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Team:Debrecen Hungary/Project
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Our team was interested at designing eukaryotic synthetic biology tools related to PoPs. PoPs (polymerase per second), the flying Dutchman of synthetic biology, is a number which represents the rate (base pair per second) at which RNA polymerase crosses past a given DNA position. Currently, no in vivo technique for measuring PoPS directly exists; it can be estimated indirectly by measuring other parameters (eg protein expression or enzyme activity). Nevertheless it is still a useful abstraction for thinking about transcription-based logic devices and it allows the engineer to define devices. Our aim, was not only to infer PoPs but to devise a way to titrate it remotely. | Our team was interested at designing eukaryotic synthetic biology tools related to PoPs. PoPs (polymerase per second), the flying Dutchman of synthetic biology, is a number which represents the rate (base pair per second) at which RNA polymerase crosses past a given DNA position. Currently, no in vivo technique for measuring PoPS directly exists; it can be estimated indirectly by measuring other parameters (eg protein expression or enzyme activity). Nevertheless it is still a useful abstraction for thinking about transcription-based logic devices and it allows the engineer to define devices. Our aim, was not only to infer PoPs but to devise a way to titrate it remotely. | ||
| - | === | + | === Nuclear Hormone Receptors === |
| + | Nuclear receptors are ligand activated transcription factors. As such, they are able to regulate the expression of their target genes by direct DNA-binding, in a ligand-dependent manner. They play a central role in endocrine signaling, regulation of embryonic and adult development and differentiation. Many nuclear receptors, are among the primary targets of drug discovery because of their diverse biological actions. Nuclear receptors bear high homology to each other and are modular into distinct domains: N-terminal regulatory domain, DNA-binding domain, a Hinge region, Ligand binding domain (LBD) and a C-terminal domain. | ||
| + | Ligand binding domain (LBD) is a well conserved domain amongst various nuclear receptors whose structure usually referred to as an alpha helical sandwich fold. The LBD shows some diversity among nuclear receptors as it is a site for receptor-specific events. It possesses transactivation ability and contains a ligand-binding pocket as well as the main interaction surfaces for other proteins. | ||
| + | The DNA-binding domain (DBD) contains two Zinc-finger motifs and is linked to the LBD by a highly flexible hinge region. This segment of the nuclear receptors holds the ability to recognize and bind to preferred or specific DNA-motifs, the response elements. | ||
| + | A partially unexpected and amazing feature of the nuclear receptors is that the above mentioned domains can be swapped. It was Vincent Giguere and his colleagues, who first devised a technique called cotransfection assay and developed it to the domain-swap technique. | ||
| - | === | + | === Apoptosis === |
| + | Apoptosis, the process of programmed cell death (PCD), is an elaborate cellular homeostasis mechanism that ensures correct development and function of multicellular organisms. Biochemical events lead to characteristic changes (morphology) and the death of cells. | ||
| + | It has a crucial role in maintaining tissue homeostasis as provides the appropriate rate of cellular turnover. Potentially dangerous, poorly differentiated or excess cells are removed by apoptosis without local inflammation from leakage of cell contents. Dying by apoptosis in many cases has primary importance: lens cells, which lost their nucleus, need the vision, the corneocytes without nucleus and organelles provide mechanical protection on the outer surface of the skin. | ||
| + | Apoptosis might occur in pathological conditions, mainly in secondary, as a part of the defensive mechanism, helping to eliminate the viral infected or malignant cells. The abnormality of the central molecules and genes of apoptosis may play a role in the development of certain diseases: degenerative disease, autoimmune disease, tumor formation and malformations. | ||
| - | + | Many pathways and signals lead to apoptosis, but there is only one mechanism that actually causes the death of a cell. After a cell receives stimulus, it undergoes organized degradation of cellular organelles by activated proteolytic caspases. A cell undergoing apoptosis shows characteristic morphology. | |
| + | === The Experiments === | ||
| + | |||
| + | |||
| + | === Part 3 === | ||
== Results == | == Results == | ||
Revision as of 16:02, 5 September 2011
| You can write a background of your team here. Give us a background of your team, the members, etc. Or tell us more about something of your choosing. | |
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Nuclear hormone receptors (NHRs) are ligand activated transcription factors. They are able to regulate the expression of their target genes by direct DNA-binding, in a ligand-dependent manner. NHRs bear high homology to each other and are modular into distinct regions: N-terminal regulatory region, DNA-binding domain, a Hinge region, Ligand binding domain (LBD) and a C-terminal region. Some Nematode NHRs can be activated by extracted oil contamination of the soil so they can use as possible oil sensors in the future. Zinc finger motifs are the tools of NHRs to bind DNA and regulate gene expression directly. These tiny elements can be tested as direct gene regulators. Controlled gene induction can also lead to programmed cell death which is a less harmful tool in order to quit non-functionable cells. | File:Debrecen Hungary team.png Your team picture |
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Contents |
Abstract
Nuclear hormone receptors (NHRs) are ligand activated transcription factors. They are able to regulate the expression of their target genes by direct DNA-binding, in a ligand-dependent manner. NHRs bear high homology to each other and are modular into distinct regions: N-terminal regulatory region, DNA-binding domain, a Hinge region, Ligand binding domain (LBD) and a C-terminal region. Some Nematode NHRs can be activated by extracted oil contamination of the soil so they can use as possible oil sensors in the future. Zinc finger motifs are the tools of NHRs to bind DNA and regulate gene expression directly. These tiny elements can be tested as direct gene regulators. Controlled gene induction can also lead to programmed cell death which is a less harmful tool in order to quit non-functionable cells.
Introduction
Synthetic Biology
Synthetic biology is a relatively new area of biological research; similar to many other new scientific fields it has many definitions. The best way to express the meaning of synthetic biology is to understand the desired end-result: engineering of complex biological systems. These systems are best thought of as analogues of everyday machinery: cogwheels, levers, timers, button and buzzers (in this case a clock), only in the case of biological systems (molecular ones) cells, DNA, proteins, lipids, sugars and RNA are the “parts” of the system.
Similar to mechanical engineering (or every other engineering branch) there is a need for standards (consensus way of doing things), abstraction (simple and unified way of thinking about the parts of a system), and modularity (how these parts interact to become the device, or several devices into a system). Thus a good definition for synthetic biology could be engineering of molecular (for the time being) biological systems according to preset standard parts. The international genetically engineered machines and associated parts registry are, to date, one of the largest registries for standard parts in use for synthetic biology. Free information can be found in the registry regarding parts, devices and modules all inputted by various teams worldwide.
Eukaryotic synthetic biology is still in its infancy. The large kingdom of metazoans includes all multicellular eukaryotes such as mammalians, arthropods and nematodes. No standard chassis (framework) exists for the animal kingdom which makes them far less popular then the famous E.coli. The protein modules derived from metazoans (like Drosophila or C. elegans) are functional in yeasts also. Very few iGEM teams (or even labs outside iGEM) have chosen to toggle the animal chassis (two notable examples are team Heidelberg 2009 and team Slovenia 2006). The amount of available compatible parts is limited, which severely restricts the options of creating complex biological devices. Nearly no imagination is required for designing tools, since their analogues already exist in the bacterial chassis. The possible use of such systems is unlimited. Field’s such as of environment, medicine, energy and research all gain to profit from the development of animal synthetic biology.
Systems requiring gene expression input in eukaryotic synthetic biology systems require a way to standardize gene expression, a complicated task. The way from gene to protein contains many steps of possible error: transcription factor binding, promoter strength, recruitment of auxiliary proteins, nuclear RNA synthesis and many more steps finally leading to translation, folding, cleaving and delivery (but hey, you have to start somewhere).
Our team was interested at designing eukaryotic synthetic biology tools related to PoPs. PoPs (polymerase per second), the flying Dutchman of synthetic biology, is a number which represents the rate (base pair per second) at which RNA polymerase crosses past a given DNA position. Currently, no in vivo technique for measuring PoPS directly exists; it can be estimated indirectly by measuring other parameters (eg protein expression or enzyme activity). Nevertheless it is still a useful abstraction for thinking about transcription-based logic devices and it allows the engineer to define devices. Our aim, was not only to infer PoPs but to devise a way to titrate it remotely.
Nuclear Hormone Receptors
Nuclear receptors are ligand activated transcription factors. As such, they are able to regulate the expression of their target genes by direct DNA-binding, in a ligand-dependent manner. They play a central role in endocrine signaling, regulation of embryonic and adult development and differentiation. Many nuclear receptors, are among the primary targets of drug discovery because of their diverse biological actions. Nuclear receptors bear high homology to each other and are modular into distinct domains: N-terminal regulatory domain, DNA-binding domain, a Hinge region, Ligand binding domain (LBD) and a C-terminal domain.
Ligand binding domain (LBD) is a well conserved domain amongst various nuclear receptors whose structure usually referred to as an alpha helical sandwich fold. The LBD shows some diversity among nuclear receptors as it is a site for receptor-specific events. It possesses transactivation ability and contains a ligand-binding pocket as well as the main interaction surfaces for other proteins.
The DNA-binding domain (DBD) contains two Zinc-finger motifs and is linked to the LBD by a highly flexible hinge region. This segment of the nuclear receptors holds the ability to recognize and bind to preferred or specific DNA-motifs, the response elements.
A partially unexpected and amazing feature of the nuclear receptors is that the above mentioned domains can be swapped. It was Vincent Giguere and his colleagues, who first devised a technique called cotransfection assay and developed it to the domain-swap technique.
Apoptosis
Apoptosis, the process of programmed cell death (PCD), is an elaborate cellular homeostasis mechanism that ensures correct development and function of multicellular organisms. Biochemical events lead to characteristic changes (morphology) and the death of cells.
It has a crucial role in maintaining tissue homeostasis as provides the appropriate rate of cellular turnover. Potentially dangerous, poorly differentiated or excess cells are removed by apoptosis without local inflammation from leakage of cell contents. Dying by apoptosis in many cases has primary importance: lens cells, which lost their nucleus, need the vision, the corneocytes without nucleus and organelles provide mechanical protection on the outer surface of the skin.
Apoptosis might occur in pathological conditions, mainly in secondary, as a part of the defensive mechanism, helping to eliminate the viral infected or malignant cells. The abnormality of the central molecules and genes of apoptosis may play a role in the development of certain diseases: degenerative disease, autoimmune disease, tumor formation and malformations.
Many pathways and signals lead to apoptosis, but there is only one mechanism that actually causes the death of a cell. After a cell receives stimulus, it undergoes organized degradation of cellular organelles by activated proteolytic caspases. A cell undergoing apoptosis shows characteristic morphology.
