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Journal of Bacteriology, March 2001, p. 1655-1662, Vol. 183, No. 5
Department of Molecular Biophysics and
Biochemistry, Yale University, New Haven, Connecticut
06520-8114,1 and Izumi Campus, Meiji
University, Suginami, Tokyo 168-0064, Japan2
Received 9 October 2000/Accepted 6 December 2000
The MS ring of the flagellar basal body of Salmonella
is an integral membrane structure consisting of about 26 subunits of a
61-kDa protein, FliF. Out of many nonflagellate fliF
mutants tested, three gave rise to intergenic suppressors in flagellar region II. The pseudorevertants swarmed, though poorly; this partial recovery of motile function was shown to be due to partial recovery of
export function and flagellar assembly. The three parental mutants were
all found to carry the same mutation, a six-base deletion corresponding
to loss of Ala-174 and Ser-175 in the predicted periplasmic domain of
the FliF protein. The 19 intergenic suppressors identified all lay in
flhA, and they consisted of 10 independent examples at the
nucleotide level or 9 at the amino acid level. Since two of the nine
corresponded to different substitutions at the same amino acid
position, only eight positions in the FlhA protein have given rise to
suppressors. Thus, FliF-FlhA intergenic suppression is a fairly rare
event. FlhA is a component of the flagellar protein export apparatus,
with an integral membrane domain encompassing the N-terminal half of
the sequence and a cytoplasmic C-terminal domain. All of the
suppressing mutations lay within the integral membrane domain. These
mutations, when placed in a wild-type fliF background, had
no mutant phenotype. In the fliF mutant background, mutant
FlhA was dominant, yielding a pseudorevertant phenotype. Wild-type FlhA
did not exert significant negative dominance in the pseudorevertant
background, indicating that it does not compete effectively with mutant
FlhA for interaction with mutant FliF. Mutant FliF was partially
dominant over wild-type FliF in both the wild-type and second-site FlhA
backgrounds. Membrane fractionation experiments indicated that the
fliF mutation, though preventing export, was mild enough to
permit assembly of the MS ring itself, and also assembly of the
cytoplasmic C ring onto the MS ring. The data from this study provide
genetic support for a model in which at least the FlhA component of the
export apparatus physically interacts with the MS ring within which it is housed.
0021-9193/01/$04.00+0 DOI: 10.1128/JB.183.5.1655-1662.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Intergenic Suppression between the Flagellar MS
Ring Protein FliF of Salmonella and FlhA, a Membrane
Component of Its Export Apparatus
*
Corresponding author. Mailing address: Department of
Molecular Biophysics and Biochemistry 0734, Yale University, P.O. Box 208114, 266 Whitney Ave., New Haven, CT 06520-8114. Phone: (203) 432-5590. Fax: (203) 432-9782. E-mail:
robert.macnab{at}yale.edu.
Present address: Protonic Nanomachine Project, ERATO, JST, Seika,
Kyoto, 619-0237, Japan.
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