Team:Valencia UPV/Project/overview

From 2014.igem.org

(Difference between revisions)

Latest revision as of 02:15, 18 October 2014

Project > Overview

Project Overview

Pests cause great economical loss in agriculture and impede an optimal use of the resources. Sexy Plant rises as a pest control strategy based on the use of mating disruption by sex pheromones. The release of sex pheromones impedes male moths to find their females, avoiding crop-damaging larvae to be born.

The Sexy Plant is engineered to produce and release three moth sex pheromones which affect a broad number of Lepidoptera species (moths). We implemented previous work by Ding et al. [1] to produce the first two pheromones (Z11-16:OH and Z11-16:OAc). Inspired by Hagström et al [2], we planned the introduction of a Fatty Acid reductase to transform Z11-16:OH to Z11-16:Ald. As result, the sexy plant will be able to produce three sex pheromones involved in moth’s mating disruption (Figure 1).

Figure 1. Biosynthetic pathway of moth sex pheromones. Sex pheromones are bordered in purple.

Biosafety is a major concern in our project. For that reason, we have developed a biosafety module which allows an easy identification of the plant by identity preservation and prevents the plant to spread its genetic material via pollen.

In addition, we decided to enable external control of pheromone production, designing a genetic switch that controls the activation of the biosynthetic pathway. This genetic switch turns on the production of pheromones when a solution containing CuSO4 is sprayed on the plant.

The different genetic modules are put together to create the complete system. The system resembles an electric circuit, as it is represented in figure 2. Our genetic circuit, once introduced in Nicotiana benthamiana creates the Sexy Plant, a plant to fight moths and avoid damages in crops.

Figure 2. Our system in an electric circuit-like format. The plant’s own metabolism is used as the energy source. The biosafety module, composed by identity preservation and male sterility parts is always under expression in the system, to ensure the security of the transgenic plant. Pheromone synthetic pathways (A,B or C, depending on the problem pest) are activated using a genetic switch sensitive to Copper Sulfate, so system will not be producing pheromones unless it is activated with Copper-rich solution.

← Go to Home Go to Modules Go to Results →

REFERENCES

Ding BJ, Hofvander P, Wang HL, Durrett TP, Stymne S, et al. (2014) A plant factory for moth pheromone production. Nat Commun 5: 3353.
Hagström A, Wang HL, Lienard MA, Lassance JM, Johansson T, et al. (2013) A moth pheromone brewery: production of (Z)-11-hexadecenol by heterologous co-expression of two biosynthetic genes from a noctuid moth in a yeast cell factory. Microb Cell Fact 12: 125.

@@ Line 4: / Line 4: @@
 <div align="center"><div id="cn-box" align="justify">
-<p><h3 class="hook" align="left"><a>Project</a> > <a>Project Overview</a></h3></p><br/>
+<p><h3 class="hook" align="left"><a>Project</a> > <a>Overview</a></h3></p><br/>
 <div align="center"><span class="coda"><roja>P</roja>roject <roja>O</roja>verview</span> </div><br/><br/>
-<p>Pests cause great economical loses in agriculture and impedes an optimal use of the resources. Treatments used nowadays have some not-so-wanted faces: Pesticides do not target specifically enough to pests and cause alterations in other organisms, and chemical production of sex pheromones is expensive and generates contaminant by-products.</p><br/>
+<p>Pests cause <a href="https://2014.igem.org/Team:Valencia_UPV/Project/modules/problem" class="normal-link-page">great economical loss</a> in agriculture and impede an optimal use of the resources. <span class="red-bold">Sexy Plant</span> rises as a pest control strategy based on the use of <a href="https://2014.igem.org/Team:Valencia_UPV/Project/modules/moths_behavior" class="normal-link-page">mating disruption</a> by sex pheromones. The release of sex pheromones impedes male moths to find their females, avoiding crop-damaging larvae to be born.</p><br/>
-			<p><span class="black-bold">Sexy Plant</span> rises as a pest control strategy based on the use of mating disruption to prevent laying. <span class="black-bold">Mating disruption</span> is based on the release of sex pheromones to the air, so male’s sex pheromones receptors are not able to trace females anymore. This strategy is effectively used in field already with chemically synthesised pheromones [1]. Our final goal is to increase the sustainability of this approach by intercropping few Sexy Plants emitting mating-disrupting pheromones.</p><br/>
+			<p>The <span class="red-bold">Sexy Plant</span> is engineered to <a href="https://2014.igem.org/Team:Valencia_UPV/Project/modules/biosynthesis" class="normal-link-page">produce</a> and <a href="https://2014.igem.org/Team:Valencia_UPV/Project/modules/release" class="normal-link-page">release</a> three moth sex pheromones which affect a broad number of Lepidoptera species (moths). We implemented previous work by Ding et al. [1] to produce the first two pheromones (<span class="red-bold">Z11-16:OH</span> and <span class="blue-bold">Z11-16:OAc</span>). Inspired by Hagström et al [2], we planned the introduction of a Fatty Acid reductase to transform <span class="red-bold">Z11-16:OH</span> to <span class="green-bold">Z11-16:Ald</span>. As result, the sexy plant will be able to produce three sex pheromones involved in moth’s mating disruption (Figure 1).</p><br/>
-			<p>Sexy Plant produces and releases in <span class="italic">Nicotiana benthamiana</span> several <span class="black-bold">sex pheromones</span> (Z11-16:OH, Z11-16:OAc and Z11-16:Ald) for a broad number of Lepidoptera species (moths). We implemented previous work on production pathways done by Ding et al. [2] into our system to produce the first two pheromones (Z11-16:OH and Z11-16:OAc). We knew about the importance of Z11-16:Aldehide to control a large number of species, so inspired by Hagström approach [3], we introduced Fatty Acid reductase to transform Z11-16:OH to Z11-16:Ald. As result, the sexy plant is able to produce three sex pheromones involved in moth’s mating disruption with 2 or 3 additional enzymes (Figure 1).</p><br/>
+<div align="center"><img src="https://static.igem.org/mediawiki/2014/4/43/VUPVOv_fig2.png" alt="pheromone_pathway" title="Pheromone Pathway" width="450px"></img></div><br/>
+			<div align="center"><p style="text-align: center; font-style: italic; font-size: 0.8em; width: 700px;"><span class="black-bold">Figure 1. Biosynthetic pathway of moth sex pheromones</span>. Sex pheromones are bordered in purple.</p></div><br/>
-<div align="center"><img src="https://static.igem.org/mediawiki/2014/d/de/Ivllobel_pathway.png" alt="pheromone_pathway" title="Pheromone Pathway"></img></div><br/>
-			<div align="justify"><p style="font-size: 0.8em; width: 700px;"><span class="black-bold">Figure 1. Biosynthetic pathway of moth sex pheromones</span>. Sex pheromones are bordered in purple. Taking palmitic acid CoA (16:CoA) as substrate, the expression of the genes AtrΔ11 (Desaturase) and HarFAR (Reductase) leads to the production of Z11-16:OH. By adding the gene EaDAcT (Acetyltransferase) in the previous system, production displaced to obtaining Z11-16:OAc. If a Fatty Acid Reductase (FAO) is included instead of EaDAcT, Z11-16:Ald will be produced. </p></div><br/>
+			<p>Biosafety is a major concern in our project. For that reason, we have developed a <a href="https://2014.igem.org/Team:Valencia_UPV/Project/modules/biosafety" class="normal-link-page">biosafety module</a> which allows an easy identification of the plant by identity preservation and prevents the plant to spread its genetic material via pollen.</p><br/>
-			<p align="left" class="black-bold">PHEROMONE BIOSYNTHESIS</p><br/>
+<div align="center"><img width="400px" src="https://static.igem.org/mediawiki/2014/6/64/VUPVOv_fig1.png"></img></div><br/>
-			<p>The first step in the creation of this synthetic plant is to define and construct the metabolic pathway of pheromone production.</p><br/>
-			<p>"Most moth species utilize Type I pheromones that consist of straight-chain compounds 10-18 carbons in length with a functional group of a primary alcohol, aldehyde, or acetate ester, and usually with several double bonds, de novo synthesized in the pheromone gland through modifications of fatty acid biosynthetic pathways" [1].</p><br/>
+			<p>In addition, we decided to enable external control of pheromone production, designing a <a href="https://2014.igem.org/Team:Valencia_UPV/Project/modules/switch" class="normal-link-page">genetic switch</a> that controls the activation of the biosynthetic pathway. This genetic switch turns on the production of pheromones when a solution containing CuSO4 is sprayed on the plant.</p><br/>
-			<p>Following the principle of modularity in synthetic biology, Hagström et al [2] produced the insect pheromone Z-11:160H in yeast from palmitic acid by introducing a delta-11 fatty-acyl desaturase and a fatty-acyl reductase from Agrotis segetum. Recently, Ding et al [3] had already done the same work taking into account that palmitate is synthesised in the chloroplast, leading to successful results.</p><br/>
-			<p>In Ding's work they use a delta 11 desaturase mRNA from Amyelois transitella (accession number JX964774); HarFAR_KKYR, an improved version of HarFAR-3 fatty acid reductase from Helicoverpa armigera (accession number JF709978) with an almost five folds conversion rate, and 1,2-diacyl-sn-glycerol:acetyl-CoA acetyltransferase from Euonymus alatus (accession
-			number GU594061). With these three enzymes we would obtain two pheromones: Z-11:16OH and Z-11:16OAc.</p><br/>
+			<p>The different genetic modules are put together to create the complete system. The system resembles an electric circuit, as it is represented in figure 2. Our genetic circuit, once introduced in <i>Nicotiana benthamiana</i> creates the <span class="red-bold">Sexy Plant</span>, a plant to fight moths and avoid damages in crops.</p><br/>
-			<p>Moreover, inspired by Hagström's discussion we also thought about using an alcohol oxidase in order to try to obtain pheromone Z11-16:Ald from Z11-16:OH. According to the SolGenomics database, there is already a fatty-acid alcohol oxidase in Nicotiana benthamiana but there was no certainty that the enzyme would work in in vivo conditions.
-			On the other hand, we learned that Candida tropicalis had a fatty alcohol oxidase that catalysed the conversion from Z11-16:Ald to Z11-16:O [4,5,6,7]. (At the moment it is not clear from which organism will the fatty acid alcohol oxidase be obtained).</p><br/>
-			<p>At the same time we were planning which pheromones could we produce, we also took into account on which insects will they have effect. We did a thorough search in the Pherobase database (http://www.pherobase.com) and we checked which insect met the conditions of having one of those as a major pheromone component and being a plague.</p><br/>
+<div align="center"><img width="400px" src="https://static.igem.org/mediawiki/2014/a/ac/VUPVOverview_figure_2.png" alt="circuit"></img></div><br/>
-			<div align="center"><img src="https://static.igem.org/mediawiki/2014/d/de/Ivllobel_pathway.png" alt="pheromone_pathway" title="Pheromone Pathway"></img></div><br/>
-			<div align="center"><p style="font-size: 0.8em; width: 70%;">Biosynthetic pathway of pheromone production. A continuous line shows the substrate and a discontinuous line shows the resulting pheromones.</p></div><br/>
+			<div align="center"><p style="text-align: justify; font-style: italic; font-size: 0.8em; width: 700px;"><span class="black-bold">Figure 2. Our system in an electric circuit-like format</span>. The plant’s own metabolism is used as the energy source. The biosafety module, composed by identity preservation and male sterility parts is always under expression in the system, to ensure the security of the transgenic plant. Pheromone synthetic pathways (A,B or C, depending on the problem pest) are activated using a genetic switch sensitive to Copper Sulfate, so system will not be producing pheromones unless it is activated with Copper-rich solution.</p></div><br/><br/><br/><br/><br/><br/>
-			<p align="left" class="black-bold">TRICHOMES</p><br/>
-			<p>Once the metabolic pathway needed to synthetize the desired pheromone has been properly described and characterized, a method that allows pheromone diffusion from the plant to the exterior environment must be implemented. With this objective in mind, we decided to express the enzymes required to produce the pheromones specifically in glandular trichomes, which are secretory structures naturally present in the plant.
-			For doing so, the enzyme coding sequence would be under the control of CSP2 promoter from Nicotiana tabacum [8], inducing their specific expression in trichomes and allowing the pheromones secretion.</p><br/>
-			<p align="left" class="black-bold">SWITCH</p><br/>
-			<p>The pheromone production would suppose an additional energy cost to the plant, therefore the objective is to activate this pathway and the subsequent pheromone production only when necessary, upon Lepidoptera mating season. The regulation mechanism chosen with that purpose has been the one naturally controlling the CUP1 metallothionein gene in S. cervisiae [9].
-			CUP1 function is to chelate copper ions to avoid its toxicity to the yeast; therefore it is only expressed under high copper concentration. This regulation mechanism relies on the expression of a transcription factor (ACE1 or CUP2) that changes its conformation under high copper concentrations allowing its interaction with a regulating region (Metal Responsive Element, MRE) in the promoter of the CUP1 gene.</p><br/>
-			<p>Taking as a reference this mechanism, Mett et al adapted it so it could be used in whole plants. Takanori Saijo and Akitsu Nagasawa [10] posteriorly improved its efficiency. We decided to take their work as an example and use a system in which the transcription factor CUP2 or ACE1 is expressed under the constitutive promoter P35S, and the enzymes responsible for the pheromone production are expressed under the regulation of a inducible promoter, composed of the MRE domain fused with the P35S minimal promoter.
-			Therefore, CUP2 transcription factor would only interact with MRE and the pheromone will be produced under high copper concentrations. A copper-rich solution will be applied to the plant by foliar spraying or soil drenching during Lepidoptera mating season allowing pheromone production on demand.</p><br/>
-			<p align="left" class="black-bold">BIOSAFETY</p><br/>
-			<p>One of the concerns about having transgenic plants in the field involves its uncontrolled spread. It is being developed a biosecurity module containing two parts: a male sterility submodule and an identity preservation submodule.</p><br/>
-			<p>The male sterility submodule is based on a barnase, an RNase gene from Bacillus amyloliquefaciens, specifically expressed in anthers under the regulation of the TA29 tapetum-specific promoter. As result, pollen from these plants will not be capable of fertilize plants.
-			The identity preservation submodule will express Rosea1, a transcription factor that activates anthocyanin synthesis pathways, leading to their accumulation, so that plants containing the biosafety module will be easily differentiated from non-transgenic plants.</p><br/>
-			<p align="left" class="black-bold">FINAL PLANT</p><br/>
-			<p>Finally, our purpose is to find an ideal plant which gathers the special characteristics to lay out the implementation of the plant as a real solution of the problem. This ideal plant is still to be determined, and will probably vary depending on the geographical situation of the crop.</p><br/><br/><br/><br/><br/>
 <div align="center">
-<a class="button-content-trans" id="goto-left" align="center"></a>
+<a class="button-content" id="goto-left" align="center" href="https://2014.igem.org/Team:Valencia_UPV"><strong>&larr; Go to Home</strong></a>
 <a class="button-content" id="goto-middle" align="center" href="https://2014.igem.org/Team:Valencia_UPV/Project/modules"><strong>Go to Modules</strong></a>
 <a class="button-content" id="goto-right" align="center" href="https://2014.igem.org/Team:Valencia_UPV/Project/results"><strong>Go to Results &rarr;</strong></a></div><br/><br/><br/><br/>
-			<p align="left" class="black-bold">REFERENCES</p><br/>
+			<p align="center" class="black-bold">REFERENCES</p><br/>
 				<div style="position: relative; left: 3%; width: 96%;"><ol>
-					<li>Matsumoto S (2010) Molecular mechanisms underlying sex pheromone production in moths. Biosci Biotechnol Biochem 74: 223-231.</li>
-					<li>Hagström A, Wang HL, Lienard MA, Lassance JM, Johansson T, et al. (2013) A moth pheromone brewery: production of (Z)-11-hexadecenol by heterologous co-expression of two biosynthetic genes from a noctuid moth in a yeast cell factory. Microb Cell Fact 12: 125.</li>
 					<li>Ding BJ, Hofvander P, Wang HL, Durrett TP, Stymne S, et al. (2014) A plant factory for moth pheromone production. Nat Commun 5: 3353.</li>
-					<li>Vanhanen S, West M, Kroon JT, Lindner N, Casey J, Cheng Q et al (2000) A consensus sequence for long-chain fatty-acid alcohol oxidases from Candida identifies a family of genes involved in lipid omega-oxidation in yeast with homologues in plants and bacteria. J Biol Chem. 275, 4445-52.</li>
+					<li>Hagström A, Wang HL, Lienard MA, Lassance JM, Johansson T, et al. (2013) A moth pheromone brewery: production of (Z)-11-hexadecenol by heterologous co-expression of two biosynthetic genes from a noctuid moth in a yeast cell factory. Microb Cell Fact 12: 125.</li>
-					<li>Eirich LD, Craft DL, Steinberg L, Asif A, Eschenfeldt WH, et al. (2004) Cloning and characterization of three fatty alcohol oxidase genes from Candida tropicalis strain ATCC 20336. Appl Environ Microbiol 70: 4872-4879.</li>
-					<li>Kemp GD, Dickinson FM, Ratledge C (1991) Activity and substrate specificity of the fatty alcohol oxidase of Candida tropicalis in organic solvents. Appl Microbiol Biotechnol. 34(4), 441-445.</li>
-					<li>Kemp GD, Dickinson FM, Ratledge (1988) Inducible long chain alcohol oxidase from alkane-grown Candida tropicalis. Appl Microbiol Biotechnol. 29(4), 370-374.</li>
-					<li>Sallaud C, Giacalone C, Topfer R, Goepfert S, Bakaher N, et al. (2012) Characterization of two genes for the biosynthesis of the labdane diterpene Z-abienol in tobacco (Nicotiana tabacum) glandular trichomes. Plant J 72: 1-17.</li>
-					<li>Mett VL, Lochhead LP, Reynolds PH (1993) Copper-controllable gene expression system for whole plants. Proc Natl Acad Sci U S A 90: 4567-4571.</li>
 				</ol></div>
 </br></br></div>