A Yeast Optogenetic Toolkit for Control of Intra- and Intercellular Signaling

Optogenetic proteins "light sensitive proteins that change activity in their photoexcited state” make it possible to harness the exquisite spatiotemporal control available with light to actuate cellular activities. We describe a yeast optogenetic toolkit (yOTK) that allows for rapid assembly of optically controlled biological circuits within the model organism Saccharomyces cerevisiae.

 

We created an optogenetic tools capable of regulating gene expression and subsequent protein expression in Saccharomyces cerevisiae in response to blue light. A CRY2/CIB1based optogenetic activator increased expression of a target gene in response to blue light in a time- and dose-dependent manner. Additionally, a flexible optogenetic tool targeted to DNA through a nuclease-dead Cas9 protein was able to repress expression of target genes in a guide-RNA dependent manner. Our optogenetic tools are designed as DNA parts so that optically tunable circuits can be rapidly created using a modular cloning (MoClo) strategy. The modular nature of the toolkit allows us to design, construct, verify, and test optically sensitive circuits rapidly, with the entire cycle taking approximately two weeks.

 

Using the toolkit we have designed synthetic transcription factors with light-controllable nuclear localization and activity. By controlling transcription factor dynamics, we hope to uncover relationships between transcription factor activity, promoter structure, and gene expression. Furthermore, we can control intracellular interactions between cells using light. The spatial control available with light allows us to create structured populations of yeast, for example, through manipulation of expression of a public good.