Difference between revisions of "Part:BBa K3788017"

 
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<partinfo>BBa_K3788017 short</partinfo>
 
<partinfo>BBa_K3788017 short</partinfo>
  
<p>This part is constituted from  -35 and -10 boxes, with a LexA box and the beggining of the CDS of <i>caa</i>.
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This part contains the LexA-responsive promoter region of the Colicin A gene, amplified from the Colicin A encoding plasmid (pRL1 or pColA).
The LexA box is a sequence that allows the binding of a tetramere of LexA that represses the trascription of the <i>caa-cal</i> operon.</p>
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<p>This repression can be stopped when the SOS response is activated, due to DNA damage that activate RecA production that induce LexA's depolymersation.</p>
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<h3> <u> Background information </u> </h3>
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<p>LexA is a transcriptional regulator that binds to the promoter regions of the numerous SOS-response regulatory genes, repressing the expression of the downstream genes. The consensus LexA binding box is CTG-N10-CAG. In response to the presence of DNA lesions (resulting from UV or mitomycin treament for exemple), the RecA protein is activated and stimulates the autocatalytic digestion of LexA, leading to the derepression of the promoter. </p>
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<br>
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<p>Colicin A is a plasmid-encoded toxin produced by some <i>E. coli</i> strains and active against other non-immune <i>E.coli</i>strains. Colicin A is encoded by the <i>caa </i>gene. The toxin production is induced by the SOS response. <i>caa </i>was demonstrated to be transcribed as an operon with the <i>cal</i> gene, encoding the lysis protein CaL (see <html><a href="https://parts.igem.org/Part:BBa_K3788018 ">BBa_K3788018</a></html>, <html><a href="https://parts.igem.org/Part:BBa_K3788016 ">BBa_K3788016</a></html> and <html><a href="https://parts.igem.org/Part:BBa_K3788015 ">BBa_K3788015</a></html> for details on CaL). Upon induction of the SOS system, Colicin A accumulates in the cell cytoplasm. The delayed production of the lysis protein results in a time-controlled lysis of the cells, and the release of colicin A in the environment. An immunity protein encoded by the <i>cai</i>gene is transcribed constitutively in the opposite direction of the <i>caa-cal</i> operon, insuring immunity of the pColA plasmid carrying strain from the lethal activity of the toxin.</p>
  
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<u>The <i>caa</i> promoter region:</u>
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<p>The -35 and -10 promoter elements, the +1 transcription site and LexA binding sites have been identified in the promoter region of <i>caa. </i></p>
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<p>Binding of LexA on the <i>caa </i>promoter was experimentaly demonstrated by footprinting experiments (1). LexA protein binds with a high affinity to an approximately 40 bp long sequence just downstream of the -10 element. The sequence of the binding site includes two overlapped SOS boxes. </p>
  
===Role in natural environment===
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[[File:T--Aix-Marseille--Organization-of-the-caa-cal-operon.png|600px|thumb|left|<b>Figure adapted from: Lloubes et al., Nucleic Acids Research 1986 (DOI: 10.1093/nar/14.6.2621)</b>. <i><u>Organization of the caa-cal operon and the cai gene</u>. The nucleotide sequences for the promoters are presented: the recognition and binding sequences for RNA polymerase are presented as yellow boxed(-35 and -10 elements). SA indicates the transcriptional start site for the caa-cal operon. Arrows above the start site indicates the direction of transcription, the bold line indicates the major band detected by SI mapping. The binding site for LexA in the operator region for caa is indicated by brackets overlining the protected region for the sense-strand and brackets underlining the sequence (not represented) for the anti-sense strand. The two overlapped consensus "SOS sequences" are presented as a pink box.</i> ]]
  
Induced by the SOS response (LexA dependent) upon DNA damages (UV, mitomycin…). This promoter allow the expression of the operon caa and cal. caa gene is coding for Colicin A (ColA) a toxin produced by E. coli strains, and active against other non-immune E.coli strains. E. coli have an imumnity protein coded by cai gene (-). cal gene is codding for a lysis protein, when this protein is produce ColA can be delivered in the extracellular medium after the E. coli cells lysis.
 
  
  

Latest revision as of 14:56, 21 October 2021


lexA-repressed promoter

This part contains the LexA-responsive promoter region of the Colicin A gene, amplified from the Colicin A encoding plasmid (pRL1 or pColA).

Background information

LexA is a transcriptional regulator that binds to the promoter regions of the numerous SOS-response regulatory genes, repressing the expression of the downstream genes. The consensus LexA binding box is CTG-N10-CAG. In response to the presence of DNA lesions (resulting from UV or mitomycin treament for exemple), the RecA protein is activated and stimulates the autocatalytic digestion of LexA, leading to the derepression of the promoter.


Colicin A is a plasmid-encoded toxin produced by some E. coli strains and active against other non-immune E.colistrains. Colicin A is encoded by the caa gene. The toxin production is induced by the SOS response. caa was demonstrated to be transcribed as an operon with the cal gene, encoding the lysis protein CaL (see BBa_K3788018, BBa_K3788016 and BBa_K3788015 for details on CaL). Upon induction of the SOS system, Colicin A accumulates in the cell cytoplasm. The delayed production of the lysis protein results in a time-controlled lysis of the cells, and the release of colicin A in the environment. An immunity protein encoded by the caigene is transcribed constitutively in the opposite direction of the caa-cal operon, insuring immunity of the pColA plasmid carrying strain from the lethal activity of the toxin.


The caa promoter region:

The -35 and -10 promoter elements, the +1 transcription site and LexA binding sites have been identified in the promoter region of caa.

Binding of LexA on the caa promoter was experimentaly demonstrated by footprinting experiments (1). LexA protein binds with a high affinity to an approximately 40 bp long sequence just downstream of the -10 element. The sequence of the binding site includes two overlapped SOS boxes.

Figure adapted from: Lloubes et al., Nucleic Acids Research 1986 (DOI: 10.1093/nar/14.6.2621). Organization of the caa-cal operon and the cai gene. The nucleotide sequences for the promoters are presented: the recognition and binding sequences for RNA polymerase are presented as yellow boxed(-35 and -10 elements). SA indicates the transcriptional start site for the caa-cal operon. Arrows above the start site indicates the direction of transcription, the bold line indicates the major band detected by SI mapping. The binding site for LexA in the operator region for caa is indicated by brackets overlining the protected region for the sense-strand and brackets underlining the sequence (not represented) for the anti-sense strand. The two overlapped consensus "SOS sequences" are presented as a pink box.




Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]