Synthetic Biology in Mammals
From Dibernardo
Synthetic biology aims at building novel biological ‘circuits’ (synthetic network) in the cell able to perform specific tasks, or to change the behaviour of the biological process in a desired way. Biologists are familiar with manipulation of genes and proteins to probe their properties and understand biological processes. Synthetic biology must also manipulate the material elements of the cell, but for the purpose of design, to build synthetic biological systems. Recent advances have been made in Synthetic Biology but mainly in vitro and in microorganisms. Our research is aimed at building novel synthetic circuits that can be used in vivo in mammalian cells. We are working on two synthetic circuits that can be packed in a lentiviral vector and delivered in vivo: (1) the “toggle switch” and (2) the “oscillator”.
The toggle switch is a novel inducible system to be used in mammalian cells. Its main characteristic is the ability to permanently “switch on” or “off” a gene of interest without any external inducer. The toggle switch is based on an epigenetic mechanism where two genes inhibit each other. Specifically in our switch a shRNA is directed against a ttR-Krab transcriptional repressor that is able to regulate the shRNA transcription. A system of differential equations was used to describe the circuit and to aid determining the strength of promoters and degradation rates allowing the system to behave as a switch. Our work shows how it is possible to engineer epigenetic expression circuits for maintenance of a desired relative level of proteins in mammalian cells.
More complex behaviors, such as periodic expression of a gene independently of the cell-cycle (“oscillator”), are also possible. To date, such a synthetic network does not exist in mammalian cells. Our research aims at developing a mammalian oscillator to transcribe a gene of interest at specific time intervals from a population of engineered cells carrying the oscillator circuit in their genome. We aim at achieving this objective by using a transcriptional activator, ttA-VP16, regulating its own transcription and the transcription of a microRNA directed against the transcriptional activator. This circuit will thus contain a positive self-feedback loop (ttA-VP16 on itself) and negative feedback loop (microRNA against ttA-VP16). These characteristics are the essential features that according to mathematical models allow a stable oscillation in the transcription of both ttA-VP16 and the microRNA,
Both the toggle switch and the oscillator, in a gene therapy setting, could be used to regulate in-vivo the administration of a given gene product with a desired behaviour.
We coordinated the European Project COBIOS that aims at using Synthetic Biology for therapy
