Ras, Rap, and Rac small GTP-binding proteins are targets for Clostridium sordellii lethal toxin glucosylation

M R Popoff; E Chaves-Olarte; E Lemichez; C von Eichel-Streiber; M Thelestam; P Chardin; D Cussac; B Antonny; P Chavrier; G Flatau; M Giry; J de Gunzburg; P Boquet

doi:10.1074/jbc.271.17.10217

Ras, Rap, and Rac small GTP-binding proteins are targets for Clostridium sordellii lethal toxin glucosylation

J Biol Chem. 1996 Apr 26;271(17):10217-24. doi: 10.1074/jbc.271.17.10217.

Authors

M R Popoff¹, E Chaves-Olarte, E Lemichez, C von Eichel-Streiber, M Thelestam, P Chardin, D Cussac, B Antonny, P Chavrier, G Flatau, M Giry, J de Gunzburg, P Boquet

Affiliation

¹ Institut Pasteur, Unité des Toxines Microbiennes, 75724 Paris, Cedex 15, France.

PMID: 8626586
DOI: 10.1074/jbc.271.17.10217

Abstract

Lethal toxin (LT) from Clostridium sordellii is one of the high molecular mass clostridial cytotoxins. On cultured cells, it causes a rounding of cell bodies and a disruption of actin stress fibers. We demonstrate that LT is a glucosyltransferase that uses UDP-Glc as a cofactor to covalently modify 21-kDa proteins both in vitro and in vivo. LT glucosylates Ras, Rap, and Rac. In Ras, threonine at position 35 was identified as the target amino acid glucosylated by LT. Other related members of the Ras GTPase superfamily, including RhoA, Cdc42, and Rab6, were not modified by LT. Incubation of serum-starved Swiss 3T3 cells with LT prevents the epidermal growth factor-induced phosphorylation of mitogen-activated protein kinases ERK1 and ERK2, indicating that the toxin blocks Ras function in vivo. We also demonstrate that LT acts inside the cell and that the glucosylation reaction is required to observe its dramatic effect on cell morphology. LT is thus a powerful tool to inhibit Ras function in vivo.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

3T3 Cells
Actin Cytoskeleton / ultrastructure
Actins / chemistry
Amino Acid Sequence
Animals
Bacterial Toxins / metabolism*
Bacterial Toxins / toxicity
Calcium-Calmodulin-Dependent Protein Kinases / metabolism
Clostridium / pathogenicity*
Epidermal Growth Factor / pharmacology
GTP Phosphohydrolases / metabolism
GTP-Binding Proteins / metabolism*
Glucose / metabolism
Glucosyltransferases / metabolism*
Guanosine Triphosphate / metabolism
HeLa Cells / drug effects
HeLa Cells / ultrastructure
Humans
Kinetics
Mice
Molecular Sequence Data
Proto-Oncogene Proteins p21(ras) / metabolism*
Threonine / metabolism
Uridine Diphosphate Glucose / metabolism
rac GTP-Binding Proteins
rap GTP-Binding Proteins

Substances

Actins
Bacterial Toxins
lethal toxin LT, Clostridium sordellii
Threonine
Epidermal Growth Factor
Guanosine Triphosphate
Glucosyltransferases
Calcium-Calmodulin-Dependent Protein Kinases
GTP Phosphohydrolases
GTP-Binding Proteins
HRAS protein, human
Proto-Oncogene Proteins p21(ras)
rac GTP-Binding Proteins
rap GTP-Binding Proteins
Glucose
Uridine Diphosphate Glucose