IDENTIFICATION OF GENES DIFFERENTIALLY EXPRESSED IN ASSOCIATION WITH METASTATIC POTENTIAL OF K-1735 MURINE MELANOMA BY MESSENGER-RNA DIFFERENTIAL DISPLAY

To identify genes differentially expressed in association with the met astatic potential of K-1735 mouse melanoma cells, the mRNA differentia l display method was applied to compare mRNAs from high- and low-metas tatic K-1735-derived cells. Three of the high- and three of the low-me tastatic clones were used to reduce the false positives in the initial screening, and Southern blot screening against reverse transcription- PCR products was used to confirm that cDNA fragments detect differenti al expression between high- and low-metastatic cells. By using 256 dif ferent combinations of modified long arbitrary primers which provide b road screening of expressed genes, approximately 12,000 cDNA fragments were amplified from mRNA of each cell line. Among them, eight genes w ere identified as being expressed in either high- or low-metastatic ce lls using Northern blot analysis. Integrin alpha 6 and two unknown gen es were expressed in high-metastatic cells, whereas beta-tropomyosin, macrophage colony-stimulating factor, inhibin/activin beta B subunit, and two unknown genes were expressed in low-metastatic cells. These re sults indicate that the acquisition of metastatic potential in tumor c ells was regulated by activation and/or inactivation of several specif ic genes, such as those for cell adhesion molecule, cytoskeletal prote in, and growth factors.