Relaxed template specificity in fowl adenovirus 1 DNA replication initiation

Abstract

The fowl adenovirus 1 (FAdV-1) isolates PHELPS and OTE are highly similar, but have striking differences in the repeat region of the inverted terminal repeat (ITR). Whilst the repeat region in OTE conforms to the conventional human adenovirus repeat region (59-CATCATC), that of PHELPS contains guanidine residues at positions 1, 4 and 7 (59-GATGATG). This implies that the FAdV-1 isolates PHELPS and OTE have either distinct template specificity at replication initiation or, alternatively, a relaxed specificity for replication initiation. In this study, the distinct sequence variation at the origin of DNA replication in the ITRs of the FAdV-1 PHELPS and OTE isolates was confirmed. Sequence analyses of the pTP and Pol genes of both PHELPS and OTE did not reveal differences that could explain the distinct template specificity. Replication assays demonstrated that linear DNA fragments flanked by either 59-CATCATC or 59-GATGATG termini replicated in cells upon infection with FAdV-1 OTE and FAdV-1 PHELPS. This was evident from the appearance of DpnI-resistant fragments in a minireplicon assay. From these data, it is concluded that FAdV-1 has relaxed, rather than changed, its template specificity at replication initiation.

Adenovirus DNA Replication

“…In the FAdV-A isolates Phelps and KUR, initiation starts with incorporation of a dGMP. Interestingly, the highly similar FAdV-A strain OTE is conventional and starts initiation by binding of dCMP (Rademaker et al 2006). Studies with minireplicons showed that the pTP and Pol of CELO-OTE and CELOPhelps do not have different specificities, but rather show relaxed template specificity.…”

“…Studies with minireplicons showed that the pTP and Pol of CELO-OTE and CELOPhelps do not have different specificities, but rather show relaxed template specificity. This allows these viruses to use 3 0 -GTAGTAG as well as 3 0 -CTACTAC as templates for replication initiation (Rademaker et al 2006). This implies the efficient formation of pTP-GMP complexes.…”

“…DBP is involved in several steps during DNA replication. Besides protecting ssDNA against nucleases, DBP acts during initiation by stimulating binding of AdV Pol and NFI, presumably by slight changes in the configuration of origin DNA (Hay 1966;van Breukelen et al 2003;van der Vliet and Hoeben 2006). Moreover, DBP lowers the Km for coupling of the first nucleotide to pTP.…”

“…These studies yielded many of the basic principles of mammalian DNA replication. AdV DNA replication was reviewed in detail in Hay (1966), Van der Vliet (1996), and Van der Vliet and Hoeben (2006). The mechanisms of AdV DNA replication have now been firmly established, and many of the roles of the key actors in the process have been elucidated in detail.…”

Functional characterization of highly processive protein-primed DNA polymerases from phages Nf and GA-1, endowed with a potent strand displacement capacity

“…The development of in vitro replication systems with purified proteins, mainly in the case of bacteriophage φ29 and adenovirus, has allowed the elucidation of the general bases of the protein-priming mechanism of DNA replication ( 2 – 4 ). Specific initiation proteins interact with the replication origins at both 5′ ends of the genome, partially opening the double helix, exposing a region of ssDNA.…”