Measurement of SOS expression in individual Escherichia coli K-12 cells using fluorescence microscopy.
Many recombination, DNA repair and DNA replication mutants have high basal levels of SOS expression as determined by a sulAp-lacZ reporter gene system on a population of cells. Two opposing models to explain how the SOS expression is distributed in these cells are: (i) the 'Uniform Expression Model (UEM)' where expression is evenly distributed in all cells or (ii) the 'Two Population Model (TPM)' where some cells are highly induced while others are not at all. To distinguish between these two models, a method to quantify SOS expression in individual bacterial cells was developed by fusing an SOS promoter (sulAp) to the green fluorescent protein (gfp) reporter gene and inserting it at attlambda on the Escherichia coli chromosome. It is shown that the fluorescence in sulAp-gfp cells is regulated by RecA and LexA. This system was then used to distinguish between the two models for several mutants. The patterns displayed by priA, dnaT, recG, uvrD, dam, ftsK, rnhA, polA and xerC mutants were explained best by the TPM while only lexA (def), lexA3 (ind-) and recA defective mutants were explained best by the UEM. These results are discussed in a context of how the processes of DNA replication and recombination may affect cells in a population differentially.
McCool
Jesse D
JD
Department of Microbiology, Morrill Science Center IV N203, University of Massachusetts at Amherst, Amherst, MA 01003, USA.
Long
Edward
E
Petrosino
Joseph F
JF
Sandler
Hilary A
HA
Rosenberg
Susan M
SM
Sandler
Steven J
SJ
D000251:Adenosine Triphosphatases / Q000235:genetics / Q000378:metabolism; D004249:DNA Damage; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism; D004260:DNA Repair; D048168:Escherichia coli K12 / Q000166:cytology / Q000235:genetics* / Q000502:physiology* / Q000528:radiation effects; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D017930:Genes, Reporter; D008856:Microscopy, Fluorescence; D008957:Models, Genetic; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D013014:SOS Response, Genetics; D014466:Ultraviolet Rays
D029968:Escherichia coli Proteins; D011993:Recombinant Fusion Proteins; D000251:Adenosine Triphosphatases; C506746:UvrD protein, E coli; C506743:priA protein, E coli; D004265:DNA Helicases