Journal Article

The <i>p53</i> tumor suppressor gene of the marsupial <i>Monodelphis domestica</i>: cloning of exons 4–11 and mutations in exons 5–8 in ultraviolet radiation-induced corneal sarcomas

Donna F. Kusewitt, Tamara E. Sherburn, Katarzyna B. Miska, Gregory B. Tafoya, James M. Gale and Robert D. Miller

in Carcinogenesis

Volume 20, issue 6, pages 963-968
Published in print June 1999 | ISSN: 0143-3334
Published online June 1999 | e-ISSN: 1460-2180 | DOI: http://dx.doi.org/10.1093/carcin/20.6.963
The p53 tumor suppressor gene of the marsupial Monodelphis domestica: cloning of exons 4–11 and mutations in exons 5–8 in ultraviolet radiation-induced corneal sarcomas

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Inactivating p53 mutations are found in many ultraviolet radiation (UVR)-induced skin tumors. We examined 12 UVR-induced corneal tumors of the marsupial Monodelphis domestica for mutations in exons 5–8 of p53 and compared their mutational spectrum with that of UVR-induced skin tumors of other species. First we cloned and characterized a cDNA extending from the middle of exon 4 through exon 11 of the Monodelphis p53 gene. Based on the sequence information obtained, primers were designed to amplify introns 4–9 of the gene; intron primers to amplify individually exons 5–8 were subsequently developed. `Cold' single strand conformational polymorphism analysis followed by reamplification of DNA with altered mobility and cycle sequencing revealed single p53 mutations in four of 12 tumors (33%), including one mutation in exon 5, two identical mutations in exon 7 and one mutation in exon 8. All mutations were at dipyrimidine sites and occurred on the non-transcribed strand. Three of the four were hallmark UVR-induced C→T alterations. Three of the mutations were found at sites corresponding to human codons 248 and 273, which are mutational hotspots in human and murine UVR-induced squamous cell carcinomas. Our findings suggest that UVR-induced corneal sarcomas in Monodelphis will be valuable in studying mechanisms of p53 mutation in UVR-induced tumors.

Keywords: SSCP, single strand conformational polymorphism; UVR, ultraviolet radiation.

Journal Article.  4746 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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