A novel cytotoxin from Clostridium difficile serogroup F is a functional hybrid between two other large clostridial cytotoxins.
The large clostridial cytotoxins (LCTs) constitute a group of high molecular weight clostridial cytotoxins that inactivate cellular small GTP-binding proteins. We demonstrate that a novel LCT (TcdB-1470) from Clostridium difficile strain 1470 is a functional hybrid between "reference" TcdB-10463 and Clostridium sordellii TcsL-1522. It bound to the same specific receptor as TcdB-10463 but glucosylated the same GTP-binding proteins as TcsL-1522. All three toxins had equal enzymatic potencies but were equally cytotoxic only when microinjected. When applied extracellularly TcdB-1470 and TcdB-10463 were considerably more potent cytotoxins than TcsL-1522. The small GTP-binding protein R-Ras was identified as a target for TcdB-1470 and also for TcsL-1522 but not for TcdB-10463. R-Ras is known to control integrin-extracellular matrix interactions from inside the cell. Its glucosylation may be a major determinant for the cell rounding and detachment induced by the two R-Ras-attacking toxins. In contrast, fibroblasts treated with TcdB-10463 were arborized and remained attached, with phosphotyrosine containing structures located at the cell-to-cell contacts and beta3-integrin remaining at the tips of cellular protrusions. These components were absent from cells treated with the R-Ras-inactivating toxins. The novel hybrid toxin will broaden the utility of the LCTs for clarifying the functions of several small GTPases, now including also R-Ras.
Chaves-Olarte
E
E
Microbiology and Tumorbiology Center, Karolinska Institutet, S-171 77 Stockholm, Sweden.
Löw
P
P
Freer
E
E
Norlin
T
T
Weidmann
M
M
von Eichel-Streiber
C
C
Thelestam
M
M
D016475:3T3 Cells; D000818:Animals; D001427:Bacterial Toxins / Q000378:metabolism* / Q000494:pharmacology; D001483:Base Sequence; D002448:Cell Adhesion / Q000187:drug effects; D016360:Clostridioides difficile / Q000378:metabolism*; D003603:Cytotoxins / Q000378:metabolism* / Q000494:pharmacology; D017931:DNA Primers; D006031:Glycosylation; D051379:Mice; D008855:Microscopy, Electron, Scanning; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D001427:Bacterial Toxins; D003603:Cytotoxins; D017931:DNA Primers; D011993:Recombinant Fusion Proteins