Writing in the journal Cell, three Boston University biomedical engineers report they have created a genetic dimmer switch that can be used to turn on, shut off, or partially activate a gene's function. The switch - developed by James Collins, Charles Cantor and Tara Deans - could help advance the field of synthetic biology, which rests on the premise that complex biological systems can be "constructed" by arranging an array of different biological components. "There are a number of technologies available to regulate gene expression, but they each come with limitations," explained Collins. "One of the central problems is you can't get a really tight 'off' state." Even when genetic switches are turned off, a trickle of the protein that is meant to be repressed still gets made. Some genetic switches get around this by entirely snipping out a gene to stop production of a specific protein, but this approach is irreversible.
To overcome these challenges, Deans came up with a design that combined two different technologies to repress or shut down gene expression. The first method, a repressor protein, sits on DNA like a roadblock, preventing any messenger RNA (mRNA) from being made. If any mRNA gets past this repressor, the second technique, interfering RNA (RNAi) attaches to the functional mRNA, rendering it useless. "I was delighted to see that when the two systems are coupled, it is possible to completely turn a gene's function off," said Deans.
Importantly, the switch is also reversible and tunable. By adding a chemical - Isopropyl-â-thiogalactopyranoside - the repressor components are blocked and the gene turns on again. The gene's activity can be tuned up or down by adjusting the amount of this chemical.
The researchers demonstrated the strength of their "off" switch by hooking it up to the gene for diphtheria toxin, then inserting it into cells. One molecule of diphtheria toxin can kill a cell, but with the genetic switch turned off, the cells survived for weeks. When the researchers flipped the switch, toxin production was triggered and the cell died.
They also showcased the switch's capability for delicately tuning gene expression by installing it alongside a gene that leads to apoptosis (programmed cell death) once a certain threshold concentration of the gene's product is reached. They gradually increased the gene's activity until they met and passed this threshold. "This tuning feature is important," said Deans, because "many diseases are not a result of missing a gene, but rather a result of too much or too little expression. With the ability to tune the level of gene expression in our switch, we could explore threshold responses and how these result in disease phenotypes."
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Source: Boston University
