Nerve growth factor-induced differentiation does not alter the biochemical properties of a mutant prion protein expressed in PC12 cells.
Insertional and point mutations in the gene encoding the prion protein (PrP) are responsible for familial prion diseases. We have previously generated lines of Chinese hamster ovary cells that express PrP molecules carrying pathogenic mutations, and found that the mutant proteins display several biochemical properties reminiscent of PrP(Sc), the infectious isoform of PrP. To analyze the properties and effects of mutant PrP molecules expressed in cells with a neuronal phenotype, we have constructed stably transfected lines of PC12 cells that synthesize a PrP molecule carrying a nine-octapeptide insertion. We report here that this mutant PrP acquires scrapie-like properties, including detergent insolubility, protease resistance, and resistance to phospholipase cleavage of its glycolipid anchor. A detergent-insoluble and phospholipase-resistant form of the mutant protein is also released spontaneously into conditioned medium. These scrapie-like biochemical properties are quantitatively similar to those seen in Chinese hamster ovary cells and are not affected by differentiation of the PC12 cells into sympathetic neurons by nerve growth factor. Moreover, there is no detectable effect of mutant PrP expression on the morphology or viability of the cells in either the differentiated or undifferentiated state. These results indicate that conversion of mutant PrP into a PrP(Sc)-like form does not depend critically on the cellular context, and they suggest that mutant PrP expressed in cultured cells, even those having the phenotype of differentiated neurons, is not neurotoxic.
Chiesa
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R
Department of Cell Biology, Washington University School of Medicine, St. Louis, Missouri, USA.
Harris
D A
DA
D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects*; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival; D015870:Gene Expression; D017261:Glycosylphosphatidylinositols / Q000378:metabolism; D051379:Mice; D016296:Mutagenesis; D020932:Nerve Growth Factor / Q000494:pharmacology*; D016716:PC12 Cells / Q000378:metabolism*; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D011328:Prions / Q000737:chemistry / Q000235:genetics*; D051381:Rats; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism
D017261:Glycosylphosphatidylinositols; D011328:Prions; D020932:Nerve Growth Factor; D010738:Type C Phospholipases; D043265:Phosphatidylinositol Diacylglycerol-Lyase