SEARCH · PubMed Health

Results for “Infectious clone”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 325 records · Page 18Linked to original sources

PubMed2003

Evolution of Soybean mosaic virus-G7 molecularly cloned genome in Rsv1-genotype soybean results in emergence of a mutant capable of evading Rsv1-mediated recognition.

Plant resistance (R) genes direct recognition of pathogens harboring matching avirluent signals leading to activation of defense responses. It has long been hypothesized that under selection pressure the infidelity of RNA virus replication together with large population size and short generation times results in emergence of mutants capable of evading R-mediated recognition. In this study, the Rsv1/Soybean mosaic virus (SMV) pathosystem was used to investigate this hypothesis. In soybean line PI 96983 (Rsv1), the progeny of molecularly cloned SMV strain G7 (pSMV-G7) provokes a lethal systemic hypersensitive response (LSHR) with up regulation of a defense-associated gene transcript (PR-1). Serial passages of a large population of the progeny in PI 96983 resulted in emergence of a mutant population (vSMV-G7d), incapable of provoking either Rsv1-mediated LSHR or PR-1 protein gene transcript up regulation. An infectious clone of the mutant (pSMV-G7d) was synthesized whose sequences were very similar but not identical to the vSMV-G7d population; however, it displayed a similar phenotype. The genome of pSMV-G7d differs from parental pSMV-G7 by 17 substitutions, of which 10 are translationally silent. The seven amino acid substitutions in deduced sequences of pSMV-G7d differ from that of pSMV-G7 by one each in P1 proteinase, helper component-proteinase, and coat protein, respectively, and by four in P3. To the best of our knowledge, this is the first demonstration in which experimental evolution of a molecularly cloned plant RNA virus resulted in emergence of a mutant capable of evading an R-mediated recognition.

Cloning, Molecular↗

PubMed2005

Amplification and cloning of near full-length HIV-2 genomes.

The genomes of human immunodeficiency virus type 2 (HIV-2), like those of HIV-1, are not only extremely variable but are also highly recombinogenic. Determination of subtypes based on partial genomes cannot predict the subtype classification of other regions of the genome owing to the frequent occurrence of recombinant genomes among subtypes. To fully understand the genetic variation and evolution of HIV-2s, full-length viral genomes need to be obtained for genetic analysis. Full-length HIV-2 genomes can also be used as infectious clones to study viral biological characteristics and as reference sequences for phylogenetic analysis. More important, all genes in the obtained genomes can be cloned into expression vectors to produce proteins that can be used as antigens for diagnostic reagents or as immunogens for vaccine development. The long-range polymerase chain reaction technique has recently become a more preferred method than the lambda phage cloning method to obtain near-full-length HIV-2 genomes.

Cloning, Molecular↗

PubMed2006

Small interfering RNA targeted to stem-loop II of the 5' untranslated region effectively inhibits expression of six HCV genotypes.

BACKGROUND: The antiviral action of interferon alpha targets the 5' untranslated region (UTR) used by hepatitis C virus (HCV) to translate protein by an internal ribosome entry site (IRES) mechanism. Although this sequence is highly conserved among different clinical strains, approximately half of chronically infected hepatitis C patients do not respond to interferon therapy. Therefore, development of small interfering RNA (siRNA) targeted to the 5'UTR to inhibit IRES mediated translation may represent an alternative approach that could circumvent the problem of interferon resistance. RESULTS: Four different plasmid constructs were prepared for intracellular delivery of siRNAs targeting the stem loop II-III of HCV 5' UTR. The effect of siRNA production on IRES mediated translation was investigated using chimeric clones between the gene for green fluorescence protein (GFP) and IRES sequences of six different HCV genotypes. The siRNA targeted to stem loop II effectively mediated degradation of HCV IRES mRNA and inhibited GFP expression in the case of six different HCV genotypes, where as siRNAs targeted to stem loop III did not. Furthermore, intracytoplasmic expression of siRNA into transfected Huh-7 cells efficiently degraded HCV genomic RNA and inhibited core protein expression from infectious full-length infectious clones HCV 1a and HCV 1b strains. CONCLUSION: These in vitro studies suggest that siRNA targeted to stem-loop II is highly effective inhibiting IRES mediated translation of the major genotypes of HCV. Stem-loop II siRNA may be a good target for developing an intracellular immunization strategy based antiviral therapy to inhibit hepatitis C virus strains that are not inhibited by interferon.

5' Untranslated Regions↗

PubMed1996

CD4 confers susceptibility to human immunodeficiency virus type 1 infection in a rat fibroblast cell line.

A new model system is delineated that will enable study of CD4 cofactors and gp120 binding proteins other than CD4. We have previously described a nontransformed rat fibroblast cell line that can efficiently produce HIV-1 upon transfection with an HIV-1 infectious clone, in contrast to other nonhuman mammalian cell lines tested. In the present study we analyzed the susceptibility to HIV-1 infection of Rat2 cells expressing the human CD4 protein. We have used the mammalian expression vector pKS286, in which HIV-1 LTR drives the expression of the CD4 gene. We show that the Rat2 cell line, expressing the human CD4 (Rat2/CD4), is susceptible to fusion with and infection by HIV-1. The virus produced by the Rat2/CD4 cells was infectious. CD4 expression in the Rat2/CD4 was down-regulated over time, similarly to HIV-1 expression in HIV-1-transfected Rat2 cells. Transfection of the Rat2/CD4 cells with a tat expression vector reestablished the CD4 expression on the surface of those cells, as expected. We conclude that the expression of a gp120 binding protein on the Rat2 cells' surface suffices to render the Rat2 cells susceptible to HIV-1 infection.

Animals↗

PubMed2005

Genetically engineered, live, attenuated vaccines protect nonhuman primates against aerosol challenge with a virulent IE strain of Venezuelan equine encephalitis virus.

Two live, attenuated strains of Venezuelan equine encephalitis virus (VEE), IE1150K and V3526, were administered to macaques to determine if they could elicit protection against an aerosol challenge with virulent VEE virus of the IE variety (VEEV-IE). These viruses were rescued from full-length cDNA clones of 68U201 (VEEV-IE variety) and Trinidad donkey (VEEV-IA/B variety), respectively, and both have a furin cleavage site deletion mutation and a second-site resuscitating mutation. Both vaccines elicited neutralizing antibodies to viruses of the homologous variety but not to viruses of the heterologous variety. Eight weeks after vaccination, the macaques were challenged by aerosol exposure to virulent 68U201. Macaques vaccinated with V3526 were protected as well as macaques inoculated with IE1009, the wild-type infectious clone of 68U201. However, IE1150K failed to significantly protect macaques relative to controls. V3526 has now been shown to protect macaques against both IA/B [Pratt WD, Davis NL, Johnston RE, Smith JF. Genetically engineered, live attenuated vaccines for Venezuelan equine encephalitis: testing in animal models. Vaccine 2003;21(25-26):3854-62] and IE strains of VEE viruses.

Aerosols↗

PubMed2006

The small envelope protein of porcine reproductive and respiratory syndrome virus possesses ion channel protein-like properties.

The small envelope (E) protein of porcine reproductive and respiratory syndrome virus (PRRSV) is a hydrophobic 73 amino acid protein encoded in the internal open reading frame (ORF) of the bicistronic mRNA2. As a first step towards understanding the biological role of E protein during PRRSV replication, E gene expression was blocked in a full-length infectious clone by mutating the ATG translational initiation to GTG, such that the full-length mutant genomic clone was unable to synthesize the E protein. DNA transfection of PRRSV-susceptible cells with the E gene knocked-out genomic clone showed the absence of virus infectivity. P129-DeltaE-transfected cells however produced virion particles in the culture supernatant, and these particles contained viral genomic RNA, demonstrating that the E protein is essential for PRRSV infection but dispensable for virion assembly. Electron microscopy suggests that the P129-DeltaE virions assembled in the absence of E had a similar appearance to the wild-type particles. Strand-specific RT-PCR demonstrated that the E protein-negative, non-infectious P129-DeltaE virus particles were able to enter cells but further steps of replication were interrupted. The entry of PRRSV has been suggested to be via receptor-mediated endocytosis, and lysomotropic basic compounds and known ion-channel blocking agents both inhibited PRRSV replication effectively during the uncoating process. The expression of E protein in Escherichia coli-mediated cell growth arrests and increased the membrane permeability. Cross-linking experiments in cells infected with PRRSV or transfected with E gene showed that the E protein was able to form homo-oligomers. Taken together, our data suggest that the PRRSV E protein is likely an ion-channel protein embedded in the viral envelope and facilitates uncoating of virus and release of the genome in the cytoplasm.

Animals↗

PubMed1995

Infection of central nervous system cells by ecotropic murine leukemia virus in C58 and AKR mice and in in utero-infected CE/J mice predisposes mice to paralytic infection by lactate dehydrogenase-elevating virus.

Certain mouse strains, such as AKR and C58, which possess N-tropic, ecotropic murine leukemia virus (MuLV) proviruses and are homozygous at the Fv-1n locus are specifically susceptible to paralytic infection (age-dependent poliomyelitis [ADPM]) by lactate dehydrogenase-elevating virus (LDV). Our results provide an explanation for this genetic linkage and directly prove that ecotropic MuLV infection of spinal cord cells is responsible for rendering anterior horn neurons susceptible to cytocidal LDV infection, which is the cause of the paralytic disease. Northern (RNA) blot hybridization of total tissue RNA and in situ hybridization of tissue sections demonstrated that only mice harboring central nervous system (CNS) cells that expressed ecotropic MuLV were susceptible to ADPM. Our evidence indicates that the ecotropic MuLV RNA is transcribed in CNS cells from ecotropic MuLV proviruses that have been acquired by infection with exogenous ecotropic MuLV, probably during embryogenesis, the time when germ line proviruses in AKR and C58 mice first become activated. In young mice, MuLV RNA-containing cells were found exclusively in white-matter tracts and therefore were glial cells. An increase in the ADPM susceptibility of the mice with advancing age correlated with the presence of an increased number of ecotropic MuLV RNA-containing cells in the spinal cords which, in turn, correlated with an increase in the number of unmethylated proviruses in the DNA extracted from spinal cords. Studies with AKXD recombinant inbred strains showed that possession of a single replication-competent ecotropic MuLV provirus (emv-11) by Fv-1n/n mice was sufficient to result in ecotropic MuLV infection of CNS cells and ADPM susceptibility. In contrast, no ecotropic MuLV RNA-positive cells were present in the CNSs of mice carrying defective ecotropic MuLV proviruses (emv-3 or emv-13) or in which ecotropic MuLV replication was blocked by the Fv-1n/b or Fv-1b/b phenotype. Such mice were resistant to paralytic LDV infection. In utero infection of CE/J mice, which are devoid of any endogenous ecotropic MuLVs, with the infectious clone of emv-11 (AKR-623) resulted in the infection of CNS cells, and the mice became ADPM susceptible, whereas littermates that had not become infected with ecotropic MuLV remained ADPM resistant.

Animals↗

PubMed1988

Agrobacterium-mediated infectivity of cloned digitaria streak virus DNA.

A monomeric clone of double-stranded DNA synthesized in vitro DNA of the geminivirus Digitaria streak (DSV) was subcloned as a tandem dimeric unit into a binary vector of Agrobacterium tumefaciens, creating a plasmid pDS2. Inoculation of digitaria sanguinalis with A. tumefaciens carrying pDS2 resulted in viral infection. The symptoms, virus particles, and DNA forms obtained were indistinguishable from those of a natural DSV infection of D. sanguinalis. Inoculations have also induced infections in Zea mays and Avena sativa. The sequence of the Agrobacterium-mediated infectious clone of DSV has been determined.

Cloning, Molecular↗

PubMed1996

A point mutation in the HIV-1 Tat responsive element is associated with postintegration latency.

Study of the mechanism of HIV-1 postintegration latency in the ACH2 cell line demonstrates that these cells failed to increase HIV-1 production following treatment with exogenous Tat. Reasoning that the defect in ACH2 cells involves the Tat response, we analyzed the sequence of tat cDNA and Tat responsive element (TAR) from the virus integrated in ACH2. Tat cDNA sequence is closely related to that of HIV LAI, and the encoded protein is fully functional in terms of long terminal repeat (LTR) transactivation. Cloning of a region corresponding to the 5'-LTR from ACH2, however, identified a point mutation (C37 -> T) in TAR. This mutation impaired Tat responsiveness of the LTR in transient transfection assays, and the measured defect was complemented in cells that had been treated with tetradecanoyl phorbol acetate or tumor necrosis factor type alpha (TNF-alpha). A compensatory mutation in TAR (G28 -> A), designed to reestablish base pairing in the TAR hairpin, restored wild-type Tat responsiveness. When the (C37 -> T) mutation was introduced in an infectious clone of HIV-1, no viral production was measured in the absence of TNF-alpha, whereas full complementation was observed when the infection was conducted in the presence of TNF-alpha or when a compensatory mutation (G28 -> A) was introduced into TAR. These experiments identify a novel mutation associated with HIV-1 latency and suggest that alterations in the Tat-TAR axis can be a crucial determinant of the latent phenotype in infected individuals.

Base Sequence↗

PubMed2004

An attenuated West Nile prototype virus is highly immunogenic and protects against the deadly NY99 strain: a candidate for live WN vaccine development.

In a short time, West Nile virus has developed into a nationwide health and veterinary problem. The high virulence of the circulating virus and related lineage 1 WN strains hinders development of an attenuated live vaccine. We describe an attenuated WN isolate, WN1415, which is a molecularly cloned descendant of the WN prototype B956 strain. The parent virus belongs to lineage 2, members of which have not been associated with epidemic or epizootic outbreaks. A set of non-conservative mutations, mostly in non-structural protein genes, distinguishes the WN1415 isolate from the parent B956 prototype strain. Immunization with WN1415 (55-550,000 pfu) established a potent immunity, which protected the majority of mice against lethal challenge with WN NY99. The attenuated nature of the isolate and its excellent growth characteristics combined with the availability of a highly stable infectious clone make the isolate an attractive candidate for live WN vaccine development.

Amino Acid Substitution↗

PubMed2001

Identification of dna components required for induction of cotton leaf curl disease.

Cotton leaf curl disease (CLCuD) is a major constraint to cotton production in Pakistan. Infectious clones of the monopartite begomovirus cotton leaf curl virus (CLCuV), associated with diseased cotton, are unable to induce typical symptoms in host plants. We have identified and isolated a single-stranded DNA molecule approximately 1350 nucleotides in length which, when coinoculated with the begomovirus to cotton, induces symptoms typical of CLCuD, including vein swelling, vein darkening, leaf curling, and enations. This molecule (termed DNA beta) requires the begomovirus for replication and encapsidation. The CLCuV/DNA 1/DNA beta complex, together with a similar complex previously identified in Ageratum conyzoides, represent members of an entirely new type of infectious, disease-causing agents. The implications of this finding to our understanding of the evolution of new disease-causing agents are discussed.

Base Sequence↗

PubMed1998

Disease induction by virus derived from molecular clones of equine infectious anemia virus.

Equine infectious anemia virus (EIAV), a macrophage-tropic lentivirus, causes persistent infections of horses. A number of biologic features, including the rapid development of acute disease, the episodic nature of chronic disease, the propensity for viral genetic variation, and the ability for many infected animals to eventually control virus replication, render EIAV a potentially useful model system for the testing of antiretroviral therapies and vaccine strategies. The utility of the EIAV system has been hampered by the lack of proviral clones that encode promptly pathogenic viral stocks. In this report, we describe the generation and characterization of two infectious molecular clones capable of causing acute clinical syndromes similar to those seen in natural infections. Virus derived from clone p19/wenv17 caused severe debilitating disease at 5 to 7 days postinfection; initial febrile episodes were fatal in two of three infected animals. Virus derived from a second clone, p19/wenv16, caused somewhat milder primary febrile episodes by 10 to 12 days postinfection in two of two infected animals. Virus derived from both clones caused persistent infections such that some animals exhibited chronic equine infectious anemia, characterized by multiple disease episodes. The two virulent clones differ in envelope and rev sequences.

Acute Disease↗

PubMed2001

West Nile virus vaccine.

Within the past 5 years, West Nile encephalitis has emerged as an important disease of humans and horses in Europe. In 1999, the disease appeared for the first time in the northeastern United States. West Nile virus (a mosquito-borne flavivirus) has flourished in the North American ecosystem and is expected to expand its geographic range. In this review, the rationale for a human and veterinary vaccine is presented and a novel approach for rapid development of a molecularly-defined, live, attenuated vaccine is described. The technology (ChimeriVax) is applicable to the development of vaccines against all flaviviruses, and products against Japanese encephalitis (a close relative of West Nile) and dengue are in or are nearing clinical trials, respectively. ChimeriVax vaccines utilize the safe and effective vaccine against the prototype flavivirus -yellow fever 17D- as a live vector. Infectious clone technology is used to replace the genes encoding the pre-membrane (prM) and envelope (E) protein of yellow fever 17D vaccine with the corresponding genes of the target virus (e.g., West Nile). The resulting chimeric virus contains the antigens responsible for protection against West Nile but retains the replication efficiency of yellow fever 17D. The ChimeriVax technology is well-suited to the rapid development of a West Nile vaccine, and clinical trials could begin as early as mid-2002. Other approaches to vaccine development are briefly reviewed. The aim of this brief review is to describe the features of West Nile encephalitis, a newly introduced infectious disease affecting humans, horses and wildlife in the United States; the rationale for rapid development of vaccines; and approaches to the development of vaccines against the disease.

Animals↗

PubMed2001

Virion-targeted viral inactivation: new therapy against viral infection.

BACKGROUND: Acquired immune deficiency syndrome (AIDS) is resistant to all current therapy. Gene therapy is an attractive alternative or additive to current, unsatisfactory AIDS therapy. MATERIALS AND METHODS: To develop an antiviral molecule targeting viral integrase (HIV IN), we generated a single-chain antibody, termed scAb, which interacted with human immunodeficiency virus type 1 (HIV-1) IN and inhibited virus replication at the integration step when expressed intracellularly. To reduce infectivity from within the virus particles, we made expression plasmids (pC-scAbE-Vpr, pC-scAbE-CA, and pC-scAbE-WXXF), which expressed the anti-HIV IN scAb fused to the N-terminus of HIV-1-associated accessory protein R (Vpr), capsid protein (CA), and specific binding motif to Vpr (WXXF), respectively. All fusion proteins were tagged with a nine-amino acid peptide derived from influenza virus hemagglutinin (HA) at the C terminus. RESULTS: The fusion molecules, termed scAbE-Vpr, scAbE-CA, and scAbE-WXXF, interacted specifically with HIV IN immobilized on a nitrocellulose membrane. Immunoblot analysis showed that scAbE-Vpr, scAbE-CA, and scAbE-WXXF were incorporated into the virions produced by cotransfection of 293T cells with HIV-1 infectious clone DNA (pLAI) and pC-scAbE-Vpr, pC-scAbE-WXXF. A multinuclear activation galactosidase indicator (MAGI) assay revealed that the virions released from 293T cells cotransfected with pLAI and pC-scAbE-Vpr, pC-scAbE-WXXF had as little 1000-fold of the infectivity of the control wild-type virions, which were produced from the 293T cells transfected with pLAI alone. Furthermore, the virions produced from the 293T cells cotransfected with pLAI and an scAb expression vector (pC-scAb) showed only 1% of the infectivity of the control HIV-1 in a MAGI assay, although scAb was not incorporated into the virions. In either instance, the total quantity of the progeny virions released from the transfected 293T cells and the patterns of the virion proteins were hardly affected by the presence of scAb, scAbE-Vpr, or scAbE-WXXF, as determined by virion-associated reverse transcriptase assay and by immunoblot analysis, respectively. Because G418-selected HeLa clones carrying the expression plasmid for scAbE-WXXF were obtained much more frequently than those for scAbE-Vpr, scAbE-WXXF was inferred to be less toxic to cells than scAbE-Vpr. The result that scAbE-WXXF with viral incorporation achieved more than a 10-fold reduction in infectivity of the progeny virions than scAb without incorporation suggests that scAbE-WXXF is a potential antiviral molecule, inhibiting replication by neutralization of HIV IN activity both within cells and within virions. Moreover, it is nontoxic to human cells. We termed this gene therapy "virion-"targeted-viral inactivation" and these molecules "packageable antiviral therapeutics." CONCLUSION: This new gene therapy has the potential for wide application in many viral infectious diseases.

Acquired Immunodeficiency Syndrome↗

PubMed2001

Genome replication and postencapsidation functions mapping to the nonstructural gene restrict the host range of a murine parvovirus in human cells.

The infection outcome of the Parvoviridae largely relies on poorly characterized intracellular factors modulated by proliferation, differentiation, and transformation of host cells. We have studied the interactions displayed by the highly homologous p and i strains of the murine parvovirus minute virus of mice (MVM), with a series of transformed cells of rat (C6) and human (U373, U87, SW1088, SK-N-SH) nervous system origin, seeking for molecular mechanisms governing parvovirus host range. The MVMp infection of C6 and U373 cells was cytotoxic and productive, whereas the other nervous cells behaved essentially as resistant to this virus. In contrast, MVMi did not complete its life cycle in any of the human nervous cells, though it efficiently killed the astrocytic tumor cells by two types of nonproductive infections: (i) normal synthesis of all viral macromolecules with a late defect in infectious virion maturation and release to the medium in U373; and (ii) high levels of accumulation of the full set of viral messenger RNAs and of both nonstructural (NS-1) and structural (VP-1 and VP-2) proteins, under a very low viral DNA amplification, in U87 and SW1088 cells. Further analyses showed that U87 was permissive for nuclear transport of MVMi proteins, leading to efficient assembly of empty viral capsids with a normal phosphorylation and VP1-to-VP2 ratio. The DNA amplification blockade in U87 occurred after conversion of the incoming MVMi genome to the monomeric replicative form, and it operated independently of the delivery pathway used by the viral particle, since it could not be overcome by transfection with cloned infectious viral DNA. Significantly, a chimeric MVMi virus harboring the coding region of the nonstructural (NS) gene replaced with that of MVMp showed a similar pattern of restriction in U87 cells as the parental MVMi virus, and it attained in U373 cultures an infectious titer above 100-fold higher under equal levels of DNA amplification and genome encapsidation. The results suggest that the activity of complexes formed by the NS polypeptides and recruited cellular factors restrict parvovirus DNA amplification in a cell type-dependent manner and that NS functions may in addition determine MVM host range acting at postencapsidation steps of viral maturation. These data are relevant for understanding the increased multiplication of autonomous parvovirus in some transformed cells and the transduction efficacy of nonreplicative parvoviral vectors, as well as a general remark on the mechanisms by which NS genes may regulate viral tropism and pathogenesis.

Animals↗

PubMed1995

Inducible and conditional inhibition of human immunodeficiency virus proviral expression by vesicular stomatitis virus matrix protein.

Besides its role in viral assembly, the vesicular stomatitis virus (VSV) matrix (M) protein causes cytopathic effects such as cell rounding (D. Blondel, G. G. Harmison, and M. Schubert, J. Virol. 64:1716-1725, 1990). DNA cotransfection assays demonstrated that VSV M protein was able to inhibit the transcription of a reporter gene (B. L. Black and D. S. Lyles, J. Virol. 66:4058-4064, 1992). We have confirmed these observations by using cotransfections with an infectious clone of human immunodeficiency virus type 1 (HIV-1) and found that the amino-terminal 32 amino acids of M protein which are essential for viral assembly were not required for this inhibition. For the study of the potential role of M protein in the shutoff of transcription from chromosomal DNA, we have isolated stable HeLa T4 cell lines which encode either a wild-type or a temperature-sensitive (ts) VSV M gene under control of the HIV-1 long terminal repeat promoter. Transcription of the M mRNA was transactivated after HIV-1 infections. A cell line which encodes the wild-type M protein was nonpermissive for either HIV-1 or HIV-2. A cell line that encodes the ts M gene was transfected with the infectious HIV-1 DNA or was infected with HIV-1 or HIV-2. In all cases, at 32 degrees C, the permissive temperature for M protein, the cells were nonpermissive for HIV replication. At 40 degrees C, the ts M protein was nonfunctional and both HIV-1 and HIV-2 were able to replicate at high levels. A comparison of the amounts of proviral HIV-1 DNAs and HIV-1 mRNAs at 10 and 36 h after HIV-1 infection demonstrated that proviral insertion had not been prevented by M protein and that the block in HIV-1 replication was at the level of proviral expression. The severe reduction of HIV-1 proviral transcripts demonstrates that the VSV M protein alone can inhibit expression from chromosomal DNA. These results strongly support the hypothesis that the VSV M protein is involved in the shutoff of host cell transcription. M protein was able to attenuate HIV-1 infections and protect the cell population from HIV-1 pathogenesis. The temperature-dependent switch from a persistent to a lytic HIV-1 infection in the presence of ts M protein could be useful for studies of HIV-1 replication and pathogenesis.

Base Sequence↗

PubMed1984

Physical mapping of the paralysis-inducing determinant of a wild mouse ecotropic neurotropic retrovirus.

We have recently shown that a molecularly cloned ecotropic retrovirus, initially isolated from the brain of a paralyzed wild mouse, retained the ability to induce hind limb paralysis when inoculated into susceptible mice (Jolicoeur et al., J. Virol. 45:1159-1163, 1983). To map the viral DNA sequences encoding the determinant of paralysis, we constructed chimeric viral DNA genomes in vitro between parental cloned infectious viral DNA genomes from this neurotropic murine leukemia virus (MuLV) and from nonneurotropic amphotropic 4070-A MuLV. Infectious chimeric MuLVs, recovered after microinjection of NIH 3T3 cells with these recombinant DNAs, were inoculated into newborn SIM.S and SWR/J mice to test the paralysis-inducing potential. We found that the 3.9-kilobase-pair SalI-ClaI fragment of the neurotropic MuLV comprising the 3' end of pol and all env sequences was sufficient to confer the paralysis-inducing potential to chimeric viruses. Therefore, this region of the neurotropic MuLV genome most likely harbors the primary determinant of paralysis.

Animals↗

PubMed1983

Thymotropism of murine leukemia virus is conferred by its long terminal repeat.

Several murine leukemia viruses (MuLV) replicate efficiently in the thymus (T+) of the mouse, whereas others are unable to replicate (T-) in this organ. To map the region of the viral genome harboring the sequences responsible for this thymotropic phenotype, we constructed viral DNA recombinants in vitro between cloned infectious viral DNAs from T- BALB/c N-tropic MuLV and from T+ BALB/c B-tropic MuLV or AKR Gross passage A MuLV. (N- and B-tropic refer to the Fv-1 host range of MuLV.) Infectious recombinant MuLVs, recovered from murine cells microinjected with these recombinant DNAs, were injected into newborn mice to test their ability to replicate in the thymus. We found that the long terminal repeat from the T+ BALB/c B-tropic or AKR Gross passage A MuLV genome was sufficient to allow replication of recombinant MuLVs in the thymus. Our sequence data suggested that the U3 tandem direct repeat was responsible for this effect. These results suggest a new role for the U3 long terminal repeat in the replication of MuLV in specific differentiated target cells.

Animals↗