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Project

The problem

Producing complex therapeutic proteins requires biosynthesis using mammalian cells to obtain the desired product. This product sometimes has some level of toxicity for the cells, limiting the productivity.

Our solution

We want to transfect a light switch along with the gene of interest. This means that the product will only be synthesized by the cells in the presence of blue light, allowing them to grow happily in the dark. The switch consists in an already existing chimeric protein, LovTAP. This protein was originally intended to act as a light-induced repressor in bacteria. The EPFL 2009 iGEM team proposed to fuse it with VP16, a viral activator, in order to convert it into a light-induced activator in mammalian cells. This year we will try to realize this idea by transfecting CHO (Chinese hamster ovary) cells with two plasmids: one coding for the LovTAP-VP16 fusion protein and another with a read-out protein preceded by a cleavage site for LovTAP-VP16. If everything goes as expected, LovTAP-VP16 will only bind the plasmid and activate the production of read-out when exposed to light.

Plan B

In addition, we will be realizing the light switch described by Fussenegger et al. In this light switch, light sensitive melanopsin triggers a cascade involving calcium ion channels that eventually leads to the transcription of the gene of interest. Fussenegger's team did this using HEK (human embryo kidney) cells, and we will try to make it work also on CHO cells.