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Journal of Bacteriology, July 2000, p. 4059-4067, Vol. 182, No. 14
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
Inactivation of the inhA-Encoded Fatty Acid Synthase
II (FASII) Enoyl-Acyl Carrier Protein Reductase Induces
Accumulation of the FASI End Products and Cell Lysis of
Mycobacterium smegmatis
Catherine
Vilchèze,1
Hector R.
Morbidoni,1
Torin R.
Weisbrod,1
Hiroyuki
Iwamoto,2
Mack
Kuo,2
James C.
Sacchettini,2 and
William R.
Jacobs Jr.1,*
Howard Hughes Medical Institute, Department
of Microbiology and Immunology, Albert Einstein College of Medicine,
Bronx, New York 10461,1 and Department
of Biochemistry and Biophysics, Texas A&M University, College
Station, Texas 778432
Received 10 February 2000/Accepted 19 April 2000
The mechanism of action of isoniazid (INH), a first-line
antituberculosis drug, is complex, as mutations in at least five different genes (katG, inhA, ahpC,
kasA, and ndh) have been found to correlate
with isoniazid resistance. Despite this complexity, a preponderance of
evidence implicates inhA, which codes for an enoyl-acyl
carrier protein reductase of the fatty acid synthase II (FASII), as the
primary target of INH. However, INH treatment of Mycobacterium
tuberculosis causes the accumulation of hexacosanoic acid
(C26:0), a result unexpected for the blocking of an
enoyl-reductase. To test whether inactivation of InhA is identical to
INH treatment of mycobacteria, we isolated a temperature-sensitive
mutation in the inhA gene of Mycobacterium
smegmatis that rendered InhA inactive at 42°C. Thermal
inactivation of InhA in M. smegmatis resulted in the
inhibition of mycolic acid biosynthesis, a decrease in hexadecanoic
acid (C16:0) and a concomitant increase of tetracosanoic acid (C24:0) in a manner equivalent to that seen in
INH-treated cells. Similarly, INH treatment of Mycobacterium
bovis BCG caused an inhibition of mycolic acid biosynthesis, a
decrease in C16:0, and a concomitant accumulation of
C26:0. Moreover, the InhA-inactivated cells, like
INH-treated cells, underwent a drastic morphological change, leading to
cell lysis. These data show that InhA inactivation, alone, is
sufficient to induce the accumulation of saturated fatty acids, cell
wall alterations, and cell lysis and are consistent with InhA being a
primary target of INH.
*
Corresponding author. Mailing address: Howard Hughes
Medical Institute, Department of Microbiology and Immunology, Albert Einstein College of Medicine, 1300 Morris Park Ave., Bronx, NY 10461. Phone: (718) 430-2888. Fax: (718) 518-0366. E-mail:
jacobs{at}aecom.yu.edu.
Journal of Bacteriology, July 2000, p. 4059-4067, Vol. 182, No. 14
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
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