ANTHRAX LETHAL FACTOR CLEAVES THE N-TERMINUS OF MAPKKS AND INDUCES TYROSINE THREONINE PHOSPHORYLATION OF MAPKS IN CULTURED MACROPHAGES/

Lethal factor (LF) is the major virulence factor produced by Bacillus anthracis. LF is sufficient to cause death in laboratory animals and c ytolysis of peritoneal macrophages and macrophage cell lines. LF conta ins the characteristic zinc binding motif of metalloproteases and indi rect evidence suggest that this hydrolytic activity is essential for i ts cytotoxicity. To identify the substrate(s) of LF, we have used the yeast two-hybrid system, employing a LF inactive mutant as bait. This approach has led to the identification of the MAP kinase kinases (MAPK Ks) Mek1 and Mek2 as proteins capable of specific interaction with LF. LF cleaves Mek1 and Mek2 within their N-terminus in vitro and in vivo , hydrolyzing a Pro8-Ile9 and a Pro10-Arg11 peptide bond in Mek1 and M ek2 respectively, The removal of the amino terminus of MAPKKs eliminat es the ''docking site'' for the MAPKs ERK1 and ERK2, which become phos phorylated in cultured macrophages following toxin challenge. The poss ible implications of these findings for the cytolysis of macrophage ce lls induced by LF are discussed. These results open the way to the des ign and screening of specific inhibitors of LF. (C) 1998 Academic Pres s.