Expression and DNA binding of budgerigar fledgling disease virus large T antigen

Abstract

Budgerigar fledgling disease virus (BFDV) represents the first non-mammalian member of the polyomavirus genus and possesses uncommon structural and biological properties. Recombinant baculoviruses were constructed to express BFDV small t antigen, large T antigens, as well as a large T deletion mutant T~ and flgalactosidase-T~ fusion proteins to high levels in infected insect cells. A recombinant virus containing a genomic copy of the BFDV early region was used for small t antigen expression, and corresponding introndeleted cDNAs for production of large T antigen derivatives. Recombinant T as well as authentic T antigen proteins from infected chicken embryo fibroblasts were purified using both immunoaffinity and DNA affinity column chromatography. We present evidence that the large T antigen interacts specifically with DNA sequences present in the non-coding region of BFDV; by indirect DNA immunoprecipitation mapping and DNase I footprinting, four regions including 12 DNA-binding sites have been determined that cover most of the BFDV non-coding region. The T antigen binding pattern observed suggests a protein-DNA interaction system considerably different from those of simian virus 40 and other polyomaviruses.

Early and late pre-mRNA processing of budgerigar fledgling disease virus 1: identification of viral RNA 5' and 3' ends and internal splice junctions

“…Even though the various species of early mRNAs have not been tested individually for their translational capacities, they are very likely to represent the small t and large T antigen messengers, respectively. This is supported by the results obtained in the baculovirus system using unspliced and spliced eDNA constructs, which code for small t and large T protein (Luo et al, 1994).…”

Characterization of Two Novel Polyomaviruses of Birds by Using Multiply Primed Rolling-Circle Amplification of Their Genomes

“…Also, the penta- nucleotide sequence GAGGC is not consistently found in the noncoding region of the avian polyomaviruses, but the palindromic sequence NCC(A/T) 6 GGN is found at least once in this region in all avian polyomaviruses. As this sequence has been shown to be specifically bound by the large T-Ag of APV (25), a similar manner of large T-Ag DNA-binding may be common in all avian polyomaviruses. Because of the biological and structural differences between polyomaviruses of mammals and birds, a grouping into two different genera within the family Polyomaviridae should be considered.…”

A Novel Virus Detected in Papillomas and Carcinomas of the Endangered Western Barred Bandicoot ( Perameles bougainville ) Exhibits Genomic Features of both the Papillomaviridae and Polyomaviridae

“…In mammalian PyVs, the large T antigen binds to this GAGGC pentanucleotide (53). In avian PyVs, however, the binding sequence for the large T antigen has been identified as the palindromic sequence TCC(A/T) 6 GGA or similar (25,29), for which related sequences were not identified in the BPCV1 sequence. Two potential GAGGC large T antigen binding sites were identified in the putative BPCV1 viral regulatory region.…”