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Team:SYSU-China/Project/Design
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The introduction of each genes about its suicide mechanisms and the reasons why we choose them are as follows: | The introduction of each genes about its suicide mechanisms and the reasons why we choose them are as follows: | ||
</p> | </p> | ||
| + | <h2>Which suicide gene?</h2> | ||
| + | |||
| + | <h3>hBax & hBax mutant</h3> | ||
| + | <p> | ||
| + | hBax is a member of the Bcl-2 related protein family from human. This Family contains pro-apoptotic and anti-apoptotic proteins, and the balance among them determines the cell survival. hBax is the pro-apoptotic protein. During apoptosis, hBax will be inserted into the mitochondrial outer membrane and form permeable channels, release pro-apoptotic signals, finally lead to apoptosis<a class="quote">[1]</a>. The pathway of apoptosis is showed in Figure 1. | ||
| + | </p> | ||
| + | <img src="https://static.igem.org/mediawiki/2013/2/2f/Which_suicide_gene_fig_1.gif" width="500"/> | ||
| + | Figure1. apoptosis pathway induced by hBax | ||
| + | <p> | ||
| + | hBax S184a<a class="quote">[3]</a> is a mutant of hBax that can constantly insert into mitochondrial outer membrane, thus we guess that it may have a stronger apoptosis-induced effect than normal hBax. | ||
| + | </p> | ||
| + | <p> | ||
| + | It have been reported that over expression of this gene can successfully induce apoptosis in both Hela cell line and HEK-293 cell line<a class="quote">[2]</a>. Due to its generality, we finally determined to use it as one of our suicide genes candidates. | ||
| + | </p> | ||
| + | <p> | ||
| + | However, our experiments results finally showed to us that, when expressed in Hep G2 cell line, which is a typical hepatoma cell line, they can not induce observable apoptosis. However, although it can not successfully kill Hep G2 cell line, we figured out that the pathway is conserved in yeast[4] ,so we also tried to transfect the gene and its mutant in yeast and finally proved that the killing effect is observable and even dose-dependent (<A HREF="http://parts.igem.org/Part:BBa_K1061006">BBa_K1061006</A>). So this extra result may be useful for other team in the future who want to apply safety device in yeast. We finally submitted the hBax as a biobrick and its mutant form as an improvement of the pre-existing part of part registry. | ||
| + | </p> | ||
| + | <p>(show results)</p> | ||
<DIV id="references"> | <DIV id="references"> | ||
Revision as of 08:41, 26 September 2013
ipsc
suicide gene?
Why suicide gene?
In our project, we try to build a circuit that can prevent the iPSC-differentiated tissues from tumor formation. Instead of using some medicines to kill them, like the traditional gene therapies did, what we want to achieve here is to make the tumor cells kill themselves, in other word, automatically commit suicide. So we decided to use an exogenous suicide gene.
The ideal suicide genes that fit our design here will be genes that can successfully induce apoptosis in cells and do not introduce any harmful effect to normal tissues. However, many apoptosis pathways are reported to be blocked in tumor cells. From recent reports and papers, we found out that there have been many genes reported to have important roles in mammalian cell apoptosis. After a long time comparison and consideration, our final candidates of suicide gene are listed below:
- hBax & hBax mutant
- delta TK
- caspase3
- rip1
- rip3
- apoptin
The introduction of each genes about its suicide mechanisms and the reasons why we choose them are as follows:
Which suicide gene?
hBax & hBax mutant
hBax is a member of the Bcl-2 related protein family from human. This Family contains pro-apoptotic and anti-apoptotic proteins, and the balance among them determines the cell survival. hBax is the pro-apoptotic protein. During apoptosis, hBax will be inserted into the mitochondrial outer membrane and form permeable channels, release pro-apoptotic signals, finally lead to apoptosis[1]. The pathway of apoptosis is showed in Figure 1.
Figure1. apoptosis pathway induced by hBax
hBax S184a[3] is a mutant of hBax that can constantly insert into mitochondrial outer membrane, thus we guess that it may have a stronger apoptosis-induced effect than normal hBax.
It have been reported that over expression of this gene can successfully induce apoptosis in both Hela cell line and HEK-293 cell line[2]. Due to its generality, we finally determined to use it as one of our suicide genes candidates.
However, our experiments results finally showed to us that, when expressed in Hep G2 cell line, which is a typical hepatoma cell line, they can not induce observable apoptosis. However, although it can not successfully kill Hep G2 cell line, we figured out that the pathway is conserved in yeast[4] ,so we also tried to transfect the gene and its mutant in yeast and finally proved that the killing effect is observable and even dose-dependent (BBa_K1061006). So this extra result may be useful for other team in the future who want to apply safety device in yeast. We finally submitted the hBax as a biobrick and its mutant form as an improvement of the pre-existing part of part registry.
(show results)
References
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[3]Amotz Nechushtan et al, Conformation of the Bax C-terminus regulates subcellular location and cell death, The EMBO Journal Vol.18 No.9 pp.2330–2341, 1999.
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[20]Lars Guelen et al,TAT-apoptin is efficiently delivered and induces apoptosis in cancer cells, Oncogene (2004) 23, 1153–1165
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Sun Yat-Sen University, Guangzhou, China
Address: 135# Xingang Rd.(W.), Haizhu Guangzhou, P.R.China
