Altering the Ad5 packaging domain affects the maturation of the ad particles
Alba Fernández, Raúl (Universitat Autònoma de Barcelona. Centre de Biotecnologia Animal i de Teràpia Gènica (CBATEG))
Cots i Rabella, Dan (Universitat Autònoma de Barcelona. Centre de Biotecnologia Animal i de Teràpia Gènica (CBATEG))
Ostapchuk, Philomena (Stony Brook University. Department of Molecular Genetics and Microbiology (Nova York, Estats Units d'Amèrica))
Bosch i Merino, Assumpció (Universitat Autònoma de Barcelona. Departament de Bioquímica i de Biologia Molecular)
Hearing, Patrick (Stony Brook University. Department of Molecular Genetics and Microbiology (Nova York, Estats Units d'Amèrica))
Chillón Rodríguez, Miguel (Universitat Autònoma de Barcelona. Centre de Biotecnologia Animal i de Teràpia Gènica (CBATEG))

Data:	2011
Resum:	We have previously described a new family of mutant adenoviruses carrying different combinations of attB/attP sequences from bacteriophage PhiC31 flanking the Ad5 packaging domain. These novel helper viruses have a significantly delayed viral life cycle and a severe packaging impairment, regardless of the presence of PhiC31 recombinase. Their infectious viral titers are significantly lower (100-1000 fold) than those of control adenovirus at 36 hours post-infection, but allow for efficient packaging of helper-dependent adenovirus. In the present work, we have analyzed which steps of the adenovirus life cycle are altered in attB-helper adenoviruses and investigated whether these viruses can provide the necessary viral proteins in trans. The entry of attB-adenoviral genomes into the cell nucleus early at early timepoints post-infection was not impaired and viral protein expression levels were found to be similar to those of control adenovirus. However, electron microscopy and capsid protein composition analyses revealed that attB-adenoviruses remain at an intermediate state of maturation 36 hours post-infection in comparison to control adenovirus which were fully mature and infective at this time point. Therefore, an additional 20-24 hours were found to be required for the appearance of mature attB-adenovirus. Interestingly, attB-adenovirus assembly and infectivity was restored by inserting a second packaging signal close to the right-end ITR, thus discarding the possibility that the attB-adenovirus genome was retained in a nuclear compartment deleterious for virus assembly. The present study may have substantive implications for helper-dependent adenovirus technology since helper attB-adenovirus allows for preferential packaging of helper-dependent adenovirus genomes.
Ajuts:	European Commission 222992 Instituto de Salud Carlos III PS09/02674 Instituto de Salud Carlos III PI081162 Agència de Gestió d'Ajuts Universitaris i de Recerca 2009/SGR-1300
Drets:	Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, la comunicació pública de l'obra i la creació d'obres derivades, fins i tot amb finalitats comercials, sempre i quan es reconegui l'autoria de l'obra original.
Llengua:	Anglès
Document:	Article ; Versió publicada
Matèria:	DNA replication ; Virions ; Viral genome ; Bacterial genomics ; Vector-borne diseases ; Genomic signal processing
Publicat a:	PloS one, Vol. 6, Issue 5 (May 2011) , p. e19564, ISSN 1932-6203

DOI: 10.1371/journal.pone.0019564
PMID: 21611162

10 p, 1.3 MB

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