Abstract:
:Adeno-associated virus (AAV) replication proteins Rep78 and Rep68 play major roles in the life cycle of AAV. We have recently provided in vivo evidence for the existence of a covalent association between Rep78 and virion single-stranded (ss) AAV DNA (K. M. R. Prasad and J.P. Trempe(1995) Virology 214, 360-370). In this work we have further characterized the Rep78 protein-AAV DNA covalent linkage. Exonuclease and primer extension analyses revealed that in the majority of isolated ssDNA, Rep78 protein is covalently linked to one of the 5' terminal thymidines. Pulse-chase experiments with radiolabeled methionine suggest that Rep protein remains associated with the virus particle for up to 8 hr after labeling in infected cells. Quantitative immunoprecipitation indicated that approximately 30% of the ssDNA remains associated with the Rep protein after cell fractionation and partial purification. When cells are infected with Rep-associated AAV particles, a significant proportion of viral DNA remains attached to Rep after entry into the nucleus. However, the linkage does not persist after nuclear entry. These observations suggest that covalently linked Rep78 protein may play a key role during wild-type AAV infections and AAV vector transductions.
journal_name
Virologyjournal_title
Virologyauthors
Prasad KM,Zhou C,Trempe JPdoi
10.1006/viro.1996.8431subject
Has Abstractpub_date
1997-03-03 00:00:00pages
183-92issue
1eissn
0042-6822issn
1096-0341pii
S0042-6822(96)98431-7journal_volume
229pub_type
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