A rapid selection for animal cell mutants with defective peroxisomes.
Chinese hamster ovary (CHO) cells take up and incorporate 9-(1'-pyrene)nonanol (P9OH) into phospholipids and neutral lipids. Exposure of P9OH-labeled cells to long wavelength ultraviolet (UV) light causes cell death, because excitation of the pyrene moiety generates reactive oxygen species. CHO mutant cells deficient in plasmalogen biosynthesis and peroxisome assembly (Zoeller, R.A. and Raetz, C.R.H. (1986) Proc. Natl. Acad. Sci. USA 83, 5170-5174) are much more resistant to P9OH/UV treatment than are wild-type cells. This phenotype is explained by a 7.5-fold reduction of P9OH incorporation into the ethanolamine-linked phospholipids in the mutant cells and 2.4- to 6-fold reduction of P9OH incorporation into all other phospholipids and triglycerides, suggesting a general defect in fatty alcohol metabolism. [U-14C]Hexadecanol incorporation into the phospholipids of the mutant cells is also impaired. In contrast, the fatty acid analog, 9-(1'-pyrene)nonanoic acid, is incorporated into cells two times more rapidly by the mutants than by the wild type. Resistance to P9OH/UV treatment affords a simple, new method for the selection of animal cell mutants defective in peroxisome biogenesis.
Morand
O H
OH
Department of Biochemistry, College of Agricultural and Life Sciences, University of Wisconsin, Madison.
Allen
L A
LA
Zoeller
R A
RA
Raetz
C R
CR
D000818:Animals; D002374:Catalase / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival / Q000528:radiation effects; D002784:Cholesterol / Q000378:metabolism; D006224:Cricetinae; D003600:Cytosol / Q000201:enzymology; D005233:Fatty Alcohols / Q000378:metabolism; D005260:Female; D005434:Flow Cytometry; D005609:Free Radicals; D008830:Microbodies / Q000502:physiology* / Q000648:ultrastructure; D009154:Mutation; D010053:Ovary; D010100:Oxygen / Q000378:metabolism; D010743:Phospholipids / Q000378:metabolism; D010777:Photochemistry; D010955:Plasmalogens / Q000096:biosynthesis*; D011721:Pyrenes / Q000378:metabolism; D014280:Triglycerides / Q000378:metabolism; D014466:Ultraviolet Rays
D005233:Fatty Alcohols; D005609:Free Radicals; D010743:Phospholipids; D010955:Plasmalogens; D011721:Pyrenes; D014280:Triglycerides; C064596:9-(1'-pyrene)nonanol; C005031:cetyl alcohol; D002784:Cholesterol; D002374:Catalase; D010100:Oxygen