Multigene Lentiviral Vectors Based on Differential Splicing and Translational Control

Zhu, Yu Cheng; Feuer, Gerold; Day, S; Wrzesinski, Stephen H.; Planelles, Vicente

doi:10.1006/mthe.2001.0469

Cited by 41 publications

(33 citation statements)

References 23 publications

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“…Plasmids-We described the lentiviral vector, pHR-GFP, in a previous report (33). pHR-VPR was derived by substituting the B7.1 cDNA in place of human T-cell lymphotrophic virus-I tax in the vector pHRЈCMV/Tax1/eGFP (34).…”

Section: Methodsmentioning

confidence: 99%

Activation of the ATR-mediated DNA Damage Response by the HIV-1 Viral Protein R

Roshal

Kim

Zhu³

et al. 2003

Journal of Biological Chemistry

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183

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DNA damage is a universal inducer of cell cycle arrest at the G 2 phase. Infection by the human immunodeficiency virus type 1 (HIV-1) also blocks cellular proliferation at the G 2 phase. The HIV-1 accessory gene vpr encodes a conserved 96-amino acid protein (Vpr) that is necessary and sufficient for the HIV-1-induced block of cellular proliferation. In the present study, we examined a recently identified DNA damage-signaling protein, the ATM-and Rad3-related protein, ATR, for its potential role in the induction of G 2 arrest by Vpr. We show that inhibition of ATR by pharmacological inhibitors, by expression of the dominant-negative form of ATR, or by RNA interference inhibits Vpr-induced cell cycle arrest. As with DNA damage, activation of ATR by Vpr results in phosphorylation of Chk1. This study provides conclusive evidence of activation of the ATR-initiated DNA damage-signaling pathway by a viral gene product. These observations are important toward understanding how HIV infection promotes cell cycle disruption, cell death, and ultimately, CD4؉ lymphocyte depletion.

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Section: Methodsmentioning

confidence: 99%

Activation of the ATR-mediated DNA Damage Response by the HIV-1 Viral Protein R

Roshal

Kim

Zhu³

et al. 2003

Journal of Biological Chemistry

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183

215

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“…Vector production and concentration HIV-1-based defective lentiviral vectors (DLV) encoding selected cis-acting elements were produced by transient transfection of human embryonic kidney 293T cells with a packaging construct, a VSV-G envelope construct and a transfer construct containing the reporter gene GFP (Figure 1) [13,33]. In brief, 24 h prior to transfection, 293T cells were seeded in 75 cm 2 tissue culture (TC) flasks at a density of about 6.5 × 10 6 cells/flask with 14 ml Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal bovine serum (FBS) and incubated at 37 °C with 5% CO 2 .…”

Section: Methodsmentioning

confidence: 99%

HIV-1-based defective lentiviral vectors efficiently transduce human monocytes-derived macrophages and suppress replication of wild-type HIV-1

et al. 2005

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Background-Human monocytes play an important role in mediating human immunodeficiency virus type 1 (HIV-1) infection of the central nervous system (CNS), and monocytes-derived macrophages (MDM) represent a major viral reservoir within the brain and other target organs. Current gene transduction of MDM is hindered by a limited efficiency. In this study we established a lentiviral vector-based technique for improved gene transfer into human MDM cultures in vitro and demonstrated significant protection of transduced MDM from super-infection with wild-type HIV-1.

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“…In gene therapy, IRES elements are used in dicistronic or multicistronic vectors to drive multigene expression from single mRNAs, e.g. a therapeutic gene together with a selectable marker or two subunits of a heterodimeric protein (21)(22)(23)(24)(25)(26). In artificial virus replicon systems, the internal insertion of an additional IRES element (most commonly the EMCV IRES) is widely used to uncouple the required constitutive expression of the viral nonstructural replication proteins from the biologic activity of the viral cis-signals under investigation (27)(28)(29)(30)(31).…”

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confidence: 99%

Picornavirus Internal Ribosome Entry Site Elements Can Stimulate Translation of Upstream Genes

Jünemann¹,

Song

Bassili

et al. 2007

Journal of Biological Chemistry

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Certain viral and cellular mRNAs initiate translation capindependently at internal ribosome entry site (IRES) elements. Picornavirus IRES elements are widely used in dicistronic or multicistronic vectors in gene therapy, virus replicon systems, and analysis of IRES function. In such vectors, expression of the upstream gene often serves as internal control to standardize the readings of IRES-driven downstream reporter activity. Picornaviral IRES elements translate optimally at up to 120 mM K ؉ concentration, whereas genes used as upstream reporters usually have lower salt optima when present in monocistronic mRNAs. However, here we show that such reporter genes are efficiently translated at higher K ؉ concentrations when placed upstream of a functional picornavirus IRES. This translation enhancement occurs in cis, is independent of the nature of the first reporter and of second reporter translation, and is conferred by the IRESs of picornaviruses but not of hepatitis C virus. A defective picornavirus IRES with a deletion killing IRES activity but leaving the binding site for initiation factor eIF4G intact retains translation enhancement activity. Translation enhancement on a capped mRNA is disabled by m 7 GDP. In addition, the C-terminal fragment of eIF4G can confer translation enhancement also on uncapped mRNA. We conclude that whenever eIF4F has been captured to a dicistronic mRNA by binding to a picornavirus IRES via its eIF4G moiety, it can be provided in cis to the 5-end of the RNA and there stimulate translation initiation, either by binding to the cap nucleotide using its eIF4E moiety or by binding to the RNA cap-independently using its eIF4G moiety. Internal ribosome entry site (IRES)3 elements are cis-acting RNA regions that confer the internal entry of ribosomes to the RNA translation start site independent of the process of capdependent scanning from the 5Ј-end of the RNA (1, 2). Such IRES elements were first discovered in picornaviruses like encephalomyocarditis virus (EMCV) (3), poliovirus (4), and foot-and-mouth disease virus (FMDV) (5). Also the translation of hepatitis C virus (HCV) (6) and of several cellular mRNAs is driven by IRES elements (2).The picornaviral IRES elements are classified in three groups according to their sequences and secondary structures, the type I elements of the entero-/rhinovirus group (including poliovirus), the type II elements of the cardio-/aphthovirus group (including EMCV and FMDV), and the type III element of hepatitis A virus (1). After infection of a susceptible cell, these IRES elements guide the small ribosomal subunit to an AUG triplet in a starting window at the 3Ј-border of the IRES (1, 2, 7). Ribosome binding to the picornavirus IRES elements is mediated by a number of cellular RNA-binding proteins (8) that fall into two groups. On one hand, all standard eukaryotic translation initiation factors (eIFs) are required (9), except the actual cap-binding protein eIF4E, a protein that associates with the large adaptor protein eIF4G (and the RNA-helicase eIF4A) i...

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Multigene Lentiviral Vectors Based on Differential Splicing and Translational Control

Cited by 41 publications

References 23 publications

Activation of the ATR-mediated DNA Damage Response by the HIV-1 Viral Protein R

Activation of the ATR-mediated DNA Damage Response by the HIV-1 Viral Protein R

HIV-1-based defective lentiviral vectors efficiently transduce human monocytes-derived macrophages and suppress replication of wild-type HIV-1

Picornavirus Internal Ribosome Entry Site Elements Can Stimulate Translation of Upstream Genes

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