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 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 10(5) 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.
Flores-Díaz
M
M
Microbiology and Tumorbiology Center, Karolinska Institutet, S-171 77 Stockholm, Sweden.
Alape-Girón
A
A
Titball
R W
RW
Moos
M
M
Guillouard
I
I
Cole
S
S
Howells
A M
AM
von Eichel-Streiber
C
C
Florin
I
I
Thelestam
M
M
D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D001427:Bacterial Toxins / Q000096:biosynthesis / Q000737:chemistry / Q000633:toxicity*; D002135:Calcium-Binding Proteins; D002417:Cattle; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D002794:Choline / Q000378:metabolism; D003013:Clostridium / Q000201:enzymology; D003016:Clostridium perfringens / Q000201:enzymology*; D006224:Cricetinae; D003412:Cricetulus; D007700:Kinetics; D008168:Lung; D008563:Membrane Lipids / Q000096:biosynthesis; D010446:Peptide Fragments / Q000494:pharmacology; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism / Q000633:toxicity; D014162:Transfection; D010738:Type C Phospholipases / Q000096:biosynthesis / Q000737:chemistry / Q000633:toxicity*; D005957:UTP-Glucose-1-Phosphate Uridylyltransferase / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism; D014532:Uridine Diphosphate Glucose / Q000172:deficiency / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D001427:Bacterial Toxins; D002135:Calcium-Binding Proteins; D008563:Membrane Lipids; D010446:Peptide Fragments; D011994:Recombinant Proteins; D005957:UTP-Glucose-1-Phosphate Uridylyltransferase; D010738:Type C Phospholipases; C040649:alpha toxin, Clostridium perfringens; D002794:Choline; D014532:Uridine Diphosphate Glucose