Multiplication of adeno-associated virus type 1 in cells coinfected with a temperature-sensitive mutant of human adenovirus type 31
Handa, H.; Shimojo, H.; Yamaguchi, K.
Virology 74(1): 1-15
ISSN/ISBN: 0042-6822 PMID: 982809 DOI: 10.1016/0042-6822(76)90123-9
A temperature-sensitive mutant (tsA13) of human adenovirus type 31 (H31), defective in viral DNA replication, was able to support growth of adeno-associated virus type 1 (AAV1) at the nonpermissive temperature (40.degree. C). With the use of this system, the multiplication of AAV1 and AAV1-specific changes were investigated. The latent period of AAV1 growth was shortened by preinfection of cells with H31tsA13 10 h before superinfection with AAV1. The rate of DNA synthesis began to rise at about 6 h postinfection (p.i.) with AAV1 and reached its maximum at 16 h p.i. In cells coinfected with H31tsA13 and AAV1, only AAV1 DNA was detected without the presence of adenovirus DNA at 40.degree. C. Replicative intermediates of AAV1 DNA were larger than AAV1 DNA in neutral and alkaline sucrose gradients. Specific inclusions induced by AAV1 were observed in the nucleus of coinfected and stained cells. Microscopic autoradiogram of coinfected cells revealed that grains (viral DNA) were found before the appearance of the inclusions of AAV1 in the interior of the nucleus. The AAV1 virion antigen 1st appeared in the nucleus at about 6 h p.i. with AAV1 and spread into the cytoplasm within 12 h p.i. EM examination of infected cells revealed that the inclusions were aggregates or crystalline arrays of AAV1 particles in the nucleus. Neither adenovirus inclusions nor particles were observed. AAV1 DNA replication proceeded in the presence of cycloheximide. The time interval between AAV1 infection and the peak of DNA synthesis became shorter, when AAV1 was superinfected at 16 h or later after infection with H31tsA13. AAV1 apparently lacks its own early protein and the lack of the early protein is complemented by a factor(s) induced in adenovirus-infected cells. [Human embryonic kidney and green monkey kidney cells were used.].