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Biomedical subjects

F Wong-Staal

Publications and source records attributed to F Wong-Staal.

15 recordsLinked to original sources
PubMed1992

Inhibition of human immunodeficiency virus type 1 expression by a hairpin ribozyme.

Ribozymes are RNAs that possess the dual properties of RNA sequence-specific recognition, analogous to conventional antisense molecules, and RNA substrate destruction via site-specific cleavage. The cleavage reaction is catalytic in that more than one substrate molecule is processed per ribozyme molecule. We have designed a hairpin ribozyme that cleaves human immunodeficiency virus type 1 (HIV-1) RNA in the leader sequence (at nucleotides +111/112 relative to the transcription initiation site). The ribozyme was tested in vitro and gave efficient and specific cleavage of RNA containing the leader sequence. To test the antiviral efficacy of this ribozyme, we have cotransfected into HeLa cells HIV-1 proviral DNA and a plasmid expressing the ribozyme from the human beta-actin promoter. HIV-1 expression was inhibited as measured by p24 antigen levels and reduced Tat activity. The antiviral effect of the ribozyme appears to be specific and results from directed RNA cleavage; activity requires both a target sequence and a functional RNA catalytic center. These results suggest that this HIV-1-directed hairpin ribozyme may be useful as a therapeutic agent.

Base Sequence↗
PubMed1992

Biological characterization of noninfectious HIV-1 particles lacking the envelope protein.

To understand the role of the HIV-1 envelope protein in the assembly of virus, we constructed a proviral clone of HIV-1 where the methionine initiator codon of the env gene was substituted with a translational stop codon. Upon DNA transfection into permissive cells in culture, this clone produces virus-like particles similar in size to parental virus but are noninfectious in human T-cells, promonocytic cells, and primary macrophages. This mutant readily recombines with a deletion mutant provirus lacking the entire gag-pol region producing a recombinant virus that is infectious. Substitution of the same initiator methionine codon with valine results in a leaky missense mutant provirus capable of a low level of Env protein synthesis that leads to a productive infection. Thus, the prototype initiation codon AUG is dispensable for virus infectivity. Further, the expression of the envelope protein is not a prerequisite for the assembly of the virus particles in the HIV-1 system. These noninfectious envelope-less particles revert readily to wild-type phenotype upon cotransfection with Env-producing plasmid DNAs.

Animals↗
PubMed1979

Detection of integrated type-C viral DNA fragments in two primates (human and gibbon) by the restriction enzyme blotting technique.

We have shown that 1. partial provirus integration can be a possible result of a natural infection, and may serve as a model in animal systems where a viral etiology is implicated but detection of a major fraction of the virus genome is rare; 2. All human DNA contains some sequences that hybridize specifically with genomes of SiSV-SiSAV, suggesting that viruses of this group have infected humans in the past and recombined with human cellular DNA. 3. Finally, DNA from uncultured leukocytes of two leukemic patients, one being HL23, which yielded the virus HL23V in culture, was shown to have virus specific fragments related to BaEV. Another human DNA sample revealed virus specific fragments related to SiSV(SiSAV). These fragments are probably acquired by infection.

Animals↗
PubMed1979

Retrovirus sequences in a leukemic gibbon and its contact: evidence for partial provirus in the nonleukemic gibbon.

Integrated viral DNA sequences were detected in tissues from two gibbon apes, a leukemic gibbon (6G-1) from whose leukocytes a distinct strain of gibbon ape leukemia virus (GaLVH) was isolated, and gibbon 6G-4, a contact of 6G-1 from the same colony that had uremia and cachexia of unknown origin. Although 6G-4 had no detectable neoplasia or viral proteins, its serum contained persistent antibody against GaLV antigens. Whereas DNA from most of the tissues of 6G-1 contained GaLV provirus, DNA from only three tissues (kidney, spleen, and liver) from 6G-4 showed detectable viral sequences, and the extent of hybridization in each case was lower than with 6G-1. After cleavage with BamHI, two virus-specific DNA fragments were detected in tissues of 6G-1. Only one of these fragments was detected in the positive tissues of 6G-4. The results indicate that: (i) 6G-4 was exposed to and infected by GaLV; (ii) early target sites for infection of gibbon by GaLV may be limited to a few tissues; and (iii) infection can be contained by integration of only partial provirus in a few tissues.

Animals↗
PubMed1977

Divergence of baboon endogenous type C virogenes in primates: genomic viral RNA in molecular hybridization experiments.

RNA purified from two related RNA tumor viruses, one isolated from a baboon, Papio anubis, and the second from cultured blood leukocytes of a patient with acute myelogenous leukemia, was labeled with 125I and hybridized to DNA from different primates. RNA from both viruses showed maximum sequence homology with genes in baboons and little homology with genes of humans. The results confirm earlier suggestions that both viruses originated by transcription of baboon virogenes, and that one was transmitted to humans in nature. Hybridization of the viral RNA to cell DNA followed complicated kinetic patterns, indicating the presence of both repeated and infrequent virogene elements. This conclusion was verified in experiments using varied DNA:RNA ratios. It is proposed that virogenes, though composed of genes repeated 10 times or more, consist of some sequences more preferentially conserved than others. The non-uniformity of virogene sequence conservation limits the use of viral probes in studies concerning certain aspects of virogene evolution.

Animals↗
PubMed1976

Interactions between 125-I-labelled RNA from RNA tumor viruses and RNA of other sources.

Fragmented 125I-labelled RNA from RNA tumor viruses was hybridized to unlabelled RNA from cells, viruses, and homoribopolymers. The viral RNA interacted with all RNA tested, except for certain homoribopolymers. Complex formation with unlabelled RNA was verified by nuclease resistance, buoyant density measurements, and thermal stability in solutions of different ionic strength. The RNAase-resistant complex involved 20-30% of the sequences in the 125I-labelled viral RNA and formed preferentially with nuclear RNA of cells. 125I-labelled hemoglobin mRNA, 125I-labelled immunoglobulin light chain (lambda2) mRNA, or 125I-labelled viral RNA from encephalomyocarditis virus (EMC) dit not from RNAase-resistant complexes with unlabelled cellular RNA.

Animals↗
PubMed1976

Proviral sequences of baboon endogenous type C RNA virus in DNA of human leukaemic tissues.

Hybridisation of RNA from a baboon endogenous type C RNA virus to DNA from tissues of leukaemic patients indicates that a virus of this type is horizontally transmitted among humans. DNA from several patients with leukaemia hybridised 70% of the hybridisable RNA from baboon endogenous type C RNA virus (BaEV) and yielded hybrids of high tm, whereas DNA from normal human tissues hybridised only 23% of the BaEV RNA, and the tm of these hybrids was lower.

Adult↗
PubMed1976

Primate type-C virus nucleic acid sequences (woolly monkey and baboon types) in tissues from a patient with acute myelogenous leukemia and in viruses isolated from cultured cells of the same patient.

Cultured peripheral blood leukocytes from a woman (patient HL23) with acute myelogenous leukemia produced type-C RNA tumor viruses (HL23V). The viruses were analyzed by molecular hybridization experiments after transmission to five secondary cell culture lines. Using the criteria of molecular hybridization, we concluded that all of the transmitted virus isolates have nucleotide sequences related to the genome of simian sarcoma virus (SiSV). In addition, in agreement with data reported elsewhere, some of the transmitted viruses also have nucleotide sequences related to those of the baboon endogenous virus (BaEV). We also used molecular hybridization to ascertain whether both viruses could have originated from the patient HL23. Utilizing [3H] cDNA complementary to RNA from the separated BaEV-related component of HL23V and hybridizing this cDNA to DNA from tissues of the patient, we detected sequences related to BaEV in DNA obtained from the patient's spleen. These BaEV DNA sequences were also detectable when 125I-labeled RNA from BaEV was used as a probe. In agreement with earlier results, however, no SiSV-related sequences were detectable in the DNA of her tissues. Cytoplasmic viral-like particles, which had a buoyant density of 1.15-1.2 g/ml and were capable of synthesizing cDNA in association with a 35S RNA in vitro, were also found in the patient's fresh uncultured leukemic blood cells. cDNA synthesized by the cytoplasmic particles contained some sequences that hybridized to RNA from SiSV and, in addition, some that hybridized to RNA from BaEV. The cDNA also hybridized significantly to DNA isolated from the spleen of patient HL23 and to cytoplasmic RNA from the patient's leukocytes. These molecular hybridization results with nucleic acids obtained from the fresh blood cells of the patient, combined with the repeated isolation of similar viruses from different blood and bone marrow samples from the same patient, suggest that the virus come directly from the leukemic cell samples. The finding of BaEV-related DNA proviral sequences in the spleen of the patient strongly supports this interpretation. The failure so far to find a complete SiSV-related provirus is perplexing, but could be attributable to the existence of such a provirus in DNA of only a small population of cells in most leukemic patient.

Base Sequence↗
PubMed1975

Murine intracisternal type A particles: a biochemical characterization.

Intracisternal A particle preparations from a murine neuroblastoma cell line (N18) and from a mineral oil-induced murine plasmacytoma (MOPC-104E) contain both an endogenous RNA-dependent DNA polymerase activity and high molecular-weight polyadenylic acid (poly[A])-containing RNA. The DNA polymerase activity is stimulated by oligo(dG)-poly(C) and oligo(dT)-poly(A) and to a lesser extent by oligo(dT)-poly(dA), in agreement with previous reports. The high-molecular-weight RNA is predominantly 35S and contains a poly(A) tract of approximately 220 nucleotides as judged by polyacrylamide gel electrophoresis. Small amounts of 70S RNA are also present. This RNA preparation contains RNA homologous to RNA from type-C particles, as judged by molecular hybridization experiments. However, since this RNA derives only in part from A-particles and in part from other cellular RNA, hybridization of A-particle endogenously synthesized DNA or reverse transcripts of A-particle RNA to purified type C viral 70S RNA may more accurately reflect the relationship of A-particle RNA to RNA from C-particles. None of these DNA transcripts hybridizes significantly to C-particle 70S RNA, although MOPC and N18 DNA transcripts share significant homology. Our interpretation of these results is that murine intracisternal A particles are not closely related genetically to the tested murine type C viruses, although an alternate possibility is that all the A-particle DNA transcripts are copied from only a small part of the genome, which is unrelated to C-particle RNA.

Base Sequence↗