Response of Bacillus subtilis to cerulenin and
acquisition of resistance.
Schujman GE, Choi KH, Altabe S, Rock CO, de Mendoza D.
Instituto de Biologia Molecular y Celular de Rosario (IBR) and Departamento de
Microbiologia, Facultad de Ciencias Bioquimicas y Farmaceuticas, Universidad
Nacional de Rosario, Suipacha 531, 2000-Rosario, Argentina.
Cerulenin is a fungal mycotoxin that potently inhibits fatty acid synthesis by
covalent modification of the active site thiol of the chain-elongation subtypes
of beta-ketoacyl-acyl carrier protein (ACP) synthases. The Bacillus subtilis
fabF (yjaY) gene (fabF(b)) encodes an enzyme that catalyzes the condensation of
malonyl-ACP with acyl-ACP to extend the growing acyl chain by two carbons. There
were two mechanisms by which B. subtilis adapted to exposure to this antibiotic.
First, reporter gene analysis demonstrated that transcription of the operon
containing the fabF gene increased eightfold in response to a cerulenin
challenge. This response was selective for the inhibition of fatty acid
synthesis, since triclosan, an inhibitor of enoyl-ACP reductase, triggered an
increase in fabF reporter gene expression while nalidixic acid did not. Second,
spontaneous mutants arose that exhibited a 10-fold increase in the MIC of
cerulenin. The mutation mapped at the B. subtilis fabF locus, and sequence
analysis of the mutant fabF allele showed that a single base change resulted in
the synthesis of FabF(b)[I108F]. The purified FabF(b) and FabF(b)[I108F]
proteins had similar specific activities with myristoyl-ACP as the substrate.
FabF(b) exhibited a 50% inhibitory concentration (IC(50)) of cerulenin of 0.1
microM, whereas the IC(50) for FabF(b)[I108] was 50-fold higher (5 microM).
These biochemical data explain the absence of an overt growth defect coupled
with the cerulenin resistance phenotype of the mutant strain.