Suppression subtractive hybridisation to isolate differentially expressed genes involved in invasiveness of melanoma cell line cultured under different conditions.
We have previously analysed the invasion capacity of different melanoma cell lines in the three-dimensional dermal equivalent. The melanoma cell line M4Beu acquired invasive behaviour upon changing its cultivation conditions before the seeding on top of the collagen lattice from single cell suspension to spheroid. Based on this phenomenon SSH was used to search for the genes related to the invasive phenotype of melanoma cells. From differentially expressed clones we focused on four: fibronectin, RhoA, COXII, and H-ras-like protein. By RT-PCR the expression of these genes were tested in different populations (monolayer, spheroids on dermal equivalent) of melanoma cell line M4Beu and three additional melanoma cell lines. The expression of fibronectin was also examined by immunohistochemistry staining of co-culture spheroids-dermal equivalent.
Eleveld-Trancíková
Dagmar
D
Department of Tumor Cell Biology, Cancer Research Institute, Slovack Academy of Sciences, 833 91 Bratislava, Slovakia. d.eleveld@ncmls.kun.nl
Kúdela
Pavol
P
Majerciak
Vladimir
V
Regendová
Mária
M
Zelník
Vladimír
V
Pastorek
Jaromír
J
Pastoreková
Silvia
S
Bízik
Jozef
J
D003001:Cloning, Molecular; D051546:Cyclooxygenase 2; D018076:DNA, Complementary / Q000378:metabolism; D005353:Fibronectins / Q000096:biosynthesis / Q000235:genetics; D005821:Genetic Techniques; D006801:Humans; D007150:Immunohistochemistry; D007527:Isoenzymes / Q000096:biosynthesis / Q000235:genetics; D008545:Melanoma / Q000235:genetics*; D008565:Membrane Proteins; D009361:Neoplasm Invasiveness; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis / Q000235:genetics; D016283:Proto-Oncogene Proteins p21(ras) / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D014407:Tumor Cells, Cultured; D020742:rhoA GTP-Binding Protein / Q000096:biosynthesis / Q000235:genetics
D018076:DNA, Complementary; D005353:Fibronectins; D007527:Isoenzymes; D008565:Membrane Proteins; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases; C501009:HRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras); D020742:rhoA GTP-Binding Protein