Establishment and characterization of KB cell lines latently infected with adeno-associated virus type 1

Handa, Hiroshi; Shiroki, Kazuko; Shimojo, Hiroto

doi:10.1016/0042-6822(77)90034-4

Cited by 94 publications

(63 citation statements)

References 18 publications

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“…Negative regulation by rep might occur very early in the AAV growth cycle to provide time to amplify the replicating pool of DNA before encapsidation of progeny genomes attains maximum rates. Also, it is possible that the regulation by rep might be important during establishment or maintenance of AAV genome integration in the cell genome in the absence of helper virus or during its subsequent rescue with helper (12,24,29).…”

Section: Resultsmentioning

confidence: 99%

Negative and positive regulation in trans of gene expression from adeno-associated virus vectors in mammalian cells by a viral rep gene product.

Tratschin¹,

Tal²,

Carter³

1986

Mol. Cell. Biol.

149

143

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We previously described use of the human parvovirus, adeno-associated virus (AAV), as a vector for transient expression in mammalian cells of the gene for chloramphenicol acetyltransferase (CAT). In the AAV vector, pTS1, the CAT gene is expressed under the control of the major AAV promoter p4o. This The human parvovirus, adeno-associated virus (AAV), replicates in mammalian cells in the presence of helper functions provided by adenovirus or herpesvirus (10). When molecular clones containing the entire AAV type 2 (AAV2) genome in bacterial plasmids are transfected into mammalian cells in the presence of adenovirus, the AAV genome is excised from the plasmid and replicated to produce infectious AAV particles (38,50,52). This facilitated both the genetic analysis of AAV (27,(51)(52)(53)59) and the development of AAV as a eucaryotic expression vector (26,60,61).The AAV genome contains three transcription promoters, P5, Pi9, and P40, which yield overlapping transcripts (11). Genetic analysis showed that a major AAV reading frame (orf-2) which is accessible from P40 transcripts codes for a major portion of the AAV capsid proteins (27,59). Mutations in this region (cap-) allow normal synthesis of duplex replicating form DNA but prevent synthesis of progeny single-stranded DNA and infectious particles (27,53,59). A second major open reading frame (orf-1) in the left half of the genome is apparently accessible from either p5 or P19 transcripts and codes for one or more proteins (rep) required for AAV DNA replication (27,53,59). Mutations in this region (rep-) are DNA negative but can be complemented. Deletion of both terminal palindromes results in a cis-dominant (ori-) defect which reflects the presence of the AAV replication origins in the terminal repeat sequences (3,4,25,51,53).

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Section: Resultsmentioning

confidence: 99%

Negative and positive regulation in trans of gene expression from adeno-associated virus vectors in mammalian cells by a viral rep gene product.

Tratschin¹,

Tal²,

Carter³

1986

Mol. Cell. Biol.

149

143

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“…AAV infection of AdStransformed hamster cells inhibits Ad gene expression but does not affect cell viability or growth rate (Ostrove et al, 1981). Similarly, intact AAV can latently infect continuous lines of human cells with high efficiency (Hoggan et al, 1972;Handa et al, 1977). Finally, in experiments to be described below, under the appropriate conditions the cells can be transformed to neomycin resistance in a co-transfection with an intact AAV rep gene.…”

Section: Additional Inhibitory Effects Of the Aa V Rep Gene Productsmentioning

confidence: 99%

“…Although human infections are common, adenoassociated virus has never been implicated as an aetiological agent (Blacklow et al, 1968 a, b). In the absence of a helper virus the dependovirion penetrates to the cell nucleus where the DNA is uncoated and is then integrated into the cell genome in a highly efficient manner (Handa et al, 1977;Cheung et al, 1980). The genome can be rescued from the integrated state after superinfection of the cell with a helper virus, again in a highly efficient manner (Hoggan et aL, 1972;Berns et al, 1975;Handa et el., 1977).…”

Section: Introductionmentioning

confidence: 99%

Parvovirus Gene Regulation

Berns

Labow

1987

Journal of General Virology

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“…The present study was an attempt to elucidate the position effect of transduced gene expression. We selected a recombinant adeno-associated virus (rAAV)-mediated gene delivering system, for the following reasons: AAV is nonpathogenic (Berns and Bohenzky 1987), integration of the viral genome is efficient (Samulski et al 1991;Kotin et al 1992), virus integration appears to have no apparent effect on cell growth or morphology (Handa et al 1977), there is a highly efficient transduction of human hematopoietic progenitor cells (Goodman et al 1994), and there is no possibility of a rearranged proviral genome due to unexpected splicing events commonly observed in the globin gene retroviral delivery system (Novak et al 1990). We constructed a series of rAAV vectors containing the human β-globin gene and various combinations of HS core elements, and another rAAV vector containing two copies of the 250-bp core fragment of the chicken β-globin insulator in both sides of the HS-globin gene cassette.…”

Section: Introductionmentioning

confidence: 99%

Position-independent human β-globin gene expression mediated by a recombinant adeno-associated virus vector carrying the chicken β-globin insulator

et al. 1999

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The position-independent expression of transgenes in target cells is an essential subject for determining effective gene therapies. The chicken β-globin insulator blocks the effects of regulatory sequences on transcriptional units at differential domains. We prepared a recombinant adeno-associated virus (rAAV) containing various combinations of the DNase I-hypersensitive site 2 (HS2), 3 (HS3), and 4 (HS4) core elements from the human β-globin locus control region (LCR), the human β-globin gene, and the herpes virus thymidine kinase promoter driven neomycinresistant gene (neo R ) (rHS432, rHS43, rHS42, rHS32, and rHS2), and also rAAV containing two copies of the 250-bp core sequence of the chicken β-globin insulator on both sides of the rHS2 (rIns/HS2/2Ins). After isolating neomycinresistant mouse erythroleukemia (MEL) cells infected with each rAAV, we analyzed the rAAV genome by Southern blots and polymerase chain reaction (PCR), using primers specific for HS core elements and the human β-globin gene. All clones contained a single copy of the rAAV genome in the chromosome, however, some of them had a rearranged proviral genome. In five clones with a single unrearranged rAAV genome for each rAAV construct, we assayed the expression of the human b-globin gene relative to the endogenous mouse β maj -globin gene, using quantitative reverse transcriptase (RT)-PCR. In clones infected with rHS432, the expression level of the human β-globin gene ranged from 51.6% to 765.6% of that in the mouse β majglobin gene. Likewise, in rHS43, the expression level ranged from 36.7% to 259.0%; in rHS42, from 47.8% to 207.0%; in rHS32, from 47.9% to 105.4%; and in rHS2, from 6.1% to 172.1%, indicating a high variability of expression level in clones infected with recombinant virus lacking the insulator. In contrast, in clones infected with rIns/HS2/Ins, the range of expression of the human β-globin gene ranged from 52.8% to 58.3% of that in the mouse β maj -globin gene. These results indicate that the insulator functioned dramatically to reduce the variability of transgene expression due to the position effect. This insulator-rAAV vector system holds promise to provide a constant level of transgene expression for gene therapy, regardless of the insertion sites on the chromosome. Key words Gene therapy · β-Globin gene · Adeno-associated virus · Position effect · Insulator IntroductionThalassemias are a group of inherited anemias characterized by the reduced production of globin chains (Weatherall and Clegg 1981). This syndrome occurs worldwide. Patients with severe phenotypes require erythrocyte transfusions that can be associated with life-threatening iron overload, despite intensive chelation (Wolfe et al. 1985). Bone marrow transplantation has been performed with some success, but this treatment is feasible in only a small percentage of affected patients (Lucarelli et al. 1990;Ferster et al. 1992). Pharmacologic approaches are intended to increase γ-globin gene transcription, leading to more effective erythropoiesis and/or d...

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Establishment and characterization of KB cell lines latently infected with adeno-associated virus type 1

Cited by 94 publications

References 18 publications

Negative and positive regulation in trans of gene expression from adeno-associated virus vectors in mammalian cells by a viral rep gene product.

Negative and positive regulation in trans of gene expression from adeno-associated virus vectors in mammalian cells by a viral rep gene product.

Parvovirus Gene Regulation

Position-independent human β-globin gene expression mediated by a recombinant adeno-associated virus vector carrying the chicken β-globin insulator

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