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PubMed1979

Adenovirus type 2 early polypeptides immunoprecipitated by antisera to five lines of adenovirus-transformed rat cells.

We have identified adenovirus type 2 (Ad2)-induced early polypeptides (EPs) and have attempted to determine which EPs are coded by each of the four early gene blocks. [35S]methionine-labeled EPs were resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Cycloheximide pretreatment followed by labeling in hypertonic medium (210 to 250 mM NaCl) facilitated the detection of EPs. Seven major (reproducible bands in autoradiograms) EPs were detected with molecular weights of 74,000 (74K), 21K, 19K, 15K, 13.5K, 11.5K, and 11K. Minor (weaker bands) EPs of 55K, 52K, 42K, 18K, 12K, 8.8K, and 8.3K were also often seen. To identify and map the genes for virus-coded EPs, we prepared antisera against five lines of adenovirus-transformed cells that retain different fractions of the viral genome. The lines were F17, 8617, F4, and T2C4 transformed by Ad2 virions and 5RK (clone I) transformed by transfection with the Ad5 HsuI-G fragment (map position 0 to 8). The early gene blocks retained and expressed (in part) as RNA in these cells were as follows: 5RK(I), block 1 (70% of left 8% of genome); F17, block 1; 8617, blocks 1 and 4; F4 blocks 1, 2, and 4; T2C4, blocks 1, 2, 3, and 4. The following major EPs were immunoprecipitated: 15K by all antisera; 53K and 14.5K by F17, T2C4, 8617, and F4 antisera; 11.5K by T2C4, 8617, and F4 antisera; 44K, 42K, 19K, and 13.5K by T2C4 antisera; 11K by 8617 antisera. Minor EPs of 28K, 18K, and 12K were precipitated by all antisera except 5RK(I). The 53K and 15K EPs were precipitated also from Ad2 early infected monkey cells by the F17 antiserum and by sera from hamsters bearing tumors induced by Ad1-simian virus 40. The relationships between some of the immunoprecipitated EPs were investigated by the partial proteolysis procedure. All 53K EPs are the "same" (i.e., highly related), all 15K EPs are the "same," and all 11.5K EPs are the "same." The 15K EP is highly related to the 14.5 K EP. Although less certain, all 28K EPs appeared related, as did all 18K EPs. The T2C4-specific 44K EP is probably a dimer of the 21K glycopolypeptide. The T2C4-specific 13.5K EP and the 8617-specific 11K EP appear unrelated to any other polypeptides. These immunoprecipitation data provide evidence that early gene block I (map position 1 to 11) may encode major 53K, 15K, and 14.5K polypeptides, and minor 28K, 18K, and 12K polypeptides, and that all or some of the gene for 15K and 14.5K lies within map position 1 to 8. The surprisingly complex pattern of polypeptides coded by early gene block I raises the possibility that some polypeptides may be coded by overlapping "spliced" mRNA's. The possible block locations of the genes for the 21K, 13.5K, and 11.5K polypeptides are discussed.

Adenoviruses, Human↗
PubMed1981

In vitro synthesis of adenovirus type 5 T antigens. II. Translation of virus-specific RNA from cells transformed by fragments of adenovirus type 5 DNA.

Virus-specific cytoplasmic RNA was isolated from rat cell lines transformed by fragments of adenovirus type 5 DNA, and the RNAs were translated in cell-free systems derived from wheat germ or rabbit reticulocytes. RNA was isolated from cell lines transformed by the following fragments: XhoI-C (leftmost 15.5%), HindIII-G (leftmost 8%), and HpaI-E (leftmost 4.5%). In addition, the adenovirus type 5-transformed human embryonic kidney line 293.C31 was investigated. The products were immunoprecipitated with serum from tumor-bearing hamsters and analyzed by electrophoresis in sodium dodecyl sulfate-polyacrylamide gels. The results show that all transformed cells investigated contain early region 1a-specific RNAs which can be translated into proteins with molecular weights of 34,000 (34K), 36K, 40K, and 42K. Transformed cells that also contain an intact early region 1b synthesized RNA which can be translated into proteins with molecular weights of 19K and 65K. Minor proteins of 15K, 16K, 17.5K, 18K, 25K, and 29K were also observed, but these proteins could not be mapped unambiguously. Cells transformed by the 8% HindIII-G apparently lack RNA encoding the 65K protein, but they do contain RNA coding for the 19K protein.

Adenoviruses, Human↗
PubMed1982

Proteins containing only half of the coding information of early region 1b of adenovirus are functional in human cells transformed with the herpes simplex virus type 1 thymidine kinase gene and adenovirus type 2 DNA.

We introduced into tk- human 143 cells adenovirus type 2 (Ad2) genes by transformation with a plasmid (p711) containing both Ad2 sequences and the herpes simplex virus type 1 (HSV-1) tk gene. p711 contained approximately the left 8% of the Ad2 genome inserted in the HindIII site of pBR322, whereas the fragment of HSV-1 containing the tk gene was inserted in the BamHI site. Three tk+ cell lines were isolated after selection in HAT medium. The arrangement of viral sequences in the three transformants was analyzed by restriction endonuclease digestion and filter hybridization. All three lines contained a single insertion of Ad2 DNA which was present at approximately one copy per cell. The arrangement of Ad2 sequences in these lines was identical to that found in the linear p711 DNA used in the transformation. S1 analysis of the Ad2-specified RNA from two of these lines indicated that the early region 1a mRNA's were synthesized, though in lower amounts than found in lytic infections. These cell lines contained only the left half of early region 1b (4.6 to 11.2), which encoded the 5' portion of the 1b mRNA's. A complex pattern of 1b RNAs was made in these cell lines. Transcription of most of these RNAs began at or near the 1b promoter and proceeded through the 1b sequences into the flanking pBR322, HSV-1, or host sequences. Since many of the RNAs were terminated or spliced in the HSV-1 (anti-sense strand) or pBR322 sequences, new RNA processing sites must be used in the formation of these mRNA's. All three lines fully complemented the 1a deletion mutant Ad5 dl312. Surprisingly, these lines also permitted the growth of 1b deletion mutants (Ad5 dl313 and Ad5 dl434), although the complementation was not always complete. Presumably the new gene product(s) which contained only part of the 1b gene provided most of the essential function(s) required for viral multiplication. Alternatively, the 1b 19-kilodalton protein which was entirely encoded by the adenovirus sequences present in these cell lines was sufficient for viral growth even in the absence of the 1b 55-kilodalton protein.

Adenoviruses, Human↗
PubMed1983

Transformation of rat cells by cyt mutants of adenovirus type 12 and mutants of adenovirus type 5.

Several mutants with much reduced oncogenicity (spontaneous mutants H12 cyt 52 and H12 cyt 70 and UV-induced mutants H12 cyt 61, H12 cyt 62, and H12 cyt 68) of the highly oncogenic adenovirus type 12 (Ad12) were studied for their ability to transform primary baby rat kidney cells. Four of the mutants showed much reduced capacity to transform cells in vitro, while H12 cyt 61 transformed cells as efficiently as the wild-type virus. Viral gene expression in several cell lines established from cultures infected by cyt mutants was studied, and it was found that viral sequences belonging to the left 16% of Ad12 were always transcribed. These results suggest that the function of the transformed state is not defective in the cyt mutants studied. Heterotypic complementation studies showed that the defect(s) in a cyt mutant can be corrected by an Ad7 function. Ad5 dl 313, with a deletion between 3.5 and 10.5 map units, transformed rat cells only at high multiplicity. These results suggest that the region E1B of adenoviruses may be required for efficient transformation of rat cells.

Adenoviruses, Human↗
PubMed1983

Control of adenovirus gene expression: cellular gene products restrict expression of adenovirus host range mutants in nonpermissive cells.

Adenovirus type 5 (Ad5) host range mutants dl312 and hr-1, with lesions in region E1A (0 to 4.5 map units) of the viral genome, fail to accumulate virus-specific early RNA during infection in HeLa cells. In a recent report, we showed that the addition of anisomycin, a stringent inhibitor of protein synthesis, at 1 h after infection of HeLa cells with hr-1 virus resulted in the accumulation of properly spliced and translatable mRNA from all early regions (M. G. Katze, H. Persson, and L. Philipson, Mol. Cell. Biol. 1:807-813, 1981). Based on these results we proposed a model in which expression of early mutant RNA was achieved through inactivation of a cellular protein normally causing a reduction in the amount of viral RNA. These studies have been extended in the present report, which shows that early viral proteins can be detected in Ad5 dl312- and Ad5 hr-1-infected HeLa cells which have been treated for several hours with anisomycin either shortly after infection or before infection. A pulse of drug treatment also resulted in expression of substantial amounts of adenovirus structural proteins after infection with both Ad5 hr-1 and Ad5 dl312, whereas in drug-free controls no late proteins were detected. The Ad5 hr-1 virus previously reported to be DNA replication negative in nonpermissive HeLa cells was found to replicate its DNA, albeit at low levels, when anisomycin was present either from 1 to 5 h postinfection or for 5 h before infection. When infectious virus production was examined in mutant-infected cells the titer of Ad5 dl312 virus was found to increase at least 500-fold in anisomycin-treated HeLa cells. Taken together, these and our previous results suggest that the block in gene expression characteristic for complementation group I Ad5 host range mutants in HeLa cells can be overcome by inactivating cellular gene products serving as negative regulators of viral gene expression.

Adenoviruses, Human↗
PubMed1992

Simian virus 40 small t antigen trans activates the adenovirus E2A promoter by using mechanisms distinct from those used by adenovirus E1A.

As reported previously for simian virus 40 small t antigen, polyomavirus small t antigen stimulates transcription directed by the adenovirus E2A and VA-I promoters during transient transfection assays. To determine whether papovaviral small t antigens might employ biochemical mechanisms during transcription activation that are either similar to or distinct from other viral trans activators, I compared the abilities of simian virus 40 small t antigen and adenovirus E1A to regulate the E2A promoter during transient transfection assays. I determined that, whereas activation of the E2A promoter by E1A involves the transcription factors ATF and EIIF, activation by small t antigen involves only EIIF. The effects of cotransfecting maximal concentrations of plasmids encoding small t antigen with E1A suggested that they activate the E2A promoter by different mechanisms. To determine whether small t antigen employs a mechanism different from that encoded in E1A domain II, domain III, or both, I compared the effects of transfecting plasmids expressing small t antigen, the 12S product of E1A, or the 13S product with a mutation in domain II on trans activation of the E2A promoter in two cellular backgrounds. On the basis of these comparisons, it appears that small t antigen does not activate transcription by a mechanism similar to either of the activities encoded in E1A. This suggests that papovavirus small t antigens belong to a distinct class of trans-acting proteins.

Adenoviridae↗
PubMed2005

Adenovirus early region 3 transgenes expressed in beta cells prevent autoimmune diabetes in nonobese diabetic mice: effects of deleting the adenovirus death protein 11.6K.

The incidence of type 1 diabetes (T1D) is decreased in nonobese diabetic mice expressing the complete cassette of adenovirus early region 3 (E3) immunomodulating genes in pancreatic beta cells. Embedded among the antiapoptotic E3 genes is one encoding an adenovirus death protein (ADP), which contributes to release of virion particles by promoting cell lysis. Because removal of this proapoptotic protein might have further enhanced the ability of E3 proteins to prevent T1D, an ADP-inactivated E3 construct was tested. Significantly, deletion of ADP did not improve the diabetes-protective effect of an E3 gene cassette.

Adenovirus E3 Proteins↗
PubMed2006

Regulatable gutless adenovirus vectors sustain inducible transgene expression in the brain in the presence of an immune response against adenoviruses.

In view of recent serious adverse events and advances in gene therapy technologies, the use of regulatable expression systems is becoming recognized as indispensable adjuncts to successful clinical gene therapy. In the present work we optimized high-capacity adenoviral (HC-Ad) vectors encoding the novel tetracycline-dependent (TetOn)-regulatory elements for efficient and regulatable gene expression in the rat brain in vivo. We constructed two HC-Ad vectors encoding beta-galactosidase (beta-gal) driven by a TetOn system containing the rtTAS(s)M2 transactivator and the tTS(Kid) repressor under the control of the murine cytomegalovirus (mCMV) (HC-Ad-mTetON-beta-Gal) or the human CMV (hCMV) promoter (HC-Ad-hTetON-beta-Gal). Expression was tightly regulatable by doxycycline (Dox), reaching maximum expression in vivo at 6 days and returning to basal levels at 10 days following the addition or removal of Dox, respectively. Both vectors achieved higher transgene expression levels compared to the expression from vectors encoding the constitutive mCMV or hCMV promoter. HC-Ad-mTetON-beta-Gal yielded the highest transgene expression levels and expressed in both neurons and astrocytes. Antivector immune responses continue to limit the clinical use of vectors. We thus tested the inducibility and longevity of HC-Ad-mediated transgene expression in the brain of rats immunized against adenovirus by prior intradermal injections of RAds. Regulated transgene expression from HC-Ad-mTetON-beta-Gal remained active even in the presence of a significant systemic immune response. Therefore, these vectors display two coveted characteristics of clinically useful vectors, namely their regulation and effectiveness even in the presence of prior immunization against adenovirus.

Adenoviruses, Human↗
PubMed1999

[Genome analysis of adenovirus type 7 and adenovirus type 11].

PURPOSE: To study the epidemiology of adenovirus type 7 (Ad 7) conjunctivitis and adenovirus type 11 (Ad 11) conjunctivitis by determining genome types and subgenome types. MATERIALS AND METHODS: For Ad 7 I used twelve strains from patients with acute viral conjunctivitis and one strain from a patient with pneumonia. For Ad 11 I used seventeen strains from patients with acute viral conjunctivitis and three strains from patients with cystitis. For Ad 7 genome typing, I used eleven DNA restriction endonucleases (REs) recognizing 6- or 7-base pair sequences and for Ad 11 genome typing, I used seven. For Ad 7 and for Ad 11 subgenome typing, I used Taq I and Hinf I which recognize 4- or 5-base pair sequences. RESULTS: The thirteen Ad 7 strains all belonged to the same genome type and subgenome type. Ad 11 strains showed six genome types. Ad 11 p was the most frequent strain. Fifteen Ad 11 p strains showed three subgenome types, but none of them was the same as the prototype. CONCLUSION: Ad 7 seems quite stable and the Ad 7 epidemic may recur again. On the other hand Ad 11 showed several different types. Ad 11 was probably not epidemic in the first half of the 1990's.

Adenoviruses, Human↗
PubMed2002

Adenovirus targeting to c-erbB-2 oncoprotein by single-chain antibody fused to trimeric form of adenovirus receptor ectodomain.

The use of adenovirus (Ad) vectors for cancer gene therapy applications is currently limited by several factors, including broad Ad tropism associated with the widespread expression of coxsackievirus and adenovirus receptor (CAR) in normal human tissues, as well as limited levels of CAR in tumor cells. To target Ad to relevant cell types, we have proposed using soluble CAR (sCAR) ectodomain fused with a ligand to block CAR-dependent native tropism and to simultaneously achieve infection through a novel receptor overexpressed in target cells. To confer Ad targeting capability on cancer cells expressing the c-erbB-2/HER-2/neu oncogene, we engineered a bispecific adapter protein, sCARfC6.5, that consisted of sCAR, phage T4 fibritin polypeptide, and C6.5 single-chain fragment variable (scFv) against c-erbB-2 oncoprotein. Incorporation of fibritin polypeptide provided trimerization of sCAR fusion proteins that, compared with monomeric sCAR protein, resulted in augmented affinity to Ad fiber knob domain and in increased ability to block CAR-dependent Ad infection. We demonstrated that sCARfC6.5 protein binds to cellular c-erbB-2 oncoprotein and mediates efficient Ad targeting via a CAR-independent pathway. As illustrated in cancer cell lines that overexpress c-erbB-2, targeted Ad, complexed with sCARfC6.5 adapter protein, provided from 1.5- to 17-fold enhancement of gene transfer compared with Ad alone and up to 130-fold increase in comparison with untargeted Ad complexed with sCARf control protein. The use of recombinant trimeric sCAR-scFv adapter proteins may augment Ad vector potency for targeting cancer cell types.

Adenoviridae↗
PubMed2002

The presence of the adenovirus E3 region improves the oncolytic potency of conditionally replicative adenoviruses.

PURPOSE: The initial development of conditionally replicative adenoviruses (CRAds) for cancer treatment has aimed at achieving selective replication in and killing of malignant cells. Other aspects such as the potentiation of the cytolytic capacity have also been investigated but still require new endeavors. As an extension of our prior work, we analyzed the effect of the E3 region, which includes the adenovirus death protein, in the context of CRAd oncolytic potency. EXPERIMENTAL DESIGN: We constructed E3-positive (E3+) and E3-negative (E3-) variants of the previously characterized CRAd, Ad5-Delta 24, and its infectivity enhanced version, Ad5-Delta 24RGD, and compared their oncolytic effect in human cancer cell lines infected with 0.01 viral particle/cell and in s.c. xenografts of A549 human lung cancer cells injected intratumorally with a single dose of 10(7) adenoviral particles in immunodeficient mice. RESULTS: The in vitro experiments showed that the E3+ viruses kill tumor cells 1.6-20 times more effectively in different cell lines. As well, the in vivo study demonstrated that the administration of E3+ CRAds resulted in a more potent oncolytic effect compared with the same dose of their E3- counterparts 35 days after virus administration. Moreover, a time course study of virus replication within the tumor xenografts established a correlation between higher in situ propagation of E3+ CRAds and tumor growth inhibition compared with E3- viruses. CONCLUSIONS: These results indicate that the presence of E3 can enhance the antitumor potency of CRAds over and above the levels conferred by the enhancement of infectivity via Arg-Gly-Asp (RGD).

Adenoviridae↗
PubMed1975

Canine adenovirus type 2-induced immunity to two canine adenoviruses in pups with maternal antibody.

Twenty-four Beagle pups with high levels of maternal antibody to canine adenovirus type 1 (CAV-1) and canine adenovirus type 2 (CAV-2) were oronasally inoculated with CAV-2 at 4 weeks of age. The CAV-2 was isolated from pharyngeal swabs on postinoculation days 2 through 6. In spite of the infection, maternal antibody continued to decrease for 4 to 8 postinoculation weeks, and then homologous CAV-2 neutralizing antibody and, to a lesser extent, CAV-1 neutralizing antibody began to increase. When these pups were challenge inoculated with CAV-1 and CAV-2 at a time when maternal antibody to CAV-1 would normally have disappeared, they were immune. In addition, 3 pups with maternal antibody to CAV-1 and CAV-2 were intramuscularly inoculated with CAV-2 at 3 weeks of age. Virus was not isolated from these pups, and maternal antibody decreased at a normal rate. These pups were not immune to challenge inoculation with CAV-1 and CAV-2.

Adenoviridae↗
PubMed1988

Effect of methyl methanesulfonate on type 5 adenovirus DNA integration and the phenotypic properties of cold-sensitive type 5 adenovirus-transformed cloned rat embryo fibroblast cells.

Pretreatment of a cloned rat embryo fibroblast cell line (CREF) with methyl methanesulfonate (MMS) prior to infection with a specific cold-sensitive type 5 adenovirus mutant, H5hr1, results in a unique carcinogen enhancement of transformation phenotype. MMS induces a dose-dependent increase in the absolute number of transformed foci in comparison with solvent-treated controls as well as an increase in transformation frequency when normalized for carcinogen-induced cell toxicity. To determine if the carcinogen enhancement of transformation phenotype was a consequence of the carcinogen altering the pattern of type 5 adenovirus (Ad5) DNA integration into the genome of CREF cells and/or if carcinogen treatment modified the phenotype of established H5hr1-transformed CREF cells, we have analyzed a series of single cell-derived H5hr1-transformed CREF cultures which were isolated from cultures pretreated with carcinogen-solvent or MMS prior to infection with H5hr1. Analysis of viral DNA integration by DNA filter-transfer hybridization (Southern blotting) indicated that MMS pretreatment did not increase the copy number of Ad5 DNA sequences which persisted in H5hr1-transformed clones or result in transformants which contained identical DNA restriction enzyme cleavage patterns. MMS-pretreated H5hr1-transformed clones also did not differ significantly from solvent-pretreated H5hr1-transformed clones in their ability to grow in agar, bind 125I-epidermal growth factor, or form tumors in athymic nude mice. MMS-pretreated H5hr1-transformed CREF clones retained a similar cold-sensitive negative regulation in the expression of the transformed cell phenotype as did H5hr1-transformed clones not exposed to carcinogens. These findings suggest that the unique carcinogen enhancement of transformation phenotype displayed by CREF cells pretreated with MMS prior to infection with H5hr1 does not result in transformants which either contain increased concentrations of Ad5 DNA or similar patterns of Ad5 DNA integration. Furthermore, carcinogen-pretreated H5hr1 transformants did not display novel phenotypes not expressed by cloned H5hr1-transformed CREF cell lines exposed to solvent prior to viral infection.

Adenoviruses, Human↗
PubMed1976

In vitro transformation of rodent cells by simian adenovirus 7 and bovine adenovirus type 3 (strain WBR-1).

Hamster, rat, and mouse embryo cells were quantitatively transformed by simian adenovirus 7 with maximum efficiencies of 2.1 X 10(5), and 2.5 X 10(5) plaque-forming unit/focus-forming unit, respectively. Bovine adenovirus type 3 did not transform rat or mouse embryo cells, but transformed hamster embryo cells with a maximum efficiency of 3.4 X 10(4) plaque-forming unit/focus-forming unit. All of the transformed cells were tumorigenic in appropriate hosts and possessed viral-specific T-antigen.

Adenoviridae↗
PubMed2002

The novel early region 3 protein E3/49K is specifically expressed by adenoviruses of subgenus D: implications for epidemic keratoconjunctivitis and adenovirus evolution.

The early transcription unit 3 (E3) of adenoviruses (Ads) encodes immunomodulatory functions. We previously described a novel gene of 49K within the E3 region of Ad19a, an Ad of subgenus D that is similar to Ad8 and Ad37 causes epidemic keratoconjunctivitis (EKC). Interestingly, 49K was reported not to be present in Ad9 and Ad17, other subgenus D Ads not causing EKC. Therefore, we investigated whether 49K is selectively expressed in EKC-causing Ads. Using specific DNA probes, we detect 49K-homologous genes in all subgenus D Ads tested. Moreover, 49K-specific antibodies recognize a high molecular weight protein in cells infected with all subgenus D serotypes irrespective of their ability to cause EKC. Sequencing of several 49K genes reveals a high homology without a distinct feature recognizable for those of EKC-associated Ad strains. Thus, E3/49K is a subgenus D specific E3 protein whose expression does not correlate with the EKC-causing phenotype and thus may rather be implicated in illnesses commonly caused by this subgenus. Interestingly, the 49K sequences of Ad19a and Ad37 are identical. To estimate the extent of the sequence identity between these two viruses, we initially sequenced the right ITR and the hexon. This analysis revealed that the right ITR of Ad19a is identical to Ad37, while the hexon sequence is Ad19p-like. This suggested that the region of identity is much larger and that Ad19a arose by recombination of Ad37 with an Ad19p-like Ad. Further sequencing mapped the crossover within the DNA binding protein. Thus, Ad19a contains a large sequence block ( approximately 13 kb), from the 100K gene to the right ITR, identical to Ad37. The implications of these findings in light of the temporal appearance of the EKC-causing Ad strains are discussed.

Adenovirus E3 Proteins↗
PubMed2005

The 100K-chaperone protein from adenovirus serotype 2 (Subgroup C) assists in trimerization and nuclear localization of hexons from subgroups C and B adenoviruses.

Recombinant hexons from subgroup C adenoviruses (Ad2 and Ad5) and from a member of subgroup B (Ad3) adenoviruses have been expressed in insect cells. When expressed alone, all three hexons were found to be insoluble and accumulated as inclusion bodies in the cytoplasm. However, co-expression of recombinant Ad2, Ad5 or Ad3 hexon with Ad2 L4-100K protein resulted in the formation of soluble trimeric hexons. EM analysis of hexons revealed that they were indistinguishable from native hexon capsomers isolated from Ad2-infected human cells, or released from partially disrupted adenovirions. This suggests that 100K acts as a chaperone for hexon folding and self-assembly into capsomer in insect cells. Since 100K protein assists in the trimerization of subgroup C hexon, and of subgroup B hexon protein, it implies that it functions in a manner that is both homo- and heterotypic. During the course of recombinant protein expression, the 100K protein was found in association with hexon monomers and trimers within the cytoplasm. In the nucleus, however, 100K was found in complexes with hexon trimers exclusively. EM observation of purified 100K protein samples showed a dumb-bell-shaped molecule compatible with a monomeric protein. EM analysis of hexon-100K protein complexes showed that interaction of hexon with the 100K protein occurred via one of the globular domains of the 100K protein molecule. Our data confirm the role of the 100K protein as a scaffold protein for hexon, and provide evidence suggesting its function in hexon nuclear import in insect cells.

Adenoviridae↗
PubMed2005

Immunization with recombinant adenovirus synthesizing the secretory form of Japanese encephalitis virus envelope protein protects adenovirus-exposed mice against lethal encephalitis.

Replication-defective recombinant adenoviruses (RAds) were constructed that synthesized the pre-membrane and envelope (E) proteins of Japanese encephalitis virus (JEV). Recombinant virus RAdEa synthesized Ea, the membrane-anchored E protein, and RAdEs synthesized Es, the secretory E protein. Compared with RAdEs, RAdEa replicated poorly in HEK 293A cells and synthesized lower amounts of E protein. Oral immunization of mice with RAds generated low titers of anti-JEV antibodies that had little JEV neutralizing activity. Intra-muscular (IM) immunization of mice with either RAd generated high titers of anti-JEV antibodies. Interestingly, RAdEa induced only low titers of JEV neutralizing antibodies. Titers were significantly higher in case of RAdEs immunization. Splenocytes from mice immunized IM with RAds secreted large amounts of interferon-gamma and moderate amounts of interleukin-5 in the presence of JEV and showed cytotoxic activity against JEV-infected cells. Naïve mice immunized IM with RAdEs showed complete protection against a lethal dose of JEV given intra-cerebrally. In order to study the effect of the pre-existing adenovirus 5 (Ad5) immunity on the outcome of the RAdEs immunization, mice were exposed to Ad5 through IM or intra-nasal (IN) routes before immunization with RAdEs. Mice exposed to Ad5 through the IN route, when immunized with RAdEs given IM, or those exposed to Ad5 through the IM route, when immunized with RAdEs given IN, were completely protected against lethal JEV challenge.

Adenoviridae↗
PubMed2003

Phylogenetic analysis of the hexon and protease genes of a fish adenovirus isolated from white sturgeon (Acipenser transmontanus) supports the proposal for a new adenovirus genus.

The organisation of the central part of the genome of a fish adenovirus (AdV) isolated from white sturgeon (Acipenser transmontanus) was studied. The putative genes identified between those of the viral DNA polymerase and the pVIII protein showed no significant difference in size or localisation compared to other known non-mastadenoviral genomes. The complete nucleotide sequences of the hexon and the viral protease genes and the intergenic region in the white sturgeon adenovirus (WSAdV-1) were compared with members of the four official AdV genera. In the case of WSAdV-1, merely two nucleotides separated the hexon and the protease genes, while in the other AdVs certain genus-specific features were recognised. In distance analyses based on complete sequence of the hexon or the protease proteins, the clear separation of five groups was seen corresponding to the four accepted AdV genera and WSAdV-1. Although there were slight differences between the topologies of the phylogenetic trees, the results unambiguously confirmed the distinctness of WSAdV-1 thus supporting the establishment of a new, fifth AdV genus.

Adenoviridae↗