Subtraction hybridization identifies a novel melanoma differentiation associated gene, mda-7, modulated during human melanoma differentiation, growth and progression.
Cultured human melanoma cells lose proliferative capacity and terminally differentiate after treatment with the combination of recombinant human fibroblast interferon (IFN-beta) and mezerein (MEZ). Subtraction hybridization of cDNA libraries prepared from actively proliferating human H0-1 melanoma cells from cDNA libraries produced from H0-1 cells treated with IFN-beta + MEZ identifies a novel melanoma differentiation-associated (mda) cDNA, mda-7, that displays elevated expression in differentiation inducer-treated H0-1 cells. mda-7 encodes a novel protein of 206 amino acids with a predicted size of 23.8 kDa. The level of mda-7 mRNA is elevated in actively proliferating normal human melanocytes versus primary and metastatic human melanomas. In the Matrigel-assisted melanoma progression model, mda-7 expression decreases in early vertical growth phase primary human melanoma cells selected for autonomous or enhanced tumor formation in nude mice. Treatment of human melanomas with IFN-beta + MEZ, and to a lesser extent with MEZ, results in growth suppression and induced or enhanced mda-7 expression. Immunoprecipitation analyses using peptide-derived rabbit polyclonal antibodies detect increases in mda-7 protein, and a higher molecular weight protein of approximately 90 to 100 kDa, in MEZ and IFN-beta + MEZ treated H0-1 cells. mda-7 is a highly conserved gene with an homologous sequence in the genome of yeast. Transfection of mda-7 expression constructs into H0-1 and C8161 human melanoma cells reduces growth and inhibits colony formation. These results confirm that mda-7 has antiproliferative properties in human melanoma cells and in this context may contribute to terminal cell differentiation. The mda-7 gene may also function as a negative regulator of melanoma progression.
Jiang
H
H
Department of Pathology, Columbia-Presbyterian Cancer Center, Columbia University, College of Physicians and Surgeons, New York, NY 10032, USA.
Lin
J J
JJ
Su
Z Z
ZZ
Goldstein
N I
NI
Fisher
P B
PB
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002454:Cell Differentiation; D002455:Cell Division; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000302:isolation & purification*; D005260:Female; D015870:Gene Expression; D016147:Genes, Tumor Suppressor; D006133:Growth Substances / Q000235:genetics*; D006801:Humans; D007378:Interleukins; D008545:Melanoma / Q000235:genetics* / Q000473:pathology; D051379:Mice; D008875:Middle Aged; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000235:genetics*; D009693:Nucleic Acid Hybridization; D011817:Rabbits; D014407:Tumor Cells, Cultured; D000099180:Interleukin-24
D018076:DNA, Complementary; D006133:Growth Substances; D007378:Interleukins; D009363:Neoplasm Proteins; D000099180:Interleukin-24