FIG. 5.

The T4S system-dependent cytoplasmic stress response is mediated by σ³². (A) Schematic representation of the experimental setup used for primer extension analyses of the groESL transcript which is represented by the waved line. For reverse transcription, a Cy5-labeled oligonucleotide (indicated by an arrow) complementary to the sequence downstream from the σ³² transcription start site was used. AUG indicates the translation start site for the GroES protein within the groESL transcript. (B) Primer extension analyses with RNA isolated 15 min after dilution into fresh medium. The signal indicated by an arrowhead corresponds to transcripts originating from the transcription start site (+1) of the groESL σ³² promoter. The four lanes on the left show the DNA sequence surrounding the σ³²-dependent transcription start site. Corresponding nucleotides in the DNA and RNA sequences are boxed. (C) Steady-state levels of σ³² protein are elevated when cells express a functional T4S system. Western blot analyses detecting σ³² (32 kDa) and maltose binding protein (45 kDa) are shown. S, stationary phase; 15, number of minutes after feeding cells with fresh medium.