infection with a Bovine Adenovirus type 3 recombinant virus expressing the firefly luciferase gene
    Journal of General Virology, 1996
    Co-Authors: Suresh K. Mittal, Dorothy M Middleton, Suresh K. Tikoo, Ludvik Prevec, Frank L Graham, Lorne A. Babiuk
    Abstract:

    The histopathology of Adenovirus pneumonia in cotton rats (Sigmodon hispidus) due to Bovine Adenovirus type 3-luciferase recombinant virus (BAd3-Luc), which has a 0.7 kb deletion from the early region 3 (E3) replaced with the firefly luciferase gene, was compared with that produced by the parental wild-type (wt) Bovine Adenovirus type 3 (BAd3). After intranasal inoculation of cotton rats with 3 × 107 p.f.u. of BAd3-Luc, the infectious virus titres in the lungs at various times post-infection were similar to those of animals infected with the parental virus. Quantitative analysis of histopathological changes and immunohistochemical staining showed that the character and severity of the lesions were indistinguishable in the two infections. Luciferase activity was detected in the lungs of BAd3-Lucinoculated animals until 4 days post-infection (p.i.). Antibodies to both BAd3 and luciferase were detected in sera collected from BAd3-Luc-infected animals until at least 6 weeks p.i. These results show that BAd3-Luc produces pulmonary lesions in cotton rats similar to those of wt BAd3 and suggest that BAd3-based vectors may be suitable for the development of live recombinant virus vaccines.

  • the e1 sequence of Bovine Adenovirus type 3 and complementation of human Adenovirus type 5 e1a function in Bovine cells
    Virus Research, 1994
    Co-Authors: Bojian Zheng, Suresh K. Mittal, Frank L Graham, Ludvik Prevec
    Abstract:

    The Bovine Adenovirus type 3 (BAV3) genome was sequenced from the left end to the HindIII site at 11%. This region comprises the entire E1 transcription unit including the open reading frames (ORF) for proteins homologous to the E1A, E1B proteins and protein IX of human Adenovirus type 5 (Ad5). A portion of the BAV3 E1A protein showed significant homology with conserved region 3 (CR3), the principal transactivation region of Ad5 E1A. The BAV3 E1A protein also contains a consensus sequence known to be important for interaction with the cellular Rb protein but lacks most of the sequence corresponding to the second exon of Ad5 E1A. Promoter sequences for BAV3 E1B were not defined though the relevant region contains a 35-base pair repeat sequence. Two ORFs define the BAV3 E1B coding unit; one with regions homologous to sequences within the Ad5 E1B 19k protein, and an overlapping ORF with significant homology to the Ad5 E1B 55k protein. The encoded BAV3 E1B proteins of 157 and 420 amino acid residues (R) have predicted unmodified molecular weights of 17,393 and 46,734 respectively. Immediately following the E1B coding region there is a transcription unit containing an SP1 binding site and TATA box followed by an ORF which encodes a protein of 125R and predicted molecular weight of 13,706 with homology to protein IX of Ad5. Five concensus poly A addition sites are located in the 350 base pairs immediately following the protein IX coding region. The homology of sequences in the Ad5 E1A CR3 region and the corresponding BAV3 protein suggested that the BAV3 protein could transactivate certain Ad5 genes normally transactivated by the Ad5 E1A product. Evidence for this hypothesis was obtained in studies in which Bovine cells in culture were coinfected with BAV3 and a human Adenovirus type 5 (Ad5) recombinant viral vector lacking the E1A region and having a lacZ reporter gene within the E3 region dependent on E1A for its expression. Coinfection resulted in the induction of beta-galactosidase activity and the increased expression of other Ad5 early (E2A 72k) and late (hexon) proteins.

Balazs Harrach - One of the best experts on this subject based on the ideXlab platform.

  • Crystal structure of the fibre head domain of Bovine Adenovirus 4, a ruminant atAdenovirus.
    Virology Journal, 2015
    Co-Authors: Thanh H. Nguyen, Balazs Harrach, Mónika Z. Ballmann, Maria Benkő, Márton Z. Vidovszky, Marta Sanz-gaitero, Abhimanyu K. Singh, Mark J. Van Raaij
    Abstract:

    Background In Adenoviruses, primary host cell recognition is generally performed by the head domains of their homo-trimeric fibre proteins. This first interaction is reversible. A secondary, irreversible interaction subsequently takes place via other Adenovirus capsid proteins and leads to a productive infection. Although many fibre head structures are known for human mastAdenoviruses, not many animal Adenovirus fibre head structures have been determined, especially not from those belonging to Adenovirus genera other than MastAdenovirus.

  • dna sequencing and analysis of the right hand part of the genome of the unique Bovine Adenovirus type 10
    Journal of General Virology, 2004
    Co-Authors: Krisztina Ursu, Balazs Harrach, Katalin Matiz, Maria Benkő
    Abstract:

    The prototype strain of Bovine Adenovirus (BAdV) type 10 and four additional isolates that were indistinguishable in serum-neutralization tests have been shown to have remarkable variation in their genome size and restriction maps. In the present study, more than 40 % of the DNA sequence of the BAdV-10 isolate with the longest genome was determined. A biased base composition resulting in low (<41 %) GC content was noticed. Analysis of the genes of the DNA-binding protein, 100K, 33K, pVIII and fibre proteins, as well as early regions E3 and E4, which are encoded by the genome fragment examined, confirmed that BAdV-10 is different from the other known BAdV types regarding its phylogenetic distance and the organization of its exceptionally short E3 region, apparently containing only two genes. A comparative analysis of the E3 and E4 regions of BAdV-10 with various animal Adenoviruses revealed interesting features accounting for the very short genome of BAdV-10. In the examined BAdV-10 isolate, duplicated sequences were localized in and around the fibre gene. Since BAdV-10 appears to be pathogenic to cattle and is genetically distant from the other BAdVs, we suggest that BAdV-10 is not a genuine Bovine virus, but has recently switched host and is now undergoing an adaptation process in its new host. In accordance with this hypothesis, the remarkable predominance of AT-rich codons along with the variable fibre gene might be signs of adaptation.

  • four new inverted terminal repeat sequences from Bovine Adenoviruses reveal striking differences in the length and content of the itrs
    Virus Genes, 2001
    Co-Authors: Balazs Harrach, Zoltan Zadori, Maria Benkő
    Abstract:

    The inverted terminal repeat (ITR) of the genome of four Bovine Adenovirus (BAdV) types have been sequenced, analysed and compared to the ITRs of other Adenoviruses. The length of ITRs of the examined BAdVs ranged between 59 and 368 base pairs, thus the presently known longest Adenovirus ITR sequence is from BAdV-10. The conserved motifs and characteristic sequence elements of the ITRs providing different binding sites for replicative proteins of viral and cellular origin seemed to be distributed according to the proposed genus classification of BAdVs. The ITRs of BAdV-10 share similarity with the members of the genus MastAdenovirus, while the ITRs of the other three sequenced serotypes (BAdV-4, 5 and strain Rus) which are candidate members of the genus AtAdenovirus are very short and contain NFI and Sp1 binding sites only. The analysis of the new ITRs implied that the nucleotide sequence of the so-called core origin is highly preserved within the mastAdenovirus genus only.

  • Analysis of the hexon gene sequence of Bovine Adenovirus type 4 provides further support for a new Adenovirus genus (AtAdenovirus).
    Journal of General Virology, 1998
    Co-Authors: Zsolt Ruzsics, Mária Benkö, W. C. Russell, Balazs Harrach
    Abstract:

    The putative hexon gene of Bovine Adenovirus type 4 (BAV-4), encoding 910 amino acid residues, has been identified and sequenced. A characteristic codon usage biased towards the use of AT-rich triplets was observed. Comparative analysis with other hexon sequences detected a high level of amino acid identity in the regions corresponding to the pedestals of the hexon. Substitutions, insertions and deletions were identified mainly in the variable regions forming the loops which are exposed on the outer surface of the virion. In these variable regions, BAV-4 shared similarity only with egg drop syndrome (EDS) virus and ovine Adenovirus isolate 287 (OAV287). The close relationship of these viruses was also demonstrated by phylogenetic analysis of the hexon gene. In addition to the two groups of the MastAdenovirus and AviAdenovirus genera, a third cluster appeared comprising BAV-4, OAV287 and EDS virus.

  • Close phylogenetic relationship between egg drop syndrome virus, Bovine Adenovirus serotype 7, and ovine Adenovirus strain 287.
    Virology, 1997
    Co-Authors: Balazs Harrach, Mária Benkö, B. M. Adair, Brian Meehan, Daniel Todd
    Abstract:

    Abstract A cloned egg drop syndrome (EDS) virus genomic DNA fragment containing the protease gene has been identified and the complete nucleotide sequence of the protease and partial nucleotide sequence of the hexon genes has been determined. Phylogenetic analysis of the protease gene has revealed EDS virus to be genetically more closely related to Bovine Adenovirus type 7 (BAV-7) and ovine Adenovirus isolate 287 (OAV287) than either of these two viruses are to other members of the genus MastAdenovirus or EDS virus is to an other member of the AviAdenovirus genus. The three viruses share further similarities in that they have a high percentage AT content in their genome and are characterized by having more compact genomes than other Adenoviruses. The protease gene from all three viruses contained the active site residues (H55–D72–C122 triad) and C104 (providing a disulfide bond to cofactor pVIc). However, P137, found in all other members of the MastAdenovirus genus, and thought to be involved in trafficking, was missing from the protease of the EDS virus, BAV-7, and OAV287. These results suggest that EDS virus should be classified along with BAV-7 and OAV287 in a separate taxon.