UDP-GLUCOSE DEFICIENCY CAUSES HYPERSENSITIVITY TO THE CYTOTOXIC EFFECT OF CLOSTRIDIUM-PERFRINGENS PHOSPHOLIPASE-C

A Chinese hamster cell line with a mutation in the UDP-glucose pyropho sphorylase (UDPG:PP) gene leading to UDP-glucose deficiency as well as a revertant cell were previously isolated. We now show that the mutan t cell is 10(5) times more sensitive to the cytotoxic effect of Clostr idium perfringens phospholipase C (PLC) than the revertant cell. To cl arify whether there is a connection between the UDP-glucose deficiency and the hypersensitivity to C. perfringens PLC, stable transfectant c ells were prepared using a wild type UDPG:PP cDNA. Clones of the mutan t transfected with a construct having the insert in the sense orientat ion had increased their UDP-glucose level, whereas those of the revert ant transfected with a UDPG:PP antisense had reduced their level of UD P-glucose compared with control clones transfected with the vector. Ex posure of these two types of transfectant clones to C. perfringens PLC demonstrated that a cellular UDP-glucose deficiency causes hypersensi tivity to the cytotoxic effect of this phospholipase. Further experime nts with genetically engineered C. perfringens PLC variants showed tha t the sphingomyelinase activity and the C-domain are required for its cytotoxic effect in UDP-glucose-deficient cells.