GPI-specific phospholipase D mRNA expression in tumor cells of different malignancy.
Glycosylphosphatidylinositol-specific phospholipase D (GPI-PLD) is a highly specific enzyme whose only known substrate is the GPI anchor of cell surface proteins. GPI-PLD measurements, however, are technically difficult since the enzyme is expressed at low levels in cells and tissues, and serum contains large amounts of inactive, latent GPI-PLD interfering with protein-based assays. We have therefore developed a semi-quantitative RT-PCR method to measure mRNA expression of all known GPI-PLD isoforms in cells and tissues. In human ovarian cancer cell lines, GPI-PLD mRNA expression correlated with GPI-PLD enzyme activity and with the shedding of the GPI-anchored tumor and prognostic markers, urokinase receptor and CA125, from the cell surface. This supports a potential role for this enzyme in the generation of circulating prognostic markers in malignant tumors. Similarly, in human epithelial cells of the skin, GPI-PLD mRNA expression increased with tumor progression. Whereas normal keratinocytes did not express significant amounts of GPI-PLD mRNA, expression was dramatically induced by serum in immortalized HaCaT keratinocytes and constitutively high and independent of serum in tumorigenic A431 epidermoid carcinoma cells. In addition, GPI-PLD expression was significantly increased in highly malignant. H-ras-transfected murine bladder carcinoma cells as compared to the low malignant, non-transfected parental cells. The competitive RT-PCR described here represents the first quantitative assay specific for cellular GPI-PLD isoforms, and our in vitro analyses suggest that GPI-PLD expression might be associated with tumor malignancy.
Xiaotong
H
H
Department of Cancer Research, Fachklinik Hornheide, University of Münster, Germany.
Hannocks
Melanie-Jane
MJ
Hampson
Ian
I
Brunner
Georg
G
D000818:Animals; D014408:Biomarkers, Tumor; D002277:Carcinoma / Q000201:enzymology / Q000473:pathology; D002295:Carcinoma, Transitional Cell / Q000201:enzymology / Q000473:pathology; D018076:DNA, Complementary / Q000235:genetics; D004273:DNA, Neoplasm / Q000235:genetics; D004790:Enzyme Induction; D004847:Epithelial Cells / Q000201:enzymology; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D011905:Genes, ras; D006801:Humans; D051379:Mice; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000502:physiology; D009369:Neoplasms / Q000201:enzymology* / Q000473:pathology; D010051:Ovarian Neoplasms / Q000201:enzymology / Q000473:pathology; D010739:Phospholipase D / Q000096:biosynthesis / Q000235:genetics* / Q000502:physiology; D011379:Prognosis; D012333:RNA, Messenger / Q000096:biosynthesis*; D012334:RNA, Neoplasm / Q000096:biosynthesis*; D012867:Skin / Q000166:cytology; D001749:Urinary Bladder Neoplasms / Q000201:enzymology / Q000473:pathology
D014408:Biomarkers, Tumor; D018076:DNA, Complementary; D004273:DNA, Neoplasm; D009363:Neoplasm Proteins; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D010739:Phospholipase D; C065017:glycoprotein phospholipase D