Fig. 5: ResR/McdR regulates protein synthesis in Mtb.
a Ribosome profile of Mtb strains. Different subunits of ribosome were fractionated by ultracentrifugation of equal amount of lysates from empty vector control, resR/mcdR(−) and resR/mcdR(−)::resR/mcdR strains of Mtb mc2 7902, after 7 days of incubation with 50 ng/ml ATc. Values of absorbance at 260 nm (A260) from a total of 30 fractions were plotted in a graph showing the status of 70 S ribosome as well as small (30 S) and large (50 S) subunits in all the three samples. b Effect of resR/mcdR silencing on initiation of protein synthesis in Mtb. Empty vector control, resR/mcdR(−) and resR/mcdR(−)::resR/mcdR strains of Mtb Erdman were incubated with 50 µg/ml puromycin for 1 hour after 7 days of ATc treatment. Lysates prepared from the respective strains were subjected to anti-puromycin immunoblotting, which reveals significant inhibition of newly translated proteins upon resR/mcdR silencing, which is restored by complementation with the wild-type copy of resR/mcdR. Immunoblotting was performed with an equal amount (30 µg) of protein lysates, as ascertained by the ponceau S staining of the membrane. The arrows on the left in b indicate the positions of molecular weight markers. Molecular weight markers were accentuated by hand as the signal faded after several washes of the blot. kDa, kilo Dalton.
