Expression and function of the CD44 glycoprotein in melanoma cell lines

We have analysed a panel of murine and human melanoma cell lines for expression of the glycoprotein CD44. All 12 cell lines examined expressed CD44 at their cell surfaces, as demonstrated by fluorocytometric analysis using monoclonal antibodies (MAbs) IM7 and F10.44.2. Northern analysis revealed three transcript sizes that were 4.5, 2.2, and 1.5 kb in the human cell lines and 4.5, 3.0, and 1.5 kb in the murine cell lines. Levels of mRNA did not correlate with level of surface expression, which was highly variable between the cell lines. RT-PCR analysis of mRNA revealed that the major band identified was the expected 792 bp fragment indicative of the CD44H haemopoietic form, compared to a 1194 bp form found in the human colorectal adenocarcinoma cell line HT29, indicative of the CD44E epithelial form. There was no evidence of variant CD44 mRNA in our panel of melanomas. Functional assays revealed no clear correlation between the level of cell surface CD44 and the ability of the melanoma cell lines to adhere to hyaluronate. Rather adherence appeared to relate to the activation status of CD44 on the different cell lines as a consequence of MAb stimulation (e.g. the 1735P line demonstrated a 46.2 +/- 5.7% adherence in an inactivated state versus 62.4 +/- 5.6% adherence in an activated state to 5 mg/ml hyaluronate) and suggests that the functional capacity of CD44 expressed by melanoma cells may be modified more by activation state than by RNA splicing.