Restoration of CDKN2A into melanoma cells induces morphologic changes and reduction in growth rate but not anchorage-independent growth reversal.
CDKN2A is a melanoma susceptibility gene that is mutated and/or deleted in familial and sporadic melanoma as well as in a range of other tumors. It encodes a cell cycle regulator, p16, whose function is to inhibit activity of cyclin-dependent kinases 4 and 6. We set out to investigate the effect of reintroducing CDKN2A into MM96L, a melanoma cell line that does not express p16, by electroporation of wt CDKN2A cDNA. Our results show that transfection of the CDKN2A cDNA has a dramatic effect on cell morphology, inducing a more dendritic phenotype resembling that of adult melanocytes. This effect on cell morphology was not cell line specific because it was reproduced in another melanoma line (SK-MEL-13), which has a homozygous deletion of CDKN2A. It was abolished by mutations that abrogate p16 function, as shown by transfection of a Pro81Leu p16 variant. Reintroduction of levels of p16 protein similar to those of cultured neonatal foreskin melanocytes decreased the growth rate of the transfected clones. Surprisingly, we did not see any effect on anchorage-independent growth or on the following melanoma markers tested by western blotting: p21/WAF1, tyrosinase-related antigen 1, HMB45, and intermediate filament antigen. These data indicate that reintroduction into melanoma cells of wild type p16 at levels similar to cultured melanocytes can induce morphologic changes and suppress growth but is not sufficient to affect anchorage-independent growth.
Castellano
M
M
Joint Experimental Oncology Program, Queensland Institute of Medical Research, Post Office Royal Brisbane Hospital, Herston, Australia.
Gabrielli
B G
BG
Hussussian
C J
CJ
Dracopoli
N C
NC
Hayward
N K
NK
D000368:Aged; D014408:Biomarkers, Tumor / Q000032:analysis; D002352:Carrier Proteins / Q000235:genetics*; D002999:Clone Cells / Q000378:metabolism; D019941:Cyclin-Dependent Kinase Inhibitor p16; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D015870:Gene Expression; D016147:Genes, Tumor Suppressor; D014644:Genetic Variation; D006801:Humans; D008545:Melanoma / Q000235:genetics / Q000473:pathology; D047428:Protein Kinase Inhibitors; D014162:Transfection; D014407:Tumor Cells, Cultured / Q000187:drug effects
D014408:Biomarkers, Tumor; D002352:Carrier Proteins; D019941:Cyclin-Dependent Kinase Inhibitor p16; D004791:Enzyme Inhibitors; D047428:Protein Kinase Inhibitors