Detection of a human DNA sequence correlated with melanocyte-like differentiation and tumour suppression after transfection into murine melanoma cells.

Since there is much indirect evidence for dominant suppressor genes for melanoma, we sought to isolate such a gene. Metastatic B16-F10 murine melanoma cells were lipofected with a normal human genomic library in a cosmid vector that also confers resistance to the drug G418. A few of the G418-resistant colonies acquired combinations of properties resembling those of normal melanocytes, including differentiated features (pigmentation, dendricity), slower growth, flat shape, monolayered colony form, stimulation of proliferation by a phorbol ester, and anchorage dependence. Four out of eight also showed reduced tumorigenicity in mice. Southern blotting indicated the presence of numerous cosmids in the melanocyte-like transfectants. DNA from one such line was used for secondary transfection. One secondary G418-resistant line showed pronounced melanocytic properties and marked tumour suppression in syngeneic and nude mice. A human repetitive sequence detected in this line was used in the polymerase chain reaction (PCR) to isolate intervening unique DNA sequences. One unique human sequence was attenuated in all tumors arising from both primary and secondary transfectants, suggesting close linkage with the sequence responsible for tumour suppression.