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J Biol Chem, Vol. 273, Issue 38, 24433-24438, September 18, 1998

UDP-glucose Deficiency Causes Hypersensitivity to the Cytotoxic Effect of Clostridium perfringens Phospholipase C

Marietta Flores-Díaz^§, Alberto Alape-Girón^§¶, Richard W. Titball^**, Michael Moos, Isabelle Guillouard^¶¶, Stewart Cole^¶¶, Angela M. Howells^**, Christoph von Eichel-Streiber, Inger Florin, and Monica Thelestam

From the  Microbiology and Tumorbiology Center, Karolinska Institutet, S-171 77 Stockholm, Sweden, ^§ Instituto Clodomiro Picado, Facultad de Microbiología and ^¶ Departamento de Bioquímica, Facultad de Medicina, Universidad de Costa Rica, San José, Costa Rica, ^** Defense Evaluation and Research Agency, Chemical and Biological Defense Establishment Porton Down, Salisbury, Wiltshire Sp4 OJQ, United Kingdom,  Institut für Medizinische Mikrobiologie und Hygiene, Verfügungsgebäude 55101 Mainz, Germany, and ^¶¶ Unité de Génetique Moléculaire Bactérienne, Institut Pasteur, 75724 Paris Cédex 15, France

A Chinese hamster cell line with a mutation in the UDP-glucose pyrophosphorylase (UDPG:PP) gene leading to UDP-glucose deficiency as well as a revertant cell were previously isolated. We now show that the mutant cell is 10⁵ times more sensitive to the cytotoxic effect of Clostridium perfringens phospholipase C (PLC) than the revertant cell. To clarify whether there is a connection between the UDP-glucose deficiency and the hypersensitivity to C. perfringens PLC, stable transfectant cells were prepared using a wild type UDPG:PP cDNA. Clones of the mutant transfected with a construct having the insert in the sense orientation had increased their UDP-glucose level, whereas those of the revertant transfected with a UDPG:PP antisense had reduced their level of UDP-glucose compared with control clones transfected with the vector. Exposure of these two types of transfectant clones to C. perfringens PLC demonstrated that a cellular UDP-glucose deficiency causes hypersensitivity to the cytotoxic effect of this phospholipase. Further experiments with genetically engineered C. perfringens PLC variants showed that the sphingomyelinase activity and the C-domain are required for its cytotoxic effect in UDP-glucose-deficient cells.

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