Isolation of canine p53 cDNA and detailed characterization of the full length canine p53 protein

Abstract

The p53 tumour suppressor protein plays a central role in the maintenance of genomic integrity. Mutations of the p53 gene are found in a number of canine cancers and many contribute to tumour formation. Here we describe isolation and expression of the complete wild type canine p53 cDNA. The encoded full length canine p53 protein displays strong sequence homology with p53 proteins from other higher vertebrates. Canine p53 protein produced in vitro was shown to recognize and bind to p53-speci®c DNA targets derived from the p21 and GADD45 promoters and to a consensus p53 binding site. We also show that canine p53 associates with oligonucleotides representing damaged DNA sites and undergoes proteolytic cleavage similar to that described for murine and human p53 proteins. Finally, we show that the canine p53 protein is able to transcriptionally activate a p53-dependent reporter gene in vivo. The results suggest that canine p53 is similar both in structure and function to human p53 and that canine cancer may provide a useful clinical model in the search for e ective anti-cancer therapies based on p53.

Detection of mutations within exons 4 to 8 of the p53 tumor suppressor gene in canine mammary glands

“…However, only 626 nucleotides have identity between the two sequences. This difference in the sequence of these species could be due to deletions in the proline-rich regions in the canine Tp53, which developed through evolution (Kraegel et al, 1995;Veldhoen and Milner, 1998). The conserved region of the nucleotide sequences allow comparative studies on human breast cancer using the canine model.…”

“…(Chu et al, 1998). Alterations in the Tp53 gene have been identified in some canine cancer types, including colorectal tumor, thyroid carcinoma (Devilee et al 1994), oral papilloma (Mayr et al, 1994), mammary tumor (Van Leeuwen et al, 1996;Chu et al, 1998;Klopfleisch andGruber, 2009), osteosarcoma (Van Leeuwen et al, 1997), circumanal gland adenoma (Mayr et al, 1997) and lymphoma (Veldhoen and Milner, 1998).…”

A novel exon within the mdm2 gene modulates translation initiation in vitro and disrupts the p53-binding domain of mdm2 protein

“…Having cloned and sequenced canine p53 (Veldhoen and Milner, 1998) we next sought to characterize canine mdm2 and its interaction with p53. A canine homologue of mdm2 cDNA was isolated by reverse transcriptase-PCR (RT ± PCR) using mRNA derived from canine mammary tumour tissue (Materials and methods).…”

Nucleotide sequence of the porcine p53 cDNA, and the detection of recombinant porcine p53 expressed in vitro with a variety of anti-p53 antibodies

“…The complete cDNA sequences of human (ZakutHouri et al, 1985), murine (Zakut-Houri et al, 1983), feline (Okuda et al, 1994), canine (Veldhoen and Milner, 1998) and bovine (Komori et al, 1996) p53 have been published. Partial sequences for equine (Pazzi et al, 1996) and asinine (Nasir and Reid, 1995) p53 have also been described.…”

“…Putative binding sites of two cross-species reactive monoclonal antibodies are highlighted in grey. The position of ®ve domains highly conserved during evolution is noted by underlining of the consensus sequence and the appropriate Roman numeral (I ± V) available anti-p53 antibodies are raised against human or mouse p53 and in previous work have been shown to be poorly reactive with recombinant p53s of other species (Veldhoen and Milner, 1998). The CM-1 polyclonal antibody detected both recombinant p53 produced by transcription/translation of the porcine p53/Pci-Neo construct and the positive control p53 (Figure 3).…”