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Journal of Virology Nov 1998Vectors derived from human immunodeficiency virus (HIV) are highly efficient vehicles for in vivo gene delivery. However, their biosafety is of major concern. Here we...
Vectors derived from human immunodeficiency virus (HIV) are highly efficient vehicles for in vivo gene delivery. However, their biosafety is of major concern. Here we exploit the complexity of the HIV genome to provide lentivirus vectors with novel biosafety features. In addition to the structural genes, HIV contains two regulatory genes, tat and rev, that are essential for HIV replication, and four accessory genes that encode critical virulence factors. We previously reported that the HIV type 1 accessory open reading frames are dispensable for efficient gene transduction by a lentivirus vector. We now demonstrate that the requirement for the tat gene can be offset by placing constitutive promoters upstream of the vector transcript. Vectors generated from constructs containing such a chimeric long terminal repeat (LTR) transduced neurons in vivo at very high efficiency, whether or not they were produced in the presence of Tat. When the rev gene was also deleted from the packaging construct, expression of gag and pol was strictly dependent on Rev complementation in trans. By the combined use of a separate nonoverlapping Rev expression plasmid and a 5' LTR chimeric transfer construct, we achieved optimal yields of vector of high transducing efficiency (up to 10(7) transducing units [TU]/ml and 10(4) TU/ng of p24). This third-generation lentivirus vector uses only a fractional set of HIV genes: gag, pol, and rev. Moreover, the HIV-derived constructs, and any recombinant between them, are contingent on upstream elements and trans complementation for expression and thus are nonfunctional outside of the vector producer cells. This split-genome, conditional packaging system is based on existing viral sequences and acts as a built-in device against the generation of productive recombinants. While the actual biosafety of the vector will ultimately be proven in vivo, the improved design presented here should facilitate testing of lentivirus vectors.
Topics: Animals; Base Sequence; Brain; DNA Primers; Genes, Reporter; Genes, rev; Genes, tat; Genetic Therapy; Genetic Vectors; Genome, Viral; HIV-1; HeLa Cells; Humans; Lentivirus; Male; Polymerase Chain Reaction; Promoter Regions, Genetic; Rats; Rats, Inbred F344; Safety; Transduction, Genetic
PubMed: 9765382
DOI: 10.1128/JVI.72.11.8463-8471.1998 -
AIDS Research and Human Retroviruses Mar 1996
Review
Topics: Animals; Antibodies, Viral; Arthritis-Encephalitis Virus, Caprine; DNA, Viral; Gene Deletion; Genes, tat; Genes, vif; Goats; Lentivirus Infections; Vaccines, Attenuated; Viral Vaccines; Virus Replication
PubMed: 8882321
DOI: 10.1089/aid.1996.12.409 -
Current Opinion in Immunology Aug 1991Immediately after infection, human immunodeficiency virus directs the synthesis of three regulatory proteins tat, rev and nef that together allow the synthesis of the... (Review)
Review
Immediately after infection, human immunodeficiency virus directs the synthesis of three regulatory proteins tat, rev and nef that together allow the synthesis of the structural proteins of the virus after a delay of several hours. Viral mRNA production is controlled by the tat gene, which appears to stimulate elongation by RNA polymerase II, and the rev gene, which allows the accumulation of unspliced or partially spliced mRNAs in the cytoplasm. The nef gene is dispensible for virus growth but may limit virus spread by downregulating the levels of cellular surface proteins such as the CD4 receptor. Virus maturation also depends critically on the protease gene which allows the orderly rearrangement of the viral core structures in newly budded virions as well as the vpu and vif genes which allow efficient production of mature envelope glycoprotein.
Topics: Chromosome Mapping; DNA, Viral; Endopeptidases; Gene Expression Regulation, Viral; Genes, nef; Genes, rev; Genes, tat; Genes, vif; Genes, vpu; HIV; HIV Antigens; Humans; RNA Splicing; Transcription, Genetic; Transcriptional Activation; Virus Replication
PubMed: 1755979
DOI: 10.1016/0952-7915(91)90016-t -
Plant Physiology and Biochemistry : PPB Dec 2018Tyrosine aminotransferase (TAT, EC 2.6.1.5) is the first key enzyme that catalyzes the reversible interconversion of tyrosine and 4-hydroxyphenylpyruvate in the...
Comprehensive genomic analysis of the TYROSINE AMINOTRANSFERASE (TAT) genes in apple (Malus domestica) allows the identification of MdTAT2 conferring tolerance to drought and osmotic stresses in plants.
Tyrosine aminotransferase (TAT, EC 2.6.1.5) is the first key enzyme that catalyzes the reversible interconversion of tyrosine and 4-hydroxyphenylpyruvate in the tyrosine-derived pathway for syntheses of important secondary metabolites and compounds. Although plant TAT genes have been proposed to be important in response to abiotic stress, there is little information about TAT genes in woody perennial tree species, especially in economic fruit trees. Based on TAT domain searching, sequence homology screening and phylogenetic analysis, we identified four TATs in apple genome. Then, we carried out a detailed phylogenetic analysis of TAT genes from multi-species, focusing on apple (Malus domestica). The result showed that the TAT family comprises three major classes corresponding to genes from angiosperms, mammals, and bacteria. Angiosperm TAT genes could be further divided into six subclasses. Analysis of intron-exon structure revealed that the typical TAT gene contains six introns and seven exons, with exons of similar size at each exon location. Promoter analysis showed that the 5'-flanking region of apple MdTATs contain multiple cis-acting elements including those implicated in light, biotic stress, abiotic stress, and hormone response. MdTATs were expressed to various levels in all apple structures and organs evaluated, and showed distinct expression patterns under water deficit stress. Ectopic expression of MdTAT2 in Arabidopsis or over-expression of MdTAT2 in apple callus tissue conferred enhanced tolerance to drought and osmotic stress. Collectively, these results suggest a role for TAT genes in drought and osmotic stresses and provide valuable information for further research of TAT genes and their function in plants.
Topics: Dehydration; Genome, Plant; Genomics; Malus; Osmotic Pressure; Plant Proteins; Tyrosine Transaminase
PubMed: 30391815
DOI: 10.1016/j.plaphy.2018.10.033 -
ELife Jan 2016A virus protein called Tat plays a dual role in HIV infection by regulating the expression of genes belonging to the virus and genes belonging to the host cells.
A virus protein called Tat plays a dual role in HIV infection by regulating the expression of genes belonging to the virus and genes belonging to the host cells.
Topics: HIV; HIV Infections; Host-Pathogen Interactions; Humans; Transcription, Genetic; Virus Replication; tat Gene Products, Human Immunodeficiency Virus
PubMed: 26783762
DOI: 10.7554/eLife.12686 -
Virus Research May 2018For the production of viral genomic RNA, HIV-1 is dependent on an early viral protein, Tat, which is required for high-level transcription. The quantity of viral RNA...
For the production of viral genomic RNA, HIV-1 is dependent on an early viral protein, Tat, which is required for high-level transcription. The quantity of viral RNA detectable in blood of HIV-1 infected individuals varies dramatically, and a factor involved could be the efficiency of Tat protein variants to stimulate RNA transcription. HIV-1 virulence, measured by set-point viral load, has been observed to increase over time in the Netherlands and elsewhere. Investigation of tat gene evolution in clinical isolates could discover a role of Tat in this changing virulence. A dataset of 291 Dutch HIV-1 subtype B tat genes, derived from full-length HIV-1 genome sequences from samples obtained between 1985-2012, was used to analyse the evolution of Tat. Twenty-two patient-derived tat genes, and the control Tat were analysed for their capacity to stimulate expression of an LTR-luciferase reporter gene construct in diverse cell lines, as well as for their ability to complement a tat-defective HIV-1 clone. Analysis of 291 historical tat sequences from the Netherlands showed ample amino acid (aa) variation between isolates, although no specific mutations were selected for over time. Of note, however, the encoded protein varied its length over the years through the loss or gain of stop codons in the second exon. In transmission clusters, a selection against the shorter Tat86 ORF was apparent in favour of the more common Tat101 version, likely due to negative selection against Tat86 itself, although random drift, transmission bottlenecks, or linkage to other variants could also explain the observation. There was no correlation between Tat length and set-point viral load; however, the number of non-intermediate variants in our study was small. In addition, variation in the length of Tat did not significantly change its capacity to stimulate transcription. From 1985 till 2012, variation in the length of the HIV-1 subtype B tat gene is increasingly found in the Dutch epidemic. However, as Tat proteins did not differ significantly in their capacity to stimulate transcription elongation in vitro, the increased HIV-1 virulence seen in recent years could not be linked to an evolving viral Tat protein.
Topics: Codon; Evolution, Molecular; Gene Expression Regulation, Viral; Genome, Viral; HIV Infections; HIV Long Terminal Repeat; HIV-1; Humans; Luciferases; Netherlands; Open Reading Frames; RNA, Viral; Transcription, Genetic; Transcriptional Activation; Viral Load; tat Gene Products, Human Immunodeficiency Virus
PubMed: 29654800
DOI: 10.1016/j.virusres.2018.04.008 -
Virus Research Dec 2002Several studies have indicated that human immunodeficiency virus type-1 (HIV-1) transactivating Tat protein is essential for proviral DNA transcription and virus...
Several studies have indicated that human immunodeficiency virus type-1 (HIV-1) transactivating Tat protein is essential for proviral DNA transcription and virus replication. In addition, it is actively released from acutely HIV-1-infected cells and interacts either with the same virus-infected and virus producing cell, or with bystander uninfected cells, influencing the expression of several genes and related cellular functions. The main goal of this paper was to determine the Tat-related expression of basic cellular genes in a permanently tat transfected CD4+ cell line, to identify the cellular genes influenced by the presence of endogenous-exogenous Tat protein. For this purpose, we analyzed, by a cDNA-membrane-array assay, cellular mRNAs expressed in serum-free cultures of lymphoblastoid CD4(+) Jurkat cells, stably transfected with a plasmid constitutively expressing tat gene, in comparison with Jurkat cells transfected with the backbone plasmid only, and parental Jurkat cells. The expression of mRNAs in permanently tat-transfected Jurkat cells showed significant differences in 24 out of 1176 analyzed genes in comparison with parental or backbone plasmid transfected cells. Most of the genes overexpressed in permanently tat-transfected Jurkat cells, belong to transcription factors, or to receptors, adaptors, and mediators of signal transduction pathways, and to factors involved in response to oxidative stress, suggesting a complex regulation of CD4(+) T-lymphoid cell survival and proliferation by HIV-1 Tat protein.
Topics: CD4-Positive T-Lymphocytes; Gene Expression Profiling; Gene Expression Regulation; Gene Products, tat; HIV-1; Humans; Jurkat Cells; Oligonucleotide Array Sequence Analysis; Proteins; RNA, Messenger; Transfection; tat Gene Products, Human Immunodeficiency Virus
PubMed: 12457987
DOI: 10.1016/s0168-1702(02)00253-8 -
FEMS Microbiology Letters Mar 2003The human immunodeficiency virus (HIV-1) (transactivator of transcription (Tat)) protein is a pleiotropic factor that induces a broad range of biological effects in... (Comparative Study)
Comparative Study Review
The human immunodeficiency virus (HIV-1) (transactivator of transcription (Tat)) protein is a pleiotropic factor that induces a broad range of biological effects in numerous cell types. At the HIV promoter, Tat is a powerful transactivator of gene expression, which acts by both inducing chromatin remodeling and by recruiting elongation-competent transcriptional complexes onto the viral LTR. Besides these transcriptional activities, Tat is released outside the cells and interacts with different cell membrane-associated receptors. Finally, extracellular Tat can be internalized by cells through an active endocytosis process. Here we discuss some of the molecular mechanisms involved in intracellular and extracellular Tat function.
Topics: Acetylation; Acetyltransferases; Bacteriophage lambda; Chromatin; Endocytosis; Extracellular Space; Gene Expression Regulation, Viral; Gene Products, tat; Genes, tat; HIV Long Terminal Repeat; HIV-1; Histone Acetyltransferases; Histones; Humans; Intracellular Fluid; Models, Biological; Positive Transcriptional Elongation Factor B; Promoter Regions, Genetic; Protein Processing, Post-Translational; Protein Serine-Threonine Kinases; RNA Polymerase II; RNA, Messenger; RNA, Viral; Receptors, Virus; Regulatory Sequences, Nucleic Acid; Saccharomyces cerevisiae Proteins; Transcriptional Activation; tat Gene Products, Human Immunodeficiency Virus
PubMed: 12644228
DOI: 10.1016/S0378-1097(03)00067-3 -
Biochemical and Biophysical Research... Mar 1995Cytokine responses are dramatically affected when HIV-1 infected cells are activated with certain antigenic stimuli. We report the effects of HIV-1 tat gene in cytokine...
Cytokine responses are dramatically affected when HIV-1 infected cells are activated with certain antigenic stimuli. We report the effects of HIV-1 tat gene in cytokine modulation, using HIV-1 tat transfected T (Jurkat) and B (Raji) cell lines. Studying the effect of tat and/or PMA + PHA on mRNA expression of 14 cytokines (IL-1 alpha, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12, TNF-alpha, TNF-beta, GM-CSF, TGF-beta, IFN-gamma and MIP-1 alpha) illustrated differential effects. In addition to the varied effects of tat on the steady state levels of cytokine mRNAs, tat induced the secretion of TNF-beta preferentially in both B and T cell lines, either by itself as in Raji B cell line or synergistically upon PMA + PHA stimulation as in Jurkat T cell line.
Topics: B-Lymphocytes; Base Sequence; Cytokines; DNA Primers; Gene Expression; Gene Expression Regulation, Viral; Genes, tat; Humans; In Vitro Techniques; Lymphocyte Activation; Molecular Sequence Data; Phytohemagglutinins; RNA, Messenger; T-Lymphocytes; Tetradecanoylphorbol Acetate; Transfection
PubMed: 7695626
DOI: 10.1006/bbrc.1995.1395 -
Current Topics in Microbiology and... 1995We have focused this chapter on interactions with two of the best characterized transregulatory genes, tax for HTLV-I/II and Tat for HIV-1. Both genes illustrate the... (Review)
Review
We have focused this chapter on interactions with two of the best characterized transregulatory genes, tax for HTLV-I/II and Tat for HIV-1. Both genes illustrate the complex interplay between retroviral regulatory genes and cellular gene regulation. In both instances a viral gene of relatively straightforward function in the viral context appears to cause extensive dysregulation of cellular genes, either directly or as a consequence of altered cellular differentiation. Understanding this viral/cellular gene cross-talk may elucidate mechanisms leading to malignant transformation autoimmune disease and to neurologic and paraneoplastic complications such as hypercalcemia for HTLV-I/II, as well as the pathogenesis of immune dysfunction and opportunistic malignancy in HIV-I/II-infected individuals. An understanding of functional mechanisms of these transregulatory viral genes will undoubtedly afford better explanations for the myriad manifestations of retroviral infection.
Topics: Animals; Gene Expression Regulation, Viral; Gene Products, tax; Genes, tat; HIV-1; Human T-lymphotropic virus 1; Human T-lymphotropic virus 2; Humans; Transcriptional Activation
PubMed: 7648877
DOI: 10.1007/978-3-642-78929-8_2