Adenovirus Transforming Protein E1A Induces c-Myc in Quiescent Cells by a Novel Mechanism

Kadeppagari, Ravi-Kumar; Natesan, Sankar; Thimmapaya, Bayar

doi:10.1128/jvi.02145-08

Cited by 16 publications

(20 citation statements)

References 35 publications

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“…E1A binds to p300, dissociates the repressor complex, and then induces Myc. In vitro studies also supported this observation (31). In this paper, we showed that blocking the Myc induction that occurs in E1A-expressing quiescent cells significantly reduces the S-phase induction.…”

Section: Discussionsupporting

confidence: 71%

“…This repressor complex binds to the upstream YY1 site of the Myc promoter to repress transcription and prevent premature G 1 exit of the growth-arrested cells. E1A induces Myc by binding to p300 and dissociating the repressor complex (31,43). Induction of Myc by this mechanism by E1A contributes to the induction of the S phase.…”

Section: E1amentioning

confidence: 99%

“…Preparation of whole-cell extracts and isolation of chromatinbound proteins. To prepare the whole-cell extracts, cells were seeded overnight at a density of 3 ϫ 10 6 cells per 100-mm-diameter dish and then starved for 32 h. They were then infected with the indicated viruses, serum stimulated where appropriate, and lysed in 300 l of buffer containing NP-40 for 15 min on ice as described earlier (26,31). The cell extracts were clarified by centrifuging at 30,000 ϫ g for 40 min.…”

Section: Cells and Virusesmentioning

confidence: 99%

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Adenovirus E1A Oncogene Induces Rereplication of Cellular DNA and Alters DNA Replication Dynamics

Singhal

Leo

Setty

et al. 2013

J Virol

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The oncogenic property of the adenovirus (Ad) transforming E1A protein is linked to its capacity to induce cellular DNA synthesis which occurs as a result of its interaction with several host proteins, including pRb and p300/CBP. While the proteins that contribute to the forced induction of cellular DNA synthesis have been intensively studied, the nature of the cellular DNA replication that is induced by E1A in quiescent cells is not well understood. Here we show that E1A expression in quiescent cells leads to massive cellular DNA rereplication in late S phase. Using a single-molecule DNA fiber assay, we studied the cellular DNA replication dynamics in E1A-expressing cells. Our studies show that the DNA replication pattern is dramatically altered in E1A-expressing cells, with increased replicon length, fork velocity, and interorigin distance. The interorigin distance increased by about 3-fold, suggesting that fewer DNA replication origins are used in E1A-expressing cells. These aberrant replication events led to replication stress, as evidenced by the activation of the DNA damage response. In earlier studies, we showed that E1A induces c-Myc as a result of E1A binding to p300. Using an antisense c-Myc to block c-Myc expression, our results indicate that induction of c-Myc in E1A-expressing cells contributes to the induction of host DNA replication. Together, our results suggest that the E1A oncogene-induced cellular DNA replication stress is due to dramatically altered cellular replication events and that E1A-induced c-Myc may contribute to these events. The adenovirus (Ad) transforming E1A protein [a 243-aminoacid E1A protein, also referred to as small E1A protein [1,2]) has the capacity to induce S phase in quiescent cells, and in the presence of activated ras or virus-encoded E1B19K or 55K proteins, E1A can transform rodent cells in culture (1, 2). The S-phase induction and cell transformation activities of the small E1A protein are genetically linked and are dependent on the N-terminal region of E1A binding to cellular protein complexes, including TRRAP/p400/GCN5, histone acetyltransferase p300/CBP, and the Rb family tumor suppressor proteins (1-4). E1A-Rb interactions result in the release of the progrowth E2F family transcription factors from the Rb-histone deacetylase (HDAC) repressor complexes and the induction of the S phase (1, 5). However, studies have shown that in order for E1A to induce S phase efficiently, it must bind to p300/CBP and Rb family proteins simultaneously, suggesting that E1A must also alter the functions of p300/CBP (3, 6).Although a large number of studies have focused on the cellular proteins that contribute to the forced induction of host DNA synthesis in E1A-expressing cells, the nature of the cellular DNA that replicates in these cells is not well understood. Previous studies have shown that the E1A-expressing cells fail to undergo proper mitosis and that such cells accumulate in the S and G 2 /M phases (7-10). Mammalian cells contain a large number of DNA replication origins, a...

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Section: Discussionsupporting

confidence: 71%

Section: E1amentioning

confidence: 99%

Section: Cells and Virusesmentioning

confidence: 99%

See 1 more Smart Citation

Adenovirus E1A Oncogene Induces Rereplication of Cellular DNA and Alters DNA Replication Dynamics

Singhal

Leo

Setty

et al. 2013

J Virol

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“…47 Our data shows how the upregulation of Chk1 correlates with an increase in the activation of this kinase (Ser345) and also with an increase in cyclin B1 levels, consistent with mitotic arrest. However, other possibilities, such as reduction in the length of G 1 phase, as has been proposed in quiescent cells due to the effects of E1a onto c-Myc and pRB 48 through specific mechanisms, 49 should be considered to fully understand the effect of E1a onto cell cycle progression. In addition, we demonstrate that E1a mediates upregulation of Chk1 at the transcriptional level trough c-Myc (see below), as q-RT-PCR and luciferase assays supports.…”

Section: Discussionmentioning

confidence: 99%

E1a promotes c-Myc-dependent replicative stress

et al. 2013

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“…A future study will investigate whether E4-ORF1 represents the undetermined viral protein responsible for the second phase of Ras/Mek/Erk signaling required for optimal Ad protein expression and virion production in Ad-infected cells (32). Multiple DNA virus proteins, including Ad E1A and E4-ORF1, are reported to increase Myc expression (6,(54)(55)(56)(57). Here we showed that E4-ORF1 sustains Myc protein expression in human epithelial cells cultured in serum and growth factor-depleted medium and that this novel E4-ORF1 function is mediated by the combined activities of EGFR, InsR/IGF1R, and PI3K (Fig.…”

Section: Discussionmentioning

confidence: 99%

Adenovirus E4-ORF1 Dysregulates Epidermal Growth Factor and Insulin/Insulin-Like Growth Factor Receptors To Mediate Constitutive Myc Expression

Kong

Kumar

Taruishi

et al. 2015

J Virol

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The E4-ORF1 protein encoded by human adenovirus stimulates viral replication in human epithelial cells by binding and activating cellular phosphatidylinositol 3-kinase (PI3K) at the plasma membrane and cellular Myc in the nucleus. In this study, we showed that E4-ORF1 hijacks the tyrosine kinase activities of cellular epidermal growth factor receptor (EGFR) and insulin receptor (InsR)/insulin-like growth factor receptor 1 (IGF1R), as well as the lipid kinase activity of PI3K, to mediate constitutive Myc protein expression. We additionally demonstrated that EGFR contributes to constitutive Myc expression through the capacity of E4-ORF1 to induce ligand-independent EGFR activation and stimulation of the Ras/Mek/Erk pathway, the latter activity of which was conserved by human adenoviruses. Results further suggested that EGFR normally forms a complex with the cellular PDZ protein Discs Large 1 (Dlg1), a component of the Dlg1:E4-ORF1:PI3K ternary complex that mediates E4-ORF1-induced PI3K activation, and that E4-ORF1 binds the Dlg1:EGFR complex and promotes the association of EGFR with InsR and IGF1R. In addition to its role in constitutive Myc expression, InsR/IGF1R also negatively regulates EGFR autophosphorylation and EGFR-mediated Ras/Mek/Erk signaling, and data suggested that E4-ORF1 binding to Dlg1 antagonizes these activities. Collectively, our findings suggest that in human epithelial cells, E4-ORF1 targets EGFR, InsR/IGF1R, and PI3K at the plasma membrane to activate cytosolic signaling pathways that sustain Myc protein levels in the nucleus. We postulate that E4-ORF1-induced constitutive Myc expression functions to ensure the formation of nuclear E4-ORF1:Myc complexes, which have been shown to activate Myc and to enhance adenovirus replication. O ver 60 human adenovirus (Ad) serotypes are classified into seven subgroups (subgroups A through G) based on oncogenic, hemagglutination, and virion structural protein properties as well as genome sequence similarity (1). Ad is a human pathogen primarily associated with mild, self-limited respiratory diseases but additionally causes gastroenteritis, conjunctivitis, cystitis, and skin rashes (1). Certain viral serotypes, such as Ad type 14 (Ad14), can cause life-threatening symptoms that require hospitalization for up to 40% of otherwise healthy individuals (2), and Ad infections of neonates and immunosuppressed patients are associated with high morbidity and mortality rates (3). However, current treatments for severe Ad infections are controversial and carry significant risks for adverse events (1). Ad is also exploited as a vector for vaccination and gene and cancer therapy as well as a tool of discovery to reveal fundamental mechanisms of the cell and mechanisms for cancer development (1).The Ad early region 4 open reading frame 1 gene (E4-ORF1) encodes a 14-kDa protein required for optimal viral replication (4-6). In humans, subgroup D Ad9 is associated with eye infections (1), whereas Ad9 inoculation into experimental animals elicits estrogen-dependent m...

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Adenovirus Transforming Protein E1A Induces c-Myc in Quiescent Cells by a Novel Mechanism

Cited by 16 publications

References 35 publications

Adenovirus E1A Oncogene Induces Rereplication of Cellular DNA and Alters DNA Replication Dynamics

Adenovirus E1A Oncogene Induces Rereplication of Cellular DNA and Alters DNA Replication Dynamics

E1a promotes c-Myc-dependent replicative stress

Adenovirus E4-ORF1 Dysregulates Epidermal Growth Factor and Insulin/Insulin-Like Growth Factor Receptors To Mediate Constitutive Myc Expression

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