Presence of a laminin-binding chondroitin sulfate proteoglycan at the cell surface of a human melanoma cell Mel-85.
Working with Mel-85 (a human melanoma cell line), we have been able to detect a laminin-binding molecule with an apparent molecular mass of 100/110 kDa (Mel-85-LBM). Reduction with beta-mercaptoethanol decreases its molecular mass but does not affect its ability to bind laminin. This laminin interaction seems to be very specific since Mel-85-LBM binds laminin, but not fibronectin, vitronectin or type I collagen in affinity chromatography experiments. The molecule has a negative net charge at physiological pH and binds laminin in a divalent cation dependent way. Mel-85-LBM was metabolically radiolabeled with sodium [35S]-sulfate and chemical beta-elimination of purified Mel-85-LBM releases chondroitin sulfate chains. Mel-85-LBM is also sensitive to chondroitinase ABC digestion. These findings show that this molecule is a chondroitin sulfate proteoglycan. The location of this proteoglycan at the cell surface is evidenced by experiments using a polyclonal antiserum raised against purified Mel-85-LBM, that specifically reacts with just one molecule by western blotting among Mel-85 total cell extract as well as produces a positive signal by flow cytometry and a fluorescence profile of Mel-85 cells adhered on laminin.
Elias
M C
MC
Ludwig Institute for Cancer Research, São Paulo, Brazil.
Veiga
S S
SS
Gremski
W
W
Porcionatto
M A
MA
Nader
H B
HB
Brentani
R R
RR
D002448:Cell Adhesion; D002462:Cell Membrane / Q000378:metabolism; D002465:Cell Movement; D011508:Chondroitin Sulfate Proteoglycans / Q000302:isolation & purification / Q000378:metabolism*; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007797:Laminin / Q000378:metabolism*; D008545:Melanoma / Q000378:metabolism* / Q000473:pathology; D011485:Protein Binding; D014407:Tumor Cells, Cultured
D011508:Chondroitin Sulfate Proteoglycans; D007797:Laminin