Date of Award
Open Access Thesis
Medical Doctor (MD)
We embarked on a study to define the role of protein-tyrosine phosphatases (PTPases) in both normal melanocytes and melanoma cells, searching for specific differences that might provide clues to the malignant transformation process in these cells and to the role of PTPases as possible tumor suppressor genes. We used Northern blotting and RNase protection assays to measure mRNA expression levels, immunoprecipitation and immunoblotting to measure protein expression, and a bioassay to measure protein PTPase activity in normal melanocyte and melanoma cell lines.
Our experiments showed that all major PTPases are equally expressed at both mRNA and protein levels, except for HPTPase O, which is expressed in two out of four melanoma cell lines tested, but not in normal melanocytes. In addition, basal levels of PTPase activity were detected in cell extracts of both normal and transformed melanocytes, and the major cytosolic PTPase, PTPlB, was identified at the protein level. PTPlB expression in both normal and malignant melanocytes was not altered by the addition of mast cell growth factor (MGF), except in the melanoma cell line 888, in which PTPlB was expressed after, but not before, MGF stimulation.
In undertaking studies of PTPase activity, we found that in a specific melanoma cell line, YU-ZAZ, stimulation by vascular endothelial growth factor increased overall PTPase activity compared to basal levels. Furthermore, specificity of PTPase activity in these whole cell extracts was demonstrated to be equal towards both reduced carboxamidomethylated and maleylated lysozyme (RCML) and myelin basic protein (MBP) phosphorylated substrates. These studies are the first to characterize the role of PTPases in normal melanocytes and melanoma cells, and may help to clarify the mode of transformation in these cells and elucidate the physiological role of PTPases in regulation of cell growth.
Ahn, James H., "Protein-Tyrosine Phosphatase in Normal and Malignant Melanocytes" (1995). Yale Medicine Thesis Digital Library. 3563.
This Article is Open Access