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Journal of Bacteriology, March 1999, p. 1380-1387, Vol. 181, No. 5
Department of Medical Microbiology, St.
George's Hospital Medical School, London SW17 ORE, United Kingdom
Received 28 September 1998/Accepted 14 December 1998
The 16-kDa protein, an
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
Transcription of the Stationary-Phase-Associated
hspX Gene of Mycobacterium tuberculosis Is
Inversely Related to Synthesis of the 16-Kilodalton
Protein
-crystallin homologue, is one of the most
abundant proteins in stationary-phase Mycobacterium
tuberculosis. Here, transcription and translation of the
hspX gene, which encodes the 16-kDa protein, have been
investigated by Northern blotting analysis, primer extension, and
sodium dodecyl sulfate-polyacrylamide gel electrophoresis with a
microaerophilic stationary-phase model. Two transcripts of about 2.5 and 1.1 kb were demonstrated by Northern blot analysis and hybridized
to the hspX gene probe. Primer extension analysis revealed
that the transcription start site is located 33 nucleotides upstream of
the hspX gene start codon. The cellular level of the
hspX mRNA was maximum in log-phase bacilli and was markedly
reduced after 20 days in unagitated culture, when the organisms had
entered the stationary phase. A third transcript of 0.5 kb was detected
0.6 kb downstream of the hspX gene; this transcript has a
transcriptional pattern completely different from that of the 1.1- and
2.5-kb products, suggesting that there may be another gene in this
region. In contrast to the high level of hspX mRNA in
log-phase bacilli, 16-kDa protein synthesis was low in log-phase
bacteria and rose to its maximum after 20 days. In both log-phase and
stationary-phase bacteria the mRNA was unstable, with a half-life of 2 min, which indicated that the transcript stability was growth rate
independent and not a general means for controlling the gene
expression. However, the cellular content of 16-kDa protein, while low
in log-phase bacteria, rose to a maximum at 10 days and remained at
this high level for up to 50 days, which indicates that this protein is
a stable molecule with a low turnover rate. These data suggest that the
regulation of hspX expression during entry into and
maintenance of stationary phase involves translation initiation
efficiency and protein stability as potential mechanisms.
*
Corresponding author. Mailing address: Department of
Medical Microbiology, St. George's Hospital Medical School, London
SW17 ORE, United Kingdom. Phone: 44 (181) 725 5725. Fax: 44 (181) 672 0234. E-mail: acoates{at}sghms.ac.uk.
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