Team:Heidelberg LSL/Project SOS


iGEM-2012HS - LSL-Heidelberg iGEM-2012HS - LSL-Heidelberg

Scientific Background - The SOS response

In order to realize our project, we took advantage of the DNA-repair machinery of the bacterium E. coli, the so-called SOS response system. The SOS system is the repair system of the cell to counteract extensive DNA damages e.g. caused by UV radiation or ionizing radiation. If UV radiation impinges on the bacterial DNA it may cause cross-linking of adjacent cytosine and thymine bases resulting in pyrimidine dimers – a process called direct DNA damage. Heavily damaged DNA is no longer able to replicate and leads to interruption of the cell cycle. The SOS response allows bacterial cells to repair damaged DNA and continue the cell cycle. In normal cells, the SOS system is turned off and it is only activated in the case of DNA damage. The central regulator of the SOS response is recA. The recA protein is activated by single-stranded DNA which is induced through strong DNA damages. Activated recA leads to the cleavage of the lexA-repressor protein that normally blocks the RNA-polymerase for transcription of repair proteins. The activated LexA can no longer bind to the DNA strain and allows the production of a cascade of about 20 different repair proteins, amongst others sulA, uvrB and dinF. Another action of the activated lexA repressor protein is regulation of its own transcription as well as transcription of recA . When DNA repair is finished and the concentration of activated recA has decreased, the system returns to its steady state.

RecA, lexA, sulA and roughly 15 other proteins are main members of the SOS response. Amongst 20 different promoters, we have chosen recA and sulA for our experiments. We used the following criteria for the selection of the optimal promoters:

  • They should be involved in only a few processes in the cell to avoid hazardous disorders.
  • The promoters should already be well known, tested and available in the parts registry.

Only recA and sulA meet these criteria. Although recA has many functions in the cell, it is a standard gene for the SOS response and well characterized. Another wanted feature for our project is the ability to detect the amount of UV radiation by changes in the intensity of color development. Therefore, we are trying to use other rec-proteins besides recA such as recB, recC etc. Our hypothesis is that these rec-proteins may be activated at different UV mediated damages.

GeneRepair Function
lexASOS repressor
recASOS regulator; SOS mutagenesis; recF-dependent recombinational repair; recB-dependent repair of double-strand gaps; cross-link repair
recNrecF-dependent recombinational repair; repair of double-strand gaps
recQrecF-dependent recombinational repair
ruvrecF-dependent recombinational repair
umuCSOS mutagenesis (Error prone repair)
umuDSOS mutagenesis (Error prone repair)
uvrAShort-patch nucleotide-excision repair; long-patch nucleotide-excision repair; cross-link repair
uvrBShort-patch nucleotide-excision repair; long-patch nucleotide-excision repair; cross-link repair
uvrDShort-patch nucleotide-excision repair; recF-dependent recombinational repair; recB-dependent repair of double-strand gaps; cross-link repair; methylation-directed mismatch repair
dinASOS mutagenesis
sulAInhibitor of cell division