TraR, a Homolog of a RNAP Secondary Channel Interactor, Modulates Transcription
Figure 3
Inhibition of the rrnB P1 Promoter by Ectopically Expressed TraR.
Differential β-galactosidase activity of the rrnB P1-lacZ promoter fusion in LB media. (A,B) Wild-type background. (A) Effects of uninduced ectopic expression of TraR (open triangles) or DksA (open squares) on rrnB P1 activity. Plasmid control indicated by open circles. Rates of β-galactosidase synthesis in exponential phase (from OD600 0.2 to 0.6) are ∼100, 100, and 84 β-gal activity/OD600 for pControl, pDksA, and pTraR, respectively. R2 values>0.95 in this linear range. (B) Induced (0.1 mM IPTG) ectopic expression of TraR (triangles) or DksA (squares) represses rrnB P1 activity. Circles, plasmid control. Rates of β-galactosidase synthesis in exponential phase (from OD600 0.2 to 0.6) are ∼95, 67, and 2.0 β-gal activity/OD600 for pControl, pDksA, and pTraR, respectively. R2 values>0.95 in this linear range. (C,D) ΔdksA background. (C) Repression of β-galactosidase activity of rrnB P1-lacZ from uninduced ectopic expression of TraR (open triangles) or DksA (open squares) in the ΔdksA cells. Plasmid control indicated by open circles. Rates of β-galactosidase synthesis in exponential phase (from OD600 0.2 to 0.6) are ∼140, 84, and 88 β-gal activity/OD600 for pControl, pDksA, and pTraR, respectively. R2 values>0.95 in this linear range. (D) Induced (0.1 mM IPTG) ectopic expression of TraR (triangles) or DksA (squares) represses rrnB P1 activity in ΔdksA. Circles, plasmid control. Rates of β-galactosidase synthesis in exponential phase (from OD600 0.2 to 0.6) are ∼140, 80, and 3.3 β-gal activity/OD600 for pControl, pDksA, and pTraR, respectively. R2 values>0.95 in this linear range. (E) TraR does not affect the Plac promoter. β-galactosidase activity assays of the wild-type lac operon. TraR (triangles) induced (0.1 mM IPTG) from a multi-copy plasmid. Control plasmid indicated by circles.