A cka-gfp Transcriptional Fusion Reveals that the Colicin K Activity Gene Is Induced in Only 3 Percent of the Population

Abstract

In prokaryotes, only a few examples of differential gene expression in cell populations have been described. Colicin production in natural populations of Escherichia coli, while providing a competitive advantage in the natural habitat, also leads to lysis of the toxin-producing cell. Colicin K synthesis has been found to be induced due to an increase in ppGpp (I. Kuhar, J. P. van Putten, D. Žgur-Bertok, W. Gaastra, and B. J. Jordi, Mol. Microbiol. 41:207-216). Using two transcriptional fusions, cka-gfp and cki-gfp, we show that at the single-cell level, the colicin K activity gene cka is expressed in only 3% of the bacterial population upon induction by nutrient starvation. In contrast, the immunity gene cki is expressed in the large majority of the cells. Expression of the cka-gfp fusion in a lexA-defective strain and in a relA spoT mutant strain indicates that differential expression of cka is established primarily at the level of transcription.

Heterogeneity in expression of the Escherichia coli colicin K activity gene cka is controlled by the SOS system and stochastic factors

“…6; Table 2). It should be noted that the morphology of cells expressing the cka gene from the cka-gfp fusion did not diVer from nonexpressing cells (Mulec et al 2003).…”

“…5). For control strain RW542 lexA542 (Def) with a defective LexA protein that cannot bind to DNA, virtually all cells showed intense Xuorescence (Mulec et al 2003). To ensure that the eVects were due to LexA activity rather than the general SOS response, a high copy number plasmid without LexA binding sequences (pUC19) was tested for its eVect on lexAgfp expression in strain AB1157L.…”

“…To test this hypothesis we increased the intracellular number of LexA binding sites which should result in titration of the LexA protein. For this purpose plasmids pERO22 and pERO19 were constructed, harbouring the wild type cka-gfp fusion (Mulec et al 2003) in pBR322 and pUC19 that are present in approximately 50 and 500 copies per cell, respectively. Expression of the cka-gfp fusion was subsequently analysed at the single cell level by Xuorescence microscopy.…”

“…The bacterial cells were prepared for Xuorescence microscopy as described previously (Mulec et al 2003). Fluorescence in single cells was detected using a Carl Zeiss Axiovert 135 microscope, equipped with a GFP Wlter (Chroma).…”

“…Therefore, only a part of the population should express the colicin activity and lysis genes. It was shown that in the stationary phase, transcription from the cka promoter is derepressed in only approximately 3% of the colicinogenic population and that the LexA protein is a decisive regulatory element repressing expression at the level of transcription in the large majority of the population (Mulec et al 2003).…”