DNA amplification and tumorigenicity of the human melanoma cell line MeWo.

Homogeneously staining regions (HSRs) were found in hypodiploid cells (40%) of a subline of the human melanoma cell line, MeWo, (MeWo-C) but were absent from the hypotetraploid cells (60%). Another subline (MeWo-B) was also shown to contain two populations of cells, 70% hypodiploid and 30% hypotetraploid. None of the MeWo-B cells contained HSRs, but all four cell types from both sublines shared marker chromosomes indicating their common origin. The hypodiploid MeWo-B cells were karyotypically similar to the hypodiploid MeWo-C cells except for the presence of the HSRs in the latter. Both MeWo-C and MeWo-B sublines were injected into BALB/c nude mice. The MeWo-C cells were markedly more tumorigenic than MeWo-B cells as judged by tumor incidence, latency, average tumor size, and tumor take values. Cytogenetic and flow cytofluorometric analyses of the tumors induced by MeWo-C cells revealed a shift in the tumor cell population from 40% to greater than 90% HSR-containing hypodiploid cells during tumor growth. Hybridization of tumor DNA to a probe (D15Z1), the sequence of which is amplified in the HSRs, also indicated an increase in the proportion of HSR-bearing cells during tumor growth. No such selective advantage was found with the hypodiploid, HSR-lacking MeWo-B cells. The results suggest that HSRs found in the human melanoma line MeWo may confer enhanced tumorigenicity to the cells containing them.

Organization of a repetitive human 1.8 kb KpnI sequence localized in the heterochromatin of chromosome 15.

We have isolated a repetitive 1.8 kb KpnI DNA sequence which is amplified in the homogeneously staining regions of a human melanoma cell line. Under low stringency conditions this sequence (D15Z1) hybridized in situ to the centromeric heterochromatin of chromosomes 1, 9, 15p, 16, and distal Yq as well as to the short arms of the other acrocentric chromosomes. Under conditions of high stringency, labelling was predominantly on the short arm of chromosome 15. D15Z1 was shown to be present at approximately 3,000 copies per haploid genome and organized in long tandem arrays showing restriction site heterogeneity. Sequences homologous to D15Z1 were highly enriched in the less dense shoulder region of a Ag+-Cs2SO4 gradient. Analysis of D15Z1 indicated that this sequence is composed of tandemly arranged imperfect repeats of the consensus 5' AATGG 3' similar to previously identified satellite III sequences. Digestion of D15Z1 with HinfI resulted in a series of restriction fragments making up a subset of the HinfI ladder components of satellites III and IV. These data suggest that D15Z1 represents a chromosome 15 specific domain of human satellites III or IV and that it makes up the major fraction of the heterochromatin of this chromosome. Possible relationships between this sequence and the cytochemical staining properties of human chromosomes with distamycin A/DAPI, D280/170, and antiserum to 5-methylcytosine are discussed.