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. 1999 Mar;73(3):2074-83.
doi: 10.1128/JVI.73.3.2074-2083.1999.

The replicative capacities of large E1B-null group A and group C adenoviruses are independent of host cell p53 status

A S Turnell  1 , R J Grand, P H Gallimore
Affiliations

The replicative capacities of large E1B-null group A and group C adenoviruses are independent of host cell p53 status

A S Turnell et al. J Virol. 1999 Mar.

Abstract

Recent reports suggest that an early region 1B (E1B) 55, 000-molecular-weight polypeptide (55K)-null adenovirus type 5 (Ad5) mutant (dl1520) can replicate to the same extent as wild-type (wt) Ad5 in cells either deficient or mutated in p53, implicating p53 in limiting viral replication in vivo. In contrast, we show here that the replicative capacity of Ad5 dl1520 is wholly independent of host cell p53 status, as is the replicative capacity of comparable Ad12 E1B 54K-null adenoviruses (Ad12 dl620 and Ad12 hr703). Furthermore, we show that there is no requirement for complex formation between p53 and Ad5 E1B 55K or Ad12 E1B 54K for a productive infection, such that wt Ad5 and wt Ad12 will both replicate in cells which are null for p53. In addition, we find that these Ad5 and Ad12 mutant viruses induce S phase irrespective of the p53 status of the cell and that, therefore, S-phase induction does not correlate with the replicative capacity of the virus. Interestingly, the replicative capacities of the large E1B-null adenoviruses correlated positively with the ability to express E1B 19K and were related to the ability to repress premature adenovirus-induced apoptosis. Infection of primary human cells indicated that Ad5 dl1520, wt Ad5, and wt Ad12 replicated better in cycling normal human skin fibroblasts (HSFs) than in quiescent HSFs. Thus, the cell cycle status of the host cell, upon infection, also influences viral yield.

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Figures

FIG. 1
FIG. 1
Replicative capacities of Ad5 dl1520 and wt Ad5 in cells differing in p53 status. Virus yields were quantified as stated in Materials and Methods, by using permissive Ad5 HEK 293 cells. (A) Virus yield from Ad5 dl1520- and wt Ad5-infected cells at 48 h p.i. Lane 1, A549 cells (wt p53); lane 2, HCT116 cells (wt p53); lane 3, C33A cells (mutant p53); lane 4, HaCaT cells (mutant p53); lane 5, HT-29 cells (mutant p53); lane 6 H1299 cells (p53 null); lane 7, JW2 cells (p53 null). (B) Same as for panel A, except values were taken at 72 h p.i. The values above each panel reflect the fold decrease in replicative capacity of Ad5 dl1520 relative to wt Ad5. Results are means from two independent experiments.
FIG. 2
FIG. 2
Replicative capacities of Ad12 dl620, Ad12 hr703, Ad12 pm700 and wt Ad12 in cells differing in p53 status. Virus yields shown at 72 h p.i., were quantified as stated in Materials and Methods, by using permissive Ad12 HER3 cells. Lane 1, A549 cells (wt p53); lane 2, HCT116 cells (wt p53); lane 3, C33A cells (mutant p53); lane 4, HaCaT cells (mutant p53); lane 5, HT-29 cells (mutant p53); lane 6 H1299 cells (p53 null); lane 7, JW2 cells (p53 null). Results are means from two independent experiments.
FIG. 3
FIG. 3
S-phase induction by Ad5 dl1520 and wt Ad5. (A) Percentages of cells in S + G2/M from Ad5 dl1520- and wt Ad5-infected cells at 48 h p.i. Lane 1, A549 cells (wt p53); lane 2, HCT116 cells (wt p53); lane 3, C33A cells (mutant p53); lane 4, HaCaT cells (mutant p53); lane 5, HT-29 cells (mutant p53); lane 6 H1299 cells (p53 null); lane 7, JW2 cells (p53 null). (B) Same as for panel A, except values were taken at 72 h p.i. Results are means from two independent experiments.
FIG. 4
FIG. 4
S-phase induction by Ad12 dl620, Ad12 hr703, Ad12 pm700, and wt Ad12. Percentages of cells in S + G2/M at 72 h p.i. are shown. Lane 1, A549 cells (wt p53); lane 2, HCT116 cells (wt p53); lane 3, C33A cells (mutant p53); lane 4, HaCaT cells (mutant p53); lane 5, HT-29 cells (mutant p53); lane 6 H1299 cells (p53 null); lane 7, JW2 cells (p53 null). Results are means from two independent experiments.
FIG. 5
FIG. 5
Expression of adenovirus early gene products at 24, 48, and 72 h p.i. in Ad5 dl1520- and wt Ad5-infected cells. (A) A549 cells (wt p53). (B) HCT116 cells (wt p53). (C) C33A cells (mutant p53). (D) HaCaT cells (mutant p53). (E) HT-29 cells (mutant p53). (F) H1299 cells (p53 null). (G) JW2 cells (p53 null). Lanes 1 to 3, Ad5 dl1520-infected cells; lanes 4 to 6, wt Ad5-infected cells. Panels i, Ad5 E1A; panels ii, Ad5 E1B 55K; panels iii, Ad5 E1B 19K.
FIG. 6
FIG. 6
Expression of adenovirus early gene products at 24, 48, and 72 h p.i. in Ad12 dl620-, Ad12 hr703-, Ad12 pm700-, and wt Ad12-infected cells. (A) A549 cells (wt p53). (B) HCT116 cells (wt p53). (C) C33A cells (mutant p53). (D) HaCaT cells (mutant p53). (E) HT-29 cells (mutant p53). (F) H1299 cells (p53 null). (G) JW2 cells (p53 null). Lanes 1 to 3, Ad12 dl620-infected cells; lanes 4 to 6, Ad12 hr703-infected cells; lanes 7 to 9, Ad12 pm700-infected cells; lanes 10 to 12, wt Ad12-infected cells. Panels i, Ad12 E1A; panels ii, Ad12 E1B 54K; panels iii, Ad12 E1B 19K.
FIG. 7
FIG. 7
Replicative capacities of Ad5 and Ad12 in cycling and quiescent HSFs. Virus yields were determined at 72 h p.i., as described in Materials and Methods, by using permissive Ad5 HEK 293 cells and Ad12 HER3 cells for Ad5 and Ad12, respectively. Lanes 1 and 2, growing HSFs; lanes 3 and 4, quiescent HSFs; lanes 1 and 3, Ad5 dl1520 and wt Ad5; lanes 2 and 4, Ad12 dl620 and wt Ad12. The values above the figure reflect the fold decrease in replicative capacity of Ad5 dl1520 relative to wt Ad5 and of Ad12 dl620 relative to wt Ad12. Results are means from two independent experiments.
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