FIG 2.

MiaA effects are independent of Hfq polarity. (A) Genetic and transcriptional organization of the complex yjeF-tsaE-amiB-mutL-miaA-hfq-hflX-hflK-hflC operon. The approximate location of the miaA::kan insertion mutation is highlighted. (B) Overnight cultures of wild-type (EM1050) and miaA rpoS750-lacZ (KMT31) translational fusion strains were grown as described in Fig. 1B and subjected to Western blot analysis using polyclonal Hfq antisera preadsorbed to crude cell lysates of hfq mutant cultures. These were the same samples analyzed in Fig. 1B. (C) Cultures of wild-type rpoS750-lacZ and miaA rpoS750-lacZ strains containing pBAD24 (KMT69; KMT54), pBAD-miaA (KMT70; KMT55), and pBAD-hfq (KMT71; KMT56) were grown in 50 ml of LB (Lennox) liquid medium supplemented with 50 μg of ampicillin/ml. These cultures were grown at 37°C in shaking water baths, without or with 0.002% arabinose to induce expression of either miaA or hfq. Cultures were grown to early stationary phase, i.e., an OD₆₀₀ of 1.5 to 2.0, and 100-μl culture aliquots were taken for β-galactosidase activity measurements. Each value represents the mean of at least three replicate experiments; the error bars represent the SEM.