Identification of an Adeno-Associated Virus Rep Protein Binding Site in the Adenovirus
            <i>E2a</i>
            Promoter

Casper, Jonathan D.; Timpe, Jennifer M.; Dignam, John David; Trempe, James P.

doi:10.1128/jvi.79.1.28-38.2005

Cited by 15 publications

(19 citation statements)

References 70 publications

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“…Since E2a and E4 proteins are essential for efficient Ad DNA amplification, we considered the possibility that Repmediated E2a and E4 transcriptional regulation contributes to decreased Ad DNA replication. Previous studies suggested that AAV Rep proteins interact with the E2A promoter and regulate its activity (7). Also, our studies indicate that E4 mRNA levels are dramatically reduced at 24 hpi during coinfection with AAV (Fig.…”

supporting

confidence: 61%

Effects of Adeno-Associated Virus on Adenovirus Replication and Gene Expression during Coinfection

Timpe

Verrill

Trempe

2006

J Virol

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Adeno-associated virus (AAV) is a nonpathogenic parvovirus that requires adenovirus (Ad) or another helper virus for a fully permissive infection. AAV-mediated inhibition of Ad is well documented, yet many details of this interaction remain unclear. In this study, we observed a maximum 50-fold decrease in infectious virus production and a 10-to 40-fold reduction in Ad DNA synthesis during coinfections with AAV. With the exception of the E3 gene, AAV decreased all steady-state Ad mRNA levels at 24 h postinfection (hpi) in a dose-dependent manner. However, not all transcription units were affected equally. E4 and late transcription were the most strongly inhibited, and E1A and E2A were the least affected. The temporal effects of AAV on Ad mRNA transcript levels also varied among the Ad genes. Ad protein expression paralleled mRNA levels at 24 hpi, suggesting that coinfecting AAV does not exert substantial effects on translation. In plasmid transfection assays, Rep78 protein most effectively limited Ad amplification, while Rep40 had no effect. Since E2a and E4 proteins are essential for efficient Ad DNA amplification, we examined the relationship between reduced E2A and E4 expression and decreased DNA amplification. Transfected Rep78 did not reduce E2A and E4 transcript levels prior to DNA replication. Also, AAV-induced inhibition of E2A and E4 mRNA production did not occur in the presence of hydroxyurea. It is therefore unlikely that decreased early gene expression is solely responsible for AAV's suppression of Ad DNA replication. Our results suggest that AAV amplification and/or Rep gene expression inhibits Ad DNA synthesis.

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supporting

confidence: 61%

Effects of Adeno-Associated Virus on Adenovirus Replication and Gene Expression during Coinfection

Timpe

Verrill

Trempe

2006

J Virol

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“…DNase I protection assays were performed as described previously (10). The AdMLP fragment was obtained by PCR amplification of a 336-bp DNA between nt 5893 and 6229 of the Ad5 genome using the following primers: 5Ј-GTCGTTGTCCACTAGG-3Јand 5Ј-GATTGTCTTTTCT GACCAG-3Ј.…”

Section: Methodsmentioning

confidence: 99%

“…AAV and the Rep proteins also regulate Ad gene expression during coinfection. The Rep78/68 proteins bind to the Ad E2a promoter, reducing the amount of E2a mRNA (10,30,40). Rep proteins interact with cellular proteins involved in transcription regulation, including Sp1 (25,45), high-mobility-group nonhistone protein 1 (15), the transcriptional coactivator PC4 (54), TATA-binding protein (26,51), a putative protein kinase (protein kinase X or PKX), and protein kinase A (PKA) (13,17).…”

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confidence: 99%

Adeno-Associated Virus Rep Protein-Mediated Inhibition of Transcription of the Adenovirus Major Late Promoter In Vitro

Needham

Casper

Kalman‐Maltese

et al. 2006

J Virol

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“…Besides inhibiting HSV-1 replication, AAV-2 Rep has also been shown to have inhibitory effects on its other helper virus, Ad [28], as well as on the host cell [161,162]. The inhibitory effect of Rep on Ad has been suggested to occur primarily at the level of DNA replication [163], although several effects on Ad gene expression have also been demonstrated [164][165][166][167]. Although it is thought that the inhibitory effect of AAV-2 on its helper viruses contributes to the replication fitness of AAV-2 in coinfected cells, it can also lead to autoinhibition.…”

Section: Effects Of Aav-2 Rep On Hsv-1 Replication and The Host Cellmentioning

confidence: 99%

Interactions Between AAV-2 and HSV-1: Implications for Hybrid Vector Design

Glauser

Fraefel

2011

Future Virol.

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Herpes simplex virus type 1 (HSV-1)-based amplicon vectors have a transgene capacity of up to 150 kbp and can efficiently transduce many different cell types in culture and in vivo without causing cytopathic effects. However, these vectors do not support long-term transgene expression. Adeno-associated virus type 2 (AAV-2) has the capacity to integrate its genome into a specific site on human chromosome 19, but AAV-2-derived gene therapy vectors have a transgene capacity of only 4.5 kb. To combine the large transgene capacity of HSV-1 with the potential for site-specific genomic integration and long-term transgene expression of AAV-2, HSV/AAV hybrid vectors have been developed. This review describes the design, applications and limitations of these hybrid vectors. However, as HSV-1 is a full helper virus for AAV-2 replication, the main focus is the analysis of the molecular mechanisms of interaction between the two viruses. The knowledge of these interactions will have direct implications on the design of novel HSV/AAV hybrid vectors.

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Identification of an Adeno-Associated Virus Rep Protein Binding Site in the Adenovirus E2a Promoter

Cited by 15 publications

References 70 publications

Effects of Adeno-Associated Virus on Adenovirus Replication and Gene Expression during Coinfection

Effects of Adeno-Associated Virus on Adenovirus Replication and Gene Expression during Coinfection

Adeno-Associated Virus Rep Protein-Mediated Inhibition of Transcription of the Adenovirus Major Late Promoter In Vitro

Interactions Between AAV-2 and HSV-1: Implications for Hybrid Vector Design

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