TY - JOUR
T1 - Sigma factor displacement from RNA polymerase during Bacillus subtilis sporulation
AU - Ju, Jingliang
AU - Mitchell, Theresa
AU - Peters, Howard
AU - Haldenwang, W. G.
PY - 1999/8
Y1 - 1999/8
N2 - As Bacillus subtilis proceeds through sporulation, the principal vegetative cell σ subunit (σ(A)) persists in the cell but is replaced in the extractable RNA polymerase (RNAP) by sporulation-specific σ 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 σ(A) and two mother cell or factors, σ(E) and σ(K), which successively replace σ(A) on RNAP. Although the relative abundance of σ(A) with respect to RNAP remained virtually unchanged during sporulation, the percentage of the detectable σ(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 σ(E), the first of the mother cell-specific σ factors, approximately 40% of the σ(A) remained associated with RNAP at T3. The level of σ(A)-RNAP cosedimentation dropped to less than 10% in a strain which synthesized a σ(E) variant (σ(ECR119)) that could bind to RNAP but was unable to direct σ(E)-dependent transcription. The E-σ(E)-to-E-σ(K) changeover was characterized by both the displacement of σ(E) from RNAP and the disappearance of σ(E) from the cell. Analyses of extracts from wild-type and mutant B. subtilis showed that the σ(K) protein is required for the displacement of σ(E) from RNAP and also confirmed that σ(K) is needed for the loss of the σ(E) protein. The results indicate that the successive appearance of mother cell σ factors, but not necessarily their activities, is an important element in the displacement of preexisting σ factors from RNAP. It suggests that competition for RNAP by consecutive sporulation σ factors may be an important feature of the holoenzyme changeovers that occur during sporulation.
AB - As Bacillus subtilis proceeds through sporulation, the principal vegetative cell σ subunit (σ(A)) persists in the cell but is replaced in the extractable RNA polymerase (RNAP) by sporulation-specific σ 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 σ(A) and two mother cell or factors, σ(E) and σ(K), which successively replace σ(A) on RNAP. Although the relative abundance of σ(A) with respect to RNAP remained virtually unchanged during sporulation, the percentage of the detectable σ(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 σ(E), the first of the mother cell-specific σ factors, approximately 40% of the σ(A) remained associated with RNAP at T3. The level of σ(A)-RNAP cosedimentation dropped to less than 10% in a strain which synthesized a σ(E) variant (σ(ECR119)) that could bind to RNAP but was unable to direct σ(E)-dependent transcription. The E-σ(E)-to-E-σ(K) changeover was characterized by both the displacement of σ(E) from RNAP and the disappearance of σ(E) from the cell. Analyses of extracts from wild-type and mutant B. subtilis showed that the σ(K) protein is required for the displacement of σ(E) from RNAP and also confirmed that σ(K) is needed for the loss of the σ(E) protein. The results indicate that the successive appearance of mother cell σ factors, but not necessarily their activities, is an important element in the displacement of preexisting σ factors from RNAP. It suggests that competition for RNAP by consecutive sporulation σ factors may be an important feature of the holoenzyme changeovers that occur during sporulation.
UR - http://www.scopus.com/inward/record.url?scp=0032817065&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0032817065&partnerID=8YFLogxK
U2 - 10.1128/jb.181.16.4969-4977.1999
DO - 10.1128/jb.181.16.4969-4977.1999
M3 - Article
C2 - 10438769
AN - SCOPUS:0032817065
VL - 181
SP - 4969
EP - 4977
JO - Journal of Bacteriology
JF - Journal of Bacteriology
SN - 0021-9193
IS - 16
ER -