Recombinant DNA techniques have been employed to isolate sequences that are over-expressed during multi-stage carcinogenesis in the mouse skin. One of these six cDNA sequences, mal-4, detected 1.9-KB transcripts that were expressed at levels 10-fold higher in squamous cell carcinomas (SCCs) in comparison to normal epidermis. A full-length cDNA for mal-4 was obtained from a λgt10 cDNA library made from an SCC-producing cell line, PDVC57. DNA sequencing of 1300 out of 1900 bp of the mal-4 cDNA and searching of Genbank showed >95% DNA sequence similarity to mouse β-actin over the entire cDNA and >98% similarity over the 3' untranslated region, which is unique to the various isoforms of actins. Southern analysis showed no amplification or rearrangement of the β-actin gene had occurred during tumor progression. The RNA:RNA hybrid protection assay was utilized to screen for the expression of mutated β-actin(s) in mouse skin tumors. No evidence for mutation was obtained in any of the tumors examined. Fluorescence microscopy of tumor sections stained with rhodamine-conjugated phalloidin showed a peripheral pattern of F-actin localization with no gross differences between papillomas and carcinomas. Approximately equal amounts of β-actin were observed following two-dimensional gel electrophoresis of proteins extracted from normal epidermis, papillomas or SCCs, indicating that the over-expression of β-actin RNA in SCCs did not result in an increased steadystate level of β-actin protein. These results suggest that alterations in actin metabolism accompany tumor progression.
ASJC Scopus subject areas
- Cancer Research