Losses of heterozygosity involving chromosomes 9 and 10 are frequent events in the development and progression of cutaneous malignant melanoma. To investigate whether specifically deleted chromosomal regions encode tumor suppressor genes (TSGs), we introduced normal chromosome 10 into the tumorigenic human metastatic melanoma cell line UACC-903 by microcell fusion. In addition, two chromosome 9 derivatives that were microdeleted in the region of the p16INK4A/p15INK4B locus were transferred to determine whether an additional melanoma TSG or TSGs reside on chromosome 9p, as indicated by previous melanoma allele loss studies. In comparison to parental cells, microcell hybrids generated with chromosomes 9 (microdeleted) and 10 displayed reduced anchorage-independent growth in soft agar and markedly reduced tumorigenicity in athymic (nu/nu) mice. These data define a TSG or TSGs that function independently of p15/p16 on chromosome 9 and provide evidence for a TSG (or TSGs) on chromosome 10 that may be important in melanoma development.