Patterning of Gene Expression Using New Photolabile Groups Applied to Light Activated RNAi

The spacing, timing, and amount of gene expression are crucial for a range of biological processes, including development. For this reason, there have been many attempts to bring gene expression under the control of light. We have previously shown that RNA interference (RNAi) can be controlled with light through the use of siRNA and dsRNA that have their terminal phosphates modified with the dimethoxy nitro phenyl ethyl (DMNPE) group. Upon irradiation, these groups photolyze and release native RNA. The main problem with light activated RNA interference (LARI) to date is that the groups used only partially block RNA interference prior to irradiation, thus limiting the utility of the approach. Here, we describe a new photocleavable group, cyclo-dodecyl DMNPE (CD-DMNPE), designed to completely block the interaction of duplexes with the cellular machinery responsible for RNA interference prior to irradiation. This allowed us to switch from normal to a near complete reduction in gene expression using light, and to construct well-defined patterns of gene expression in cell monolayers. Because this approach is built on the RNA interference pathway, it benefits from the ability to quickly identify duplexes that are effective at low or subnanomolar concentrations. In addition, it allows for the targeting of endogenous genes without additional genetic manipulation. Finally, because of the regiospecificity of CD-DMNPE, it allows a standard duplex to be quickly modified in a single step. The combination of its efficacy and ease of application will allow for the facile control of the spacing, timing, and degree of gene expression in a range of biological systems.