Carcinogenesis, Vol. 20, No. 6, 963-968,
June 1999
© 1999 Oxford University Press
Cancer Biology |
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
Department of Cell Biology and Physiology, Room 149, Basic Medical Sciences Building, Health Sciences Center, University of New Mexico, Albuquerque, NM 87131 and
1 Department of Biology, Castetter Hall, University of New Mexico, Albuquerque, NM 78131, USA
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.
Abbreviations: SSCP, single strand conformational polymorphism; UVR, ultraviolet radiation.
2 To whom correspondence should be addressed Email: dkusewitt{at}salud.unm.edu
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