Efficient Functional Pseudotyping of Oncoretroviral and Lentiviral Vectors by Venezuelan Equine Encephalitis Virus Envelope Proteins

Kolokoltsov, Andrey A.; Weaver, Scott C.; Davey, Robert A.

doi:10.1128/jvi.79.2.756-763.2005

Cited by 33 publications

(33 citation statements)

References 37 publications

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“…Transcription is driven by the viral long terminal repeat (LTR) promoter/enhancer elements that flank the transgene and is dependent solely on normal cellular transcription mechanisms. Pseudotyped viruses were made essentially as described previously (20). Briefly, 293FT cells (Invitrogen) were transfected with plasmids encoding MLV Gag-Pol and VSV G together with a plasmid encoding the EGFP-tagged construct flanked by MLV LTRs and a 5Ј packaging sequence.…”

Section: Methodsmentioning

confidence: 99%

Critical Role for the Host GTPase-Activating Protein ARAP2 in InlB-Mediated Entry of Listeria monocytogenes

et al. 2010

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Listeria monocytogenes is a Gram-positive, food-borne bacterial pathogen capable of causing gastroenteritis, meningitis, or abortions (39, 32). Listeria induces its own internalization (entry) into nonphagocytic mammalian cells, a process that likely plays an important role in traversal of the intestinal, placental, and blood-brain barriers (7,14,16,23). One of the pathways of Listeria entry is mediated by interaction of the bacterial surface protein InlB with its host receptor, the Met receptor tyrosine kinase (16, 37).InlB-Met interaction triggers activation (tyrosine phosphorylation) of the Met receptor and subsequent rearrangements in the F-actin cytoskeleton of the mammalian cell (16,29). These cytoskeletal changes remodel the host cell surface, resulting in engulfment of adherent Listeria. One of the host mammalian proteins that acts downstream of Met to promote F-actin rearrangements and bacterial entry is type IA phosphoinositide (PI) 3-kinase (8,17,18). This PI 3-kinase is a heterodimeric enzyme comprised of an 85-kDa regulatory subunit and a 110-kDa catalytic subunit (4, 11). p85-p110 generates phosphatidylinositol 4,5-bisphosphate [PI(4,5)P 2 )] and phosphatidylinositol 3,4,5-trisphosphate [PI(3,4,5)P 3 ], which are lipid second messengers that regulate a variety of biological processes, including growth, survival, and motility of mammalian cells. A plethora of downstream "target" proteins that bind PI(4,5)P 2 and/or PI(3,4,5)P 3 and mediate the biological effects of p85-p110 have been identified (4, 24). However, mammalian proteins that act downstream of type IA PI 3-kinase to control Listeria uptake have yet to found.In this work, we demonstrate that the human GTPase-activating protein (GAP) ARAP2 is required for InlB-mediated cytoskeletal changes and entry of Listeria. ARAP2 is known to bind PI(3,4,5)P 3 , resulting in upregulation of a GAP domain that inactivates the mammalian GTPase Arf6 (42). We provide genetic evidence indicating that the ArfGAP domain of ARAP2 stimulates Listeria entry by antagonizing Arf6. ARAP2 has several functional domains in addition to its ArfGAP domain. One of these domains in ARAP2 interacts with the mammalian GTPase RhoA (42). Our genetic data demonstrate that this "RhoA binding" (RB) domain also plays a critical role in bacterial entry. Surprisingly, pharmacological experiments indicate that the RB domain controls Listeria uptake through an unknown mechanism that does not involve Rho proteins. Our work indicates a key role for host ARAP2 in InlB-mediated entry of Listeria. One of the likely ways that type IA PI 3-kinase controls entry of Listeria is through regulation of ARAP2. MATERIALS AND METHODSBacterial strains, mammalian cell lines, and media. The wild-type Listeria monocytogenes strain EGD and the isogenic ⌬inlB mutant strain containing an in-frame deletion of the inlB gene were previously described (10). These strains were grown in brain heart infusion (BHI) broth (Difco) and prepared for infection as described previously (10,18).The human epithelial cell lin...

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Section: Methodsmentioning

confidence: 99%

Critical Role for the Host GTPase-Activating Protein ARAP2 in InlB-Mediated Entry of Listeria monocytogenes

et al. 2010

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“…The 5Ј-long terminal repeat functions as a strong promoter upon chromosomal integration of proviral DNA. The pFB plasmid was modified to contain a cassette comprising an ECMV internal ribosome entry site followed by a gene encoding ␤-galactosidase (modified with a nuclear localization signal), which enabled retrovirus titer and transcript expression levels to be determined by staining for ␤-galactosidase (22). Retroviruses were made by simultaneous transfection of HEK293FT cells (Invitrogen) with the dnSrc expression plasmid and plasmids encoding murine leukemia retrovirus gag-pol and vesicular stomatitis virus envelope protein.…”

Section: Methodsmentioning

confidence: 99%

Src Regulates Constitutive Internalization and Rapid Resensitization of a Cholecystokinin 2 Receptor Splice Variant

Chao

Ives

Goluszko

et al. 2005

Journal of Biological Chemistry

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] i were used to assess receptor activity. Src kinase activity was inhibited by transducing cells with a retrovirus containing a dominant-negative mutant Src (A430V). The subcellular location of enhanced green fluorescent protein-tagged receptors was monitored using laser scanning confocal microscopy. Both receptor variants desensitized at the same rate; however, CCK 2i4sv R resensitized five times faster than CCK 2 R. Without agonist, 80% of CCK 2i4sv R is located in an intracellular compartment. In contrast, 80% of CCK 2 R was located on the plasma membrane. Treatment with inverse agonist (YM022) or expression of dominant-negative Src blocked the constitutive internalization of CCK 2i4sv R, resulting in its accumulation on the plasma membrane. Expression of dominant-negative Src slowed the rate of CCK 2i4sv R resensitization. Inhibition of Src did not affect G17-induced internalization of either receptor variant. Constitutive internalization of CCK 2i4sv R increases its rate of resensitization by creating an intracellular pool of receptors that can rapidly recycle back to the plasma membrane.Receptor desensitization and resensitization are essential mechanisms for maintaining physiologically appropriate cellular responses to extracellular stimuli. For G protein-coupled receptors (GPCR 2 (s)), these processes are regulated, in part, by the intracellular domains of the receptors. Within seconds to minutes of agonist binding, the activated receptor is desensitized when specific amino acid residues within the 3il and/or C-terminal tail domain of the receptor are phosphorylated, creating binding sites for proteins, such as ␤-arrestins and Src kinase, which block further receptor interactions with heterotrimeric G proteins and initiate receptor internalization (1-3). Resensitization involves the dephosphorylation of the intracellular domains and recycling of the intact receptor back to the plasma membrane where it can once again bind ligand (4, 5). Dysregulation of these processes can lead to disease (6, 7). CCK 2 R is a GPCR that exists as various splice variant isoforms (8). Previously, we reported the identification and cloning of a variant, called CCK 2i4sv R, which, as a result of intron 4 retention, contains an additional 69 amino acid residues in its 3il domain (9). In contrast to CCK 2 R, which is widely expressed in normal tissues of the gastrointestinal tract and nervous system, CCK 2i4sv R appears to be the predominant variant expressed by some hyperplastic polyps, colorectal and pancreatic cancers, and pancreatic cancer-derived cell lines (9 -12). Several lines of evidence support a role for CCK 2i4sv R in the progression of these cancers. First, CCK 2i4sv R, but not CCK 2 R, coimmunoprecipitates with activated Src kinase in an agonist-independent manner (13). Increased Src activity is associated with the development and progression of breast, brain, pancreas, and colon cancers (14,15). Second, expression of CCK 2i4sv R stimulates cell growth in vitro (9) and HEK293 tumor growth in vivo in a Src-dependen...

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“…without causing significant tissue damage [18][19][20][21][22]. This, coupled with the fact that some target the nervous system [23] and the idea that most humans do not have pre-existing antibodies against them due to their restricted geography [23] The alphaviral genome encodes several non-structural and structural proteins that are…”

Section: A C C E P T E D Accepted Manuscriptmentioning

confidence: 99%

Selective transduction of astrocytic and neuronal CNS subpopulations by lentiviral vectors pseudotyped with Chikungunya virus envelope

et al. 2017

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This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. NHAs was observed. In vivo stereotaxic delivery in striatum, thalamus and hippocampus respectively in the adult rat brain revealed localised transduction restricted to striatal astrocytes and hippocampal dentate granule neurons. Transduction of different subtypes of granule neurons from precursor to post-mitotic stages of differentiation was evident in the sub-granular zone and dentate granule cell layer. No significant inflammatory response was observed, but comparable to that of VSV-G pseudotyped lentiviral vectors. Robust longterm expression followed for three months post-transduction along with absence of neuroinflammation, coupled to the selective and unique neuron/glial tropism indicates that these vectors could be useful for modelling and gene therapy studies in the CNS.

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Efficient Functional Pseudotyping of Oncoretroviral and Lentiviral Vectors by Venezuelan Equine Encephalitis Virus Envelope Proteins

Cited by 33 publications

References 37 publications

Critical Role for the Host GTPase-Activating Protein ARAP2 in InlB-Mediated Entry of Listeria monocytogenes

Critical Role for the Host GTPase-Activating Protein ARAP2 in InlB-Mediated Entry of Listeria monocytogenes

Src Regulates Constitutive Internalization and Rapid Resensitization of a Cholecystokinin 2 Receptor Splice Variant

Selective transduction of astrocytic and neuronal CNS subpopulations by lentiviral vectors pseudotyped with Chikungunya virus envelope

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