Constructing chimeric type 12/type 5 adenovirus E1A genes and using them to identify an oncogenic determinant of adenovirus type 12

Telling, Glenn C.; Williams, Jim

doi:10.1128/jvi.68.2.877-887.1994

Cited by 25 publications

(9 citation statements)

References 80 publications

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“…Moreover, their findings implied that CR2 of Ad12 E1A, which is critical for the transforming activity of both E1A proteins, is not involved in the differential transforming efficiencies of Ad5 and Ad12 and that cellular proteins whose interactions with CR2 of Ad5 E1A are critical for transformation interact equally well with CR2 of Ad5 E1A and Ad12 E1A (27), further supporting our idea that the E1A transforming function may be localized in E1A regions different from known cellular protein binding sites (28). In this context, the Ad12 spacer region bordered by CR2 and CR3 in concert with Ad12 E1A sequences to the left of CR2 is of particular interest and was discussed as an important determinant of viral tumorigenicity (28,55). Interestingly, this region or a similar region which is characterized by a stretch of alanine residues was identified in highly oncogenic adenoviruses.…”

Section: Discussionsupporting

confidence: 75%

E1A 12S and 13S of the transformation-defective adenovirus type 12 strain CS-1 inactivate proteins of the RB family, permitting transactivation of the E2F-dependent promoter

Pützer

Rumpf

Rega

et al. 1997

J Virol

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The transformation-defective Vero cell host range mutant CS-1 of the highly oncogenic adenovirus type 12 (Ad12) (Ad12-CS-1) has a 69-bp deletion in the early region 1A (E1A) gene that removes the carboxy-terminal half of conserved region 2 and the amino-terminal half of the Ad12-specific so-called spacer that seems to play a pivotal role in the oncogenicity of the virus. Despite its deficiency in immortalizing and transforming primary rodent cells, we found that the E1A 13S protein of Ad12-CS-1 retains the ability to bind p105-RB, p107, and p130 in nuclear extract binding assays with glutathione S-transferase-E1A fusion proteins and Western blot analysis. Like wild-type E1A, the mutant protein was able to dissociate E2F from retinoblastoma-related protein-containing complexes, as judged from gel shift experiments with purified 12S and 13S proteins from transfection experiments with an E1A expression vector or from infection with the respective virus. Moreover, in transient expression assays, the 12S and 13S products of wild-type Ad12 and Ad12-CS-1 were shown to transactivate the Ad12 E1A promoter containing E2F-1 and E2F-5-motifs, respectively, in a comparable manner. The same results were obtained from transfection assays with the E2F motif-dependent E2 promoter of adenovirus type 5 or the human dihydrofolate reductase promoter. These data suggest that efficient infection by Ad12 and the correlated virus-induced reprogramming of the infected cells, including the induction of cell cycle-relevant mechanisms (e.g. E2F activation), can be uncoupled from the transformation properties of the virus.

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Section: Discussionsupporting

confidence: 75%

E1A 12S and 13S of the transformation-defective adenovirus type 12 strain CS-1 inactivate proteins of the RB family, permitting transactivation of the E2F-dependent promoter

Pützer

Rumpf

Rega

et al. 1997

J Virol

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“…1B). Between conserved regions 2 (CR2) and 3 (CR3), the 13S E1A protein of subgroup A virus Ad12 possesses an alanine-rich spacer region which is, in part, responsible for the highly oncogenic phenotype of this virus (33,53). In contrast, the Ad9 13S E1A protein was found to contain a non-alanine-rich spacer region similar to the one present in 13S E1A proteins of weakly oncogenic subgroup B adenoviruses (Fig.…”

Section: Gene Organization and Predicted Polypeptides Of The Ad9 E1 Rmentioning

confidence: 97%

“…(B) Comparison of the Ad9 13S, 12S, and 10S E1A polypeptide sequences. CR1, CR2, spacer region, and CR3 are indicated (22,33,48,53).…”

Section: Fig 1 (A)mentioning

confidence: 99%

Early Region 1 Transforming Functions Are Dispensable for Mammary Tumorigenesis by Human Adenovirus Type 9

Thomas

Shin²,

Jiang

et al. 1999

J Virol

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Some human adenoviruses are tumorigenic in rodents. Subgroup A and B human adenoviruses generally induce sarcomas in both male and female animals, and the gene products encoded within viral early region 1 (E1 region) are both necessary and sufficient for this tumorigenicity. In contrast, subgroup D human adenovirus type 9 (Ad9) induces estrogen-dependent mammary tumors in female rats and requires the E4 region-encoded ORF1 oncoprotein for its tumorigenicity. Considering the established importance of the viral E1 region for tumorigenesis by adenoviruses, we investigated whether this viral transcription unit is also necessary for Ad9 to generate mammary tumors. The nucleotide sequence of the Ad9 E1 region indicated that the gene organization and predicted E1A and E1B polypeptides of Ad9 are closely related to those of other human adenovirus E1 regions. In addition, an Ad9 E1 region plasmid demonstrated focus-forming activity in both low-passage-number and established rat embryo fibroblasts, whereas a large deletion within either the E1A or E1B gene of this plasmid diminished transforming activity. Surprisingly, we found that introducing the same transformation-inactivating E1A and E1B deletions into Ad9 results in mutant viruses that retain the ability to elicit mammary tumors in rats. These results are novel in showing that Ad9 represents a unique oncogenic adenovirus in which the E4 region, rather than the E1 region, encodes the major oncogenic determinant in the rat.

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“…This indicates that the six alanines in the spacer region of Ad12 E1A are necessary for stable transformation of primary mouse cells. Interestingly, the same region is thought to be essential for the oncogenic properties of E1A, as judged from Ad5/12 hybrid virus experiments (Telling and Williams, 1994;Williams, 1995).…”

Section: Transformation Of Primary Mouse Cellsmentioning

confidence: 99%

“…1A) and exhibit a considerable homology to transcriptional suppressor proteins found in the embryonic development of Drosophila (Licht et al, 1990). Ad5/Ad12 hybrid virus experiments have shown that the integrity of the spacer is important for the oncogenicity of the virus (Jelinek et al, 1994;Telling and Williams, 1994). Despite its additional oncogenic properties, the transforma-tion rate of primary rat cells induced by transfection with Ad12 E1A/E1B was ϳ80% lower as for Ad2 E1A/Ad12 E1B (Leclére et al, 1993).…”

Section: Introductionmentioning

confidence: 99%

Two Domains within the Adenovirus Type 12 E1A UniqueSpacerHave Disparate Effects on the Interaction of E1A with P105-Rb and the Transformation of Primary Mouse Cells

1999

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Transformation of primary rodent cells by functions of the adenovirus type 12 (Ad12) early region 1 (E1) is reduced severalfold compared with transformation by E1 of Ad2. We analyzed whether the unique spacer region of Ad12 E1A that borders the conserved region (CR) 2 and represents an oncogenic determinant of Ad12 E1A is involved in this impaired transformation property, putatively by modulating transformation-relevant biological E1A functions. We show that a mutant (E1ASpm1) that lacks 12 amino-terminal residues of the spacer binds p105-Rb and p130 as Ad12 E1A wild type (E1Awt), whereas a second spacer mutant (E1ASpm2) that lacks an adjacent stretch of six alanines exhibits highly reduced binding to p105-Rb. The binding of this mutant to the p130 pocket protein is, however, little impaired. E1ASpm1 diminishes the formation of the p105-Rb-E2F complex more efficiently than E1Awt or, least efficient, E1ASpm2. These properties of the spacer mutants to target and to disintegrate the p105-Rb-E2F complex correspond with their ability to transform primary mouse cells in combination with E1B: E1ASpm1 (plus Ad12 E1B)-transfected cells could be easily established as cell lines, comparable to Ad12 E1Awt- or Ad2 E1Awt-transfected cells. In contrast, cells transfected with E1ASpm2 or Ad12 E1AdelCR2 (lacking the entire CR2) died within 6-10 weeks after replating, although foci were formed in all cases. Of note, the E1ASpm1-transformed cells grow as fast as the Ad2 E1Awt-transformed cells, with a doubling rate of 15 h, whereas the doubling of the Ad12 E1Awt-transformed cells takes approximately 120 h. Moreover, in the established cell lines, the affinity of E1ASpm1 to p105-Rb was higher than with that of E1Awt. Our data suggest the presence of a transformation-suppressing domain within the carboxyl-terminal 12 residues of the Ad12 E1A-unique spacer, whereas the hydrophobic stretch of six alanines in the spacer is required for stable transformation.

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Constructing chimeric type 12/type 5 adenovirus E1A genes and using them to identify an oncogenic determinant of adenovirus type 12

Cited by 25 publications

References 80 publications

E1A 12S and 13S of the transformation-defective adenovirus type 12 strain CS-1 inactivate proteins of the RB family, permitting transactivation of the E2F-dependent promoter

E1A 12S and 13S of the transformation-defective adenovirus type 12 strain CS-1 inactivate proteins of the RB family, permitting transactivation of the E2F-dependent promoter

Early Region 1 Transforming Functions Are Dispensable for Mammary Tumorigenesis by Human Adenovirus Type 9

Two Domains within the Adenovirus Type 12 E1A UniqueSpacerHave Disparate Effects on the Interaction of E1A with P105-Rb and the Transformation of Primary Mouse Cells

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