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Ut wisi enim ad minim veniam, quis nostrud exerci tation ullamcorper suscipit lobortis nisl ut aliquip ex ea commodo consequat. Duis autem vel eum iriure dolor in hendrerit in vulputate velit esse molestie consequat, vel illum dolore eu feugiat nulla facilisis at vero eros et accumsan et iusto odio dignissim qui blandit praesent luptatum zzril delenit augue duis dolore te feugait nulla facilisi. </p> <img src="images/img/slide02.jpg" width="100%"> <p> Investigationes demonstraverunt lectores legere me lius quod ii legunt saepius. Claritas est etiam processus dynamicus, qui sequitur mutationem consuetudium lectorum. Lorem ipsum dolor sit amet, consectetuer adipiscing elit, sed diam nonummy nibh euismod tincidunt ut laoreet dolore magna aliquam erat volutpat. Ut wisi enim ad minim veniam, quis nostrud exerci tation ullamcorper suscipit lobortis nisl ut aliquip ex ea commodo consequat. Duis autem vel eum iriure dolor in hendrerit in vulputate velit esse molestie consequat, vel illum dolore eu feugiat nulla facilisis at vero eros et accumsan et iusto odio dignissim qui blandit praesent luptatum zzril delenit augue duis dolore te feugait nulla facilisi. Nam liber tempor cum soluta nobis eleifend option congue nihil imperdiet doming id quod mazim placerat facer possim assum. </p> </div> <div class="split1_3r"> <h2>Twitter</h2> <a class="twitter-timeline" href="https://twitter.com/iGEM_UniSalento" data-widget-id="356106138686722049">Tweets di @iGEM_UniSalento</a> <script>!function(d,s,id){var js,fjs=d.getElementsByTagName(s)[0],p=/^http:/.test(d.location)?'http':'https';if(!d.getElementById(id)){js=d.createElement(s);js.id=id;js.src=p+"://platform.twitter.com/widgets.js";fjs.parentNode.insertBefore(js,fjs);}}(document,"script","twitter-wjs");</script> </div> <div class="clear"></div>  <div class="nosplit"> <h2>Introduction to NickBusters Project</h2> <p> Nickel is one of the most widespread heavy metals in the ecosystem, and it is essential for many organisms, from bacteria to higher forms of life. However, as all the heavy metals, even a small amount of Nickel higher than the essential dose could be toxic, leading to various noxious effects <a href="#biblio" style="color: #8ac101">[1-2]</a>: then the need to remove its excess from many substrates. <b>Nowadays bacteria-mediated</b> bioremediation from inorganic substances seems to be a considerably relevant frontier in microbic biotechnologies <a href="#biblio" style="color: #8ac101">[3]</a>. Our project aims to develop a living system, in an easy monitorable bacterial platform, who would work as a <b>Nickel detector</b> and a <b>Nickel remediation</b> system. The whole system is based on genetic parts from the pathogen <i>Helicobacter pylori</i>, whose evolution led to a stronger reliance on Nickel ions for host interaction and pathogenesis. <b>Sensing</b> relies on nickel regulator <i>NikR</i> from <i>H. pylori</i> (<i>HpNikR</i>): though homolog of <i>E.coli NikR</i>, it has a stronger dependance on Nickel ions and a wider range of regulated elements, being a rare case of pleiotropic regulator in bacteria. As <i>HpNikR</i> could contemporaneously activate and repress gene expression <a href="#biblio" style="color: #8ac101">[4-5-6]</a>, we thought of developing a synthetic gene cluster under <i>HpNikR</i> - regulated promoters. With a <b>double regulation</b>, one of the promoters being activated while the other being repressed, the system allows to follow up Nickel sensing as a fluorescence signal from fluorescent proteins. Thus, adding bacteria to a nickel containing medium results in <b>switching off a basal occuring green fluorescence</b> and in <b>switching on a red fluorescence signal</b>. </p> <br/> <img src="images/img/welcome1.png" alt="" width="100%"> <br/> <p> The Nickel <b>storage</b> system is even taken from our pathogenic friend: <i>Hpn</i> protein is a Nickel binding protein, whose role is to store the Nickel ions inside the cell and to supply them to other proteic interactors, from Nickel dependent heat shock proteins to pathogenesis factors as urease <a href="#biblio" style="color: #8ac101">[7]</a>. <i>Hpn</i> can be expressed in a heterologous model system (<i>E.coli</i>), increasing bacterial resistance to toxic dose of Nickel ions and allowing Ni++ intracellular accumulation <a href="#biblio" style="color: #8ac101">[8]</a>. We thought to split the <b>sense and store machineries in two different bacterial populations</b>, making them communicate through Quorum Sensing <a href="#biblio" style="color: #8ac101">[9]</a>: we used the <i>LuxI/LuxR</i> system from <i>Alivibrio fischerii</i>. The first population senses nickel concentration in medium ( and translates it in a fluorescence signal), then activating <i>Acyl-Homoserine-Lactones</i> (<i>AHLs</i>) production. The second population receives the <i>QS</i> signal and sums it up to the nickel detection signal (as in an AND gate), activating <i>Hpn</i> (the storage protein) expression and <i>AHLs</i> signal amplification. </p> <br/> <img src="images/img/welcome2.png" alt="" width="100%"> <br/> <br/> </div> <div class="clear"></div>  <img src="images/img/dna.png" alt="" style="margin-bottom: -100px; margin-left: 360px; position: absolute">  </div>  <div id="content">  <div class="nosplit"> <h2><a name="biblio">Bibliography</a></h2> <ol> <li>Denkhaus and Salnikow, Nickel essentiality, toxicity and carcinogenity; Critical Reviews in Oncology/Hematology, 42 (2002) 35–56</li> <li>“Nickel, Elemental” on TOXNET- Databases on toxicology, hazardous chemicals, environmental health, and toxic releases.</li> <li>Gadd, Metals, minerals and microbes: geomicrobiology and bioremediation; Microbiology, 156 (2010), 609–643</li> <li>Dosanjh and Michel, Microbial nickel metalloregulation: NikRs for nickel ions; Current Opinion in Chemical Biology, 10 (2006), 123–130</li> <li>Contreras et al., Characterization of the roles of NikR, a nickel-responsive pleiotropic autoregulator of Helicobacter pylori; Molecular Microbiology, 49 (4) (2003), 947–963</li> <li>Muller et al., Hierarchical regulation of the NikR-mediated nickel response in Helicobacter pylori; Nucleic Acids Research, 39 (17) (2011), 7564–7575</li> <li>Seshadri et al., Roles of His-Rich Hpn and Hpn-Like Proteins in Helicobacter pylori Nickel Physiology; Journal of Bacteriology, 189 (11) (2007), 4120–4126</li> <li>Ge et al., Expression and characterization of a histidine-rich protein, Hpn: potential for Ni2+ storage in Helicobacter pylori; Biochemistry Journal 393 (2006), 285–293</li> <li>Miller, Bassler, Quorum Sensing in bacteria; Annual Review of Microbiology 55 (2001), 165-99.</li> </ol> </div>  </div>  </div> </div> <div class="clear"></div>  <footer></footer>   <p id="back-top"> <a href="#top"><span></span>Back to Top</a> </p>  </body> </html>