Selective effects of E1B-defective adenoviruses and adenovirus E1A mutants in deficient mouse primary embryonic fibroblasts.
E1B-defective adenoviruses have been described as exerting selective cytopathic effects on transformed cells. Previously, we showed that adenovirus dl118, lacking both E1B proteins, very efficiently kills most human malignant cell lines. In order to study whether these selective effects were due to selective replication of dl118 in cells harboring specific genetic alterations, we compared the viability of various deficient mouse primary fibroblasts. We studied mouse embryonic fibroblasts (MEFs) derived from p16, p21, p27 and p53 knockout mice, as well as wild-type MEFs. We infected them with 100 p.f.u. of adenoviruses adl118, adwt300, and adenoviruses carrying the E1A mutant 922 (the E1a product only binds to the p300 and related proteins) and Ad646 (the E1A product binds to the pRb and related proteins). The percentage of infectivity was evaluated with an adenovirus carrying the green fluorescent protein (AdGFP). With AdGFP, clear green fluorescent signals were detected in more than 70% of the cells after 3 days of infection. After infection with several adenoviruses, we observed that E1A mutant 922 killed all the MEFs. Conversely, the E1a mutant Ad646 exerted its major effects on control wild-type MEFs. Moreover, Adl118 killed the wtMEFs and other MEFs slightly more efficiently than did wtAd, but less than Ad922. No viral replication was detected by adding the obtained supernatants to HEK293 cells. Due to the absence of significant viral replication on these cells, the results could be interpreted as direct effects of E1A and E1A mutant proteins on the different mouse cells carrying diverse genetic alterations.
Martin Duque
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P
Department of Pathology, Clinica Puerta de Hierro, Madrid, Spain.
Parada
C
C
Guinea
J
J
Sanchez-Prieto
R
R
Ramon Y Cajal
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S
D000256:Adenoviridae / Q000502:physiology*; D017500:Adenovirus E1A Proteins / Q000235:genetics*; D017501:Adenovirus E1B Proteins / Q000235:genetics*; D000818:Animals; D002460:Cell Line; D002470:Cell Survival; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000378:metabolism; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000378:metabolism; D003673:Defective Viruses / Q000502:physiology*; D004622:Embryo, Mammalian / Q000166:cytology; D005347:Fibroblasts / Q000378:metabolism / Q000821:virology*; D005434:Flow Cytometry; D049452:Green Fluorescent Proteins; D006801:Humans; D007124:Immunoenzyme Techniques; D008164:Luminescent Proteins / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D008840:Microfilament Proteins / Q000378:metabolism; D009124:Muscle Proteins; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D014779:Virus Replication
D017500:Adenovirus E1A Proteins; D017501:Adenovirus E1B Proteins; C493956:CDKN1A protein, human; C493957:Cdkn1a protein, mouse; D019941:Cyclin-Dependent Kinase Inhibitor p16; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D008164:Luminescent Proteins; D008840:Microfilament Proteins; D009124:Muscle Proteins; C058992:Tagln protein, mouse; D016159:Tumor Suppressor Protein p53; D049452:Green Fluorescent Proteins