Effect of deletions in adenovirus early region 1 genes upon replication of adeno-associated virus

Abstract

The growth of adeno-associated virus (AAV) is dependent upon helper functions provided by adenovirus. We investigated the role of adenovirus early gene region 1 in the AAV helper function by using six adenovirus type 5 (Ad5) host range mutants having deletions in early region 1. These mutants do not grow in human KB cells but are complemented by and grow in a line of adenovirus-transformed human embryonic kidney cells (293 cells); 293 cells contain and express the Ad5 early region 1 genes. Mutants having extensive deletions of adenovirus early region 1a (dl312) or regions 1a and 1b (dl313) helped AAV as efficiently as wild-type adenovirus in 293 cells, but neither mutant helped in KB cells. No AAV DNA, RNA, or protein synthesis was detected in KB cells in the presence of the mutant adenoviruses. Quantitative blotting experiments showed that at 20 h after infection with AAV and either dl312 or dl313 there was less than one AAV genome per cell. In KB cells infected with AAV alone, the unreplicated AAV genomes were detected readily. Apparently, infection with adenovirus mutant dl312 or dl313 results in degradation of most of the infecting AAV genomes. We suggest that at least an adenovirus region 1b product (and perhaps a region 1a product also) is required for AAV DNA replication. This putative region 1b function appears to protect AAV DNA from degradation by an adenovirus-induced DNase. We also tested additional Ad5 mutants (dl311, dl314, sub315, and sub316). All of these mutants were inefficient helpers, and they showed varying degrees of multiplicity leakiness. dl312 and dl313 complemented each other for the AAV helper function, and each was complemented by Ad5ts125 at the nonpermissive temperature. The defect in region 1 mutants for AAV helper function acts at a different stage of the AAV growth cycle than the defect in the region 2 mutant ts125.

Detection of Infectious Adeno-Associated Virus Particles in Human Cervical Biopsies

“…Adenovirus has been described as helper for AAV by coinfecting cultured cells of different origin (Carter and Laughlin, 1983;Janik et al, 1981;Laughlin et al, 1982;Richardson and Westphal, 1984). Other helpers of AAV including herpes simplex virus types I and II (Mishra and Rose, 1990;Weindler and Heilbronn, 1991;Buller et al, 1981), cytomegalovirus (McPherson et al, 1985), and vacciniavirus (Schlehofer et al, 1986) were found to provide AAV helper functions in vitro.…”

Parvoviruses: agents of distinct pathogenic and molecular potential

“…However, as shown by Tratschin et al [64] for the P19 promotor, it might also activate dependovirus genes directly. Early region EIB seems to be important for efficient accumulation of AAV RF DNA, possibly by protecting vDNA against degradation via a so-far undefined function expressed in adenovirus-infected cells [65]. In addition, Ostrove and Berns [66] reported that E1B is required for rescue of AAV DNA from the integrated state in human Detroit 6 cells but not for DNA replication.…”

The adeno-associated virus (AAV) Rep protein acts as both a repressor and an activator to regulate AAV transcription during a productive infection

“…Several reports have demonstrated that in the absence of helper virus, AAV transcripts are not detected (12,27,28). The absence of p5 activity may be attributed, in part, to the cellular YY1 protein.…”