Latest revision as of 00:52, 27 October 2007

SMB is composed of a germline and a soma. Germ cells are responsible for reproduction while the sterile somatic cells, are essential for the germline as the soma exports the DAP compound, supporting germline growth. The special feature of our system, the coexistence of two independent cell types, one dedicated to reproduction and the other sterile, makes it a potentially interesting tool for synthetic biology.

The SMB could also be used to explore basic questions in fundamental biology. If submitted to long time growth, in a chemostat for instance, the SMB could become a model to study how the genome of a cell can evolve leading to changes that do not directly affect the cells phenotype. This system could mimic evolutionary phenomena that are typicall of multicellular organisms.

System improvement through directed evolution

The engineered SMB may be further tuned by laboratory evolution. A key advantage of the system is that selection pressure could be tuned to either force faster reproduction (e.g., improving DAP production and/or export from the soma or alternatively for a more efficient use of DAP within the germline) or for improvement of soma-specific functions (see below). In other words, the soma-germline SMB dichotomy provides us with the possibility to select for functions that are otherwise deleterious for the unicellular organism.

Applications

We envision two immediate applications to the Synthetic Multicellular Bacterium. It could be used as a “metabolic plant” or as “security device”

Decoupling reproduction faculty and transgene expression

In the SMB system, any given gene or gene circuit can be selectively expressed in germ cells or somatic cells. Knowing the soma is unreplicative, we are disrupting the connection between the reproduction of the system and the expression of the transgene, leading to potentially interesting applications.

We would like to develop two extensions to the SMB : metabolic engineering and secured genetically modified organisms for environmental release.

Security device

We came up with the idea of modifying the synthetic organism in order to allow, on demand, the full differentiation of the germline into soma in a secured fashion. Why would we want to do that? This possibility is to be considered in the face of biohazard risks. Release of GM organisms in the environment poses ethical as well as technical questions. “Biological security” should be a prime concern in synthetic biology.

If a task is to be transiently performed by a GM organism in the open environment, a major problem is persistence of the GM organism, with potential risks of proliferation and transgene dissemination. A possible solution, already used for some GM crops, is to render the organisms sterile. Can this solution be implemented for GM bacteria, in bioremediation systems for instance? Using a modified version of our SMB, the answer is yes. The SMB comprises two cell types, one being the soma, a group of cells that have a longer lifetime than wild type E.coli cells & are unable to replicate. The soma cells have the required characteristics for transient use in open environment.

The next consideration is then: can full differentiation be induced in the SMB? In order to achieve this, two events should be initiated:

Induction of a massive differentiation of the germ line into soma cell
Followed by selective death of the persistent germ line cells.

For details on proposed technical solutions to this challenge, SEE HERE

Metabolic engineering

A bacterial multicellular organism could allow optimizing the production of compounds deleterious to the cell. If one attempts to optimize the production of such a compound in a classical bacteria (E.coli for instance), trade-off appears between the production of your compound and the growth of the cell. The synthetic multicellular organism could in part bypass this problem by partially decoupling growth from synthesis of the exogenous compound. It could indeed be modified so that only the soma produces the noxious molecule. If a mean exists to screen for maximum production of this molecule, you could then select the germline whose soma would have the best exocompound production yield. As long as the production doesn't impair too much the capacity of the soma to feed the germline, the optimization can go on. Thus, a trade-off also exists in this case, but it would very well less stringent that in the wild type E.coli case.

Of course, all this is only possible if the germ line is not affected by the products released by the soma. Compounds that could be optimized in this way are thus constrained to molecules that can be noxious if accumulated in the cell, but that do not affect it too much if in the medium. For instance, if the product can be gradually filtered out of the medium.

E. Colight: towards a new slim diet

Project

In an attempt to integrate all the proposed applications of the SMB, we advanced towards the construction of Ecolight. Bringing together the synthetic organism (SMB), noxious compound synthesis ("metabolic plant"), and the security device. This would include optimizing the production of triglycerides in the soma cells of our synthetic multicellular bacterium. Soma isolation according to the "security device" would be included. The super triglycerides-producing-not-able-to-divide soma cells could then be ingested. The fatty acids they would store would be as many fatty acids you will not absorb! Eat fat, don't get fat!

E.colight project is also made up with 2 independent parts. First we have to create the SMB to implement the secured module. Second E.coli has to absorb free fatty acid and to synthesize triglycerides in the cytoplasm.

References:
- 2006 Physicians' Desk Reference (PDR). Thomson PDR
- BBa_I718002

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@@ Line 2: / Line 2: @@
-== System improvement through directed evolution ==
+SMB is composed of a germline and a soma. Germ cells are responsible for reproduction while the sterile somatic cells, are essential for the germline as the soma exports the DAP compound, supporting germline growth. The special feature of our system, the coexistence of two independent cell types, one dedicated to reproduction and the other sterile, makes it a potentially interesting tool for synthetic biology.
-==Applications==
+The SMB could also be used to explore basic questions in fundamental biology. If submitted to long time growth, in a chemostat for instance,  the SMB could become a model to study how the genome of a cell can evolve leading to changes that do not directly affect the cells phenotype. This system could mimic evolutionary phenomena that are typicall of multicellular organisms.
-===Metabolic engineering===
-A bacterial multicellular organism could allow optimizing the production of compounds deleterious to the cell. If you try to optimize the production of such a compound in a classical bacteria (E.coli for instance), you will reach a trade off between the production of your compound and the growth of the cell because the reproduction capacity is linked to the expression of the toxic transgene. The synthetic organism could in part bypass this problem by partially decoupling growth from synthesis of the exogenous compound. It could indeed be modified so that the soma only will produce the noxious molecule and the germ line will multiply. If there is a way to screen for the production of this molecule, we can then select the germ line whose soma would have the best yield. As long as the production doesn't impair too much the capacity of the soma to feed the germ line, the optimization can go on. Thus, there is also a trade off in this case, but it might very well be more favorable to the optimization than the trade off between production and growth. At least, this is worth testing.
+= System improvement through directed evolution =
-Of course, all this is only possible if the germ line is not affected by the production of the soma. Compounds that could be optimized in this way are thus constrained to molecules that can be noxious in the cell, but that do not affect it to much if in the medium.
+The engineered SMB may be further tuned by laboratory evolution. A key advantage of the system is that selection pressure could be tuned to either force faster reproduction (e.g., improving DAP production and/or export from the soma or alternatively for a more efficient use of DAP within the germline) or for improvement of soma-specific functions (see below). In other words, the soma-germline SMB dichotomy provides us with the possibility to select for functions that are otherwise deleterious for the unicellular organism.
+=Applications=
-''' E. Colight: optimization of triglycerides synthesis in the soma of the synthetic organism: towards a new slim diet '''
+We envision two immediate applications to the Synthetic Multicellular Bacterium. It could be used as a “metabolic plant” or as “security device”
-Finally, the last part of our project would be to bring together the synthetic organism, the optimization of a compound noxious to the bacteria, and the security device. To do so, we'd like to optimize the production of triglycerides in the soma cells of our synthetic organism. We would then be able to differentiate all the germ line of the synthetic organism into soma in a secured way. Those super triglycerides-producing-not-able-to-divide cells could then be ingested. The fatty acids they would stock would be as many fatty acids you will not absorb! Eat fat, don't get fat !
-Triglycerides are made of a molecule of glycerol esterified by three fatty acids. When ingested they are hydrolysed by lipase in the stomach and the duodenum, into glycerol and free fatty acids. Enterocytes are only able to absorb free fatty acids and glycerol. So they combined them in the cytoplasm into triglycerides that will be free in the lymphatic system then in the blood within fatty vesicles called chylomicrons.
+== Decoupling reproduction faculty and transgene expression ==
-When we are becoming fat, it is because we have more input than output energy, lipid energy. Knowing that gut is full with bacteria forming the gut microflora (we have 1013 of our cells in our body and 1014 bacteria with the majority in the gut!), we could imaging having a somatic cell bacteria able to absorb fatty acids and form triglycerides intracellular inclusion. These triglycerides are not able to be absorbed by enterocytes! Input lipid is decreased! Eat fat, don't get fat! E.coli is the most used bacterium in synthetic biology and... belongs to the gut microflora! We could also imagine genetically engineering E.coli to stock triglycerides into inclusions! Knowing that 40% of E.coli are renewed every day, these triglyceride-fulled bacteria will leave the gut with feces! With the security device described above we should have a safe GMO with two hours life expectancy!
+In the SMB system, any given gene or gene circuit can be selectively expressed in germ cells or somatic cells. Knowing the soma is unreplicative, we are disrupting the connection between the reproduction of the system and the expression of the transgene, leading to potentially interesting applications.
+<br><br>
+We would like to develop two extensions to the SMB : metabolic engineering and secured genetically modified organisms for environmental release.
 ===Security device===
-'''secure isolation of the soma => release in the environment'''
+We came up with the idea of modifying the synthetic organism in order to allow, on demand, the full differentiation of the germline into soma in a secured fashion. Why would we want to do that?
+This possibility is to be considered in the face of biohazard risks. Release of GM organisms in the environment poses ethical as well as technical questions. “Biological security” should be a prime concern in synthetic biology.
+If a task is to be transiently performed by a GM organism in the open environment, a major problem is persistence of the GM organism, with potential risks of proliferation and transgene dissemination. A possible solution, already used for some GM crops, is to render the organisms sterile. Can this solution be implemented for GM bacteria, in bioremediation systems for instance? Using a modified version of our SMB, the answer is yes. The SMB comprises two cell types, one being the soma, a group of cells that have a longer lifetime than wild type E.coli cells & are unable to replicate. The soma cells have the required characteristics for transient use in open environment.
+The next consideration is then: can full differentiation be induced in the SMB? In order to achieve this, two events should be initiated:
+* Induction of a massive differentiation of the germ line into soma cell
+* Followed by selective death of the persistent germ line cells.
+For details on proposed technical solutions to this challenge, [[Security device|'''SEE HERE''']]
+===Metabolic engineering===
+A bacterial multicellular organism could allow optimizing the production of compounds deleterious to the cell. If one attempts to optimize the production of such a compound in a classical bacteria (E.coli for instance), trade-off appears between the production of your compound and the growth of the cell. The synthetic multicellular organism could in part bypass this problem by partially decoupling growth from synthesis of the exogenous compound. It could indeed be modified so that only the soma produces the noxious molecule. If a mean exists to screen for maximum production of this molecule, you could then select the germline whose soma would have the best exocompound production yield. As long as the production doesn't impair too much the capacity of the soma to feed the germline, the optimization can go on. Thus, a trade-off also exists in this case, but it would very well less stringent that in the wild type E.coli case.
+Of course, all this is only possible if the germ line is not affected by the products released by the soma. Compounds that could be optimized in this way are thus constrained to molecules that can be noxious if accumulated in the cell, but that do not affect it too much if in the medium. For instance, if the product can be gradually filtered out of the medium.
+== E. Colight: towards a new slim diet ==
+=== Project ===
+In an attempt to integrate all the proposed applications of the SMB, we advanced towards the construction of Ecolight.  Bringing together the synthetic organism (SMB), noxious compound synthesis ("metabolic plant"), and the security device.  This would include optimizing the production of triglycerides in the soma cells of our synthetic multicellular bacterium. Soma isolation according to the "security device" would be included. The super triglycerides-producing-not-able-to-divide soma cells could then be ingested. The fatty acids they would store would be as many fatty acids you will not absorb! Eat fat, don't get fat! <br>
+E.colight project is also made up with 2 independent parts. First we have to create the SMB to implement the secured module. Second E.coli has to absorb free fatty acid and to synthesize triglycerides in the cytoplasm.
+[[Image: securitydevice4.jpg|center|600px]]
+'''References:''' <br>
+- 2006 Physicians' Desk Reference (PDR). Thomson PDR <br>
+- <bbpart>BBa_I718002</bbpart>
-We also came up with the idea of modifying the synthetic organism so that we would be able to induce the differentiation of all the germ line into soma in a secured way. Why would we want to do that ? Well, in a security perspective, we may not want to release a transgenic strain in the environment. But it would be acceptable to do so if we are sure that the cells we release are not able to proliferate. In this view, we could release in the environment, what would correspond to our "soma". Those cells aren't able to divide and their effect is limited in time since they will die after approximatively 2h. Indeed, one of the problems with GMO is the selective advantage that the gene gives. If it is greater than for the wild type, the GMO will spread and replace the wild type in the ecosystem. For example, if we think about the gut flora, a genetic engineered E.coli could spread and replace the wild type E.coli causing diseases (diarrhoea...). That could be possible because the reproductive faculty is linked to the gene given: the gene gives a better reproductive capacity of the GMO. One way to avoid this problem is decoupling the reproduction faculty and the expression of the gene: the organism is either able to multiply or able to express the gene. So expressing the transgene can not be responsible of a selective advantage against the wild type.
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From 2007.igem.org