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Journal of Bacteriology, August 1999, p. 4969-4977, Vol. 181, No. 16
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Sigma Factor Displacement from RNA Polymerase during Bacillus subtilis Sporulation

Jingliang Ju, Theresa Mitchell, Howard Peters III,dagger and W. G. Haldenwang*

Department of Microbiology, University of Texas Health Science Center at San Antonio, San Antonio, Texas 78284-7758

Received 14 October 1998/Accepted 2 June 1999

As Bacillus subtilis proceeds through sporulation, the principal vegetative cell sigma  subunit (sigma A) persists in the cell but is replaced in the extractable RNA polymerase (RNAP) by sporulation-specific sigma  factors. To explore how this holoenzyme changeover might occur, velocity centrifugation techniques were used in conjunction with Western blot analyses to monitor the associations of RNAP with sigma A and two mother cell sigma  factors, sigma E and sigma K, which successively replace sigma A on RNAP. Although the relative abundance of sigma A with respect to RNAP remained virtually unchanged during sporulation, the percentage of the detectable sigma A which cosedimented with RNAP fell from approximately 50% at the onset of sporulation (T0) to 2 to 8% by 3 h into the process (T3). In a strain that failed to synthesize sigma E, the first of the mother cell-specific sigma  factors, approximately 40% of the sigma A remained associated with RNAP at T3. The level of sigma A-RNAP cosedimentation dropped to less than 10% in a strain which synthesized a sigma E variant (sigma ECR119) that could bind to RNAP but was unable to direct sigma E-dependent transcription. The E-sigma E-to-E-sigma K changeover was characterized by both the displacement of sigma E from RNAP and the disappearance of sigma E from the cell. Analyses of extracts from wild-type and mutant B. subtilis showed that the sigma K protein is required for the displacement of sigma E from RNAP and also confirmed that sigma K is needed for the loss of the sigma E protein. The results indicate that the successive appearance of mother cell sigma  factors, but not necessarily their activities, is an important element in the displacement of preexisting sigma  factors from RNAP. It suggests that competition for RNAP by consecutive sporulation sigma  factors may be an important feature of the holoenzyme changeovers that occur during sporulation.

* Corresponding author. Mailing address: Department of Microbiology, University of Texas Health Science Center at San Antonio, San Antonio, TX 78284-7758. Phone: (210) 567-3957. Fax: (210) 567-6612. E-mail: Haldenwang{at}UTHSCSA.EDU.

dagger Present address: Gene Regulation and Chromosome Biology Laboratory, ABL Basic Research Program, NCI-FCRDC, Frederick, MD 21702.

Journal of Bacteriology, August 1999, p. 4969-4977, Vol. 181, No. 16
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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