Latest revision as of 02:34, 29 October 2011

Team IGEM Paris 2011

The Bacillus subtilis sin operon

The sin operon of Bacillus subtilis can be turned into a bistable switch [1]. This cellular system has evolved to deal with short-term adaptation to environmental fluctuation by implementation of the appropriate stress responses such as sporulation.

Sporulation is a highly regulated phenomenon as it has a crucial role in stress response and is therefore under a huge selective pressure. It has been demonstrated that the sin operon is central to the timing and early dynamics of this network [2].

As shown by the scheme this operon is under control of the repressor SinR. Derepression is mediated by the antirepressor SinI [3]. Sporulation is also dependent on the master regulator Spo0A [4].

Overexpression of KinA, a kinase protein which phosphorylates Spo0AP, can trigger sporulation with high efficiency in cells in the exponential phase in rich medium by artificial induction [5]. Then, the activated form of Spo0AP induces the expression of SinI, which in turn inactivates SinR. The combined effect of positive regulation by Spo0AP and the inactivation of the negative regulator SinR triggers the sporulation pathway. We use SpoIIA gene as a reporter since it is is one of the first to be expressed when sporulation occurs [6].

In our design we want to diffuse kinA through the nanotubes in order to trigger sporulation in neighbouring cells, that are supposed to produce gfp once spoIIA will be activated.

For this design we used strains IIA/GFP (Veening et al 2006) and also IDJ011 Phyperspank-

References

C. a Voigt, D. M. Wolf, and A. P. Arkin, “The Bacillus subtilis sin operon: an evolvable network motif.,” Genetics, vol. 169, no. 3, pp. 1187-202, Mar. 2005.
J.-willem Veening, O. P. Kuipers, S. Brul, K. J. Hellingwerf, and R. Kort, “Effects of Phosphorelay Perturbations on Architecture , Sporulation , and Spore Resistance in Biofilms of Bacillus subtilis ‡,” Society, vol. 188, no. 8, pp. 3099-3109, 2006.
Y. Chai, F. Chu, R. Kolter, and R. Losick, “Bistability and biofilm formation in Bacillus subtilis.,” Molecular microbiology, vol. 67, no. 2, pp. 254-63, Jan. 2008.
M. Fujita and R. Losick, “Evidence that entry into sporulation in Bacillus subtilis is governed by a gradual increase in the level and activity of the master regulator Spo0A,” Genes & Development, pp. 2236-2244, 2005.
P. Eswaramoorthy, D. Duan, J. Dinh, A. Dravis, S. N. Devi, and M. Fujita, “The threshold level of the sensor histidine kinase KinA governs entry into sporulation in Bacillus subtilis.,” Journal of bacteriology, vol. 192, no. 15, pp. 3870-82, Aug. 2010.
M. Microbiology, “Chapter 3 Phosphatases modulate the bistable sporulation gene expression pattern in Bacillus subtilis,” Molecular Microbiology, vol. 1494, pp. 1481-1494, 2005.

@@ Line 2: / Line 2: @@
 <html>
 <h1>The Bacillus subtilis sin operon</h1>
-<h2>Introduction</h2>
+<p>The sin operon of Bacillus subtilis can be turned into a bistable switch <a href="https://2011.igem.org/Team:Paris_Bettencourt/SinOp#references">[1]</a>. This cellular system has evolved to deal with short-term adaptation to environmental fluctuation by implementation of the appropriate stress responses such as sporulation.</p>
-<p>The sin operon of bacillus subtilis can be tuned in a bistable switch <a href="https://2011.igem.org/Team:Paris_Bettencourt/SinOp#references">[1]</a>. This cellular systems has evolved to deal with short-term adaptation to environmental fluctuation by implementation of the appropriate stress responses. Sin operon is involved in many stress situation, such as biofilm formation, sporulation, etc.</p>
+<p>Sporulation is a highly regulated phenomenon as it has a crucial role in stress response and is therefore under a huge selective pressure. It has been demonstrated that the sin operon is central to the timing and early dynamics of this network <a href="https://2011.igem.org/Team:Paris_Bettencourt/SinOp#references">[2]</a>.
 <center><img src="https://static.igem.org/mediawiki/2011/7/7a/Sin_op1.png" style="width:600px"></center>
-<p>This operon is under control of the repressor SinR. Derepression is mediated by the antirepressor SinI, which binds to SinR with a 1:1 stoichiometry.</p>
+<p>As shown by the scheme this operon is under control of the repressor SinR. Derepression is mediated by the antirepressor SinI <a href="https://2011.igem.org/Team:Paris_Bettencourt/SinOp#references">[3]</a>. Sporulation is also dependent on the master regulator Spo0A <a href="https://2011.igem.org/Team:Paris_Bettencourt/SinOp#references">[4]</a>.</p>
-<p>Biofilm formation and sporulation are also connected in that both processes are dependent on Spo0A, the master regulator for entry into  sporulation <a href="https://2011.igem.org/Team:Paris_Bettencourt/SinOp#references">[2]</a>.</p>
+<p>Overexpression of KinA, a kinase protein which phosphorylates Spo0AP, can trigger sporulation with high efficiency in cells in the exponential phase in rich medium by artificial induction <a href="https://2011.igem.org/Team:Paris_Bettencourt/SinOp#references">[5]</a>. Then, the activated form of Spo0AP induces the expression of SinI, which in turn inactivates SinR. The combined effect of positive regulation by Spo0AP and the inactivation of the negative regulator SinR triggers the sporulation pathway. We use SpoIIA gene as a reporter since it is is one of the first to be expressed when sporulation occurs <a href="https://2011.igem.org/Team:Paris_Bettencourt/SinOp#references">[6]</a>.</p>
-<h2>Sporulation</h2>
+<p>In our design we want to diffuse kinA through the nanotubes in order to trigger sporulation in neighbouring cells, that are supposed to produce gfp once spoIIA will be activated.</p>
-<p>Sporulation is a dramatic response to stress and is a particularly expensive endeavor for the cell, in terms of both time and materials. The exact conditions and timing for sporulation are likely to be under strong selective pressure as both premature and belated spore production can have disastrous effects on cell growth and survival. Thus, an intricate phosphorelay and a transcriptional regulatory network carefully control the onset of sporulation .The sin operon is central to the timing and early dynamics of this network.</p>
-<center><img src="https://static.igem.org/mediawiki/2011/7/7b/Sin_op2.png" style="width:600px"></center>
-<p>Sporulation can be triggered with high efficiency in cells in the exponential phase of growth in rich medium by artificial induction of the synthesis of KinA, a kinase protein which phosphorylates Spo0AP. The accumulation of Spo0AP induces the expression of SinI, which binds to and inactivates SinR. The combined effect of positive regulation by Spo0AP and the inactivation of the negative regulator SinR activates the sporulation pathway. SpoIIA gene is early express in this situation.</p>
 <center><img src="https://static.igem.org/mediawiki/2011/2/28/SinOp.003.png" style="width:750px;"></center>
 <center><img src="https://static.igem.org/mediawiki/2011/5/5f/SinOp.004.png" style="width:750px;"></center>
-<h2>Biofilm formation</h2>
+<p>For this design we used strains IIA/GFP  (Veening et al 2006) and also IDJ011 Phyperspank-</p>
-<p>Biofilms of Bacillus subtilis consist of long chains of cells that are held together in bundles by an extracellular matrix of exopolysaccharide The exopolysaccharide is produced by enzymes encoded by the epsA-O operon operons are under the control of the repressor SinR. Derepression is mediated by the antirepressor SinI.</p>
-<p>In B.subtilis, the matrix consists of an exopolysaccharide, which is produced under the direction  of the 15-gene operon epsA-O.</p>
-<center><img src="https://static.igem.org/mediawiki/2011/2/23/SinOp.001.png" style="width:750px;"></center>
-<center><img src="https://static.igem.org/mediawiki/2011/b/ba/SinOp.002.png" style="width:750px;"></center>
-<h2>Perspectives</h2>
-<p>One area we want to investigate in the future is the synchronization observed in biofilm formation and sporulation. We would very much like to see if nanotubes could be responsible for this synchronization or at least what their impact on such phenomena is.</h2>
 <div id="citation_box">
 <p id="references">References</p>
 <ol>
-<li>Voigt, Christopher Wolf, Denise Arkin, Adam P 2005</li>
+<li>C. a Voigt, D. M. Wolf, and A. P. Arkin, “The Bacillus subtilis sin operon: an evolvable network motif.,” Genetics, vol. 169, no. 3, pp. 1187-202, Mar. 2005.</li>
-<li>Sonenshein, 2000; Branda et al., 2001; Hamon and Lazazzera, 2001</li>
+<li>J.-willem Veening, O. P. Kuipers, S. Brul, K. J. Hellingwerf, and R. Kort, “Effects of Phosphorelay Perturbations on Architecture , Sporulation , and Spore Resistance in Biofilms of Bacillus subtilis ‡,” Society, vol. 188, no. 8, pp. 3099-3109, 2006.</li>
+<li>Y. Chai, F. Chu, R. Kolter, and R. Losick, “Bistability and biofilm formation in Bacillus subtilis.,” Molecular microbiology, vol. 67, no. 2, pp. 254-63, Jan. 2008.</li>
+<li>M. Fujita and R. Losick, “Evidence that entry into sporulation in Bacillus subtilis is governed by a gradual increase in the level and activity of the master regulator Spo0A,” Genes & Development, pp. 2236-2244, 2005.</li>
+<li>P. Eswaramoorthy, D. Duan, J. Dinh, A. Dravis, S. N. Devi, and M. Fujita, “The threshold level of the sensor histidine kinase KinA governs entry into sporulation in Bacillus subtilis.,” Journal of bacteriology, vol. 192, no. 15, pp. 3870-82, Aug. 2010.</li>
+<li>M. Microbiology, “Chapter 3 Phosphatases modulate the bistable sporulation gene expression pattern in Bacillus subtilis,” Molecular Microbiology, vol. 1494, pp. 1481-1494, 2005.</li>
 <ol>
 </div>

Team:Paris Bettencourt/SinOp

From 2011.igem.org

Latest revision as of 02:34, 29 October 2011

The Bacillus subtilis sin operon