Synthetic Biology Parts, Devices and Applications

176 8 Design of Ligand‐Controlled Genetic Switches Based on RNA Interference

RNP motif

Tr igger protein

Top view

RNA motif

(a)

(b)

Switch X with

trigger protein

Accessible: not functional Inaccessible: functional

Knockdown

Dicer Dicer

Knockdown

target gene target gene

Switch Y with

trigger protein

Switch X with

trigger protein

of of

Switch Y with

trigger protein

~2.6

Å/bp

30°/bp

dsRNA

+

Figure 8.3 3D design scheme of the protein‐responsive shRNA switch. (a) Protein‐responsive

shRNA switches are three‐dimensionally designed by connecting an RNP motif to the

corresponding dsRNA in silico. After specifying the position of the devices with reference to

the Dicer cleavage sites and comparing the locations of the bound proteins of the X and Y

switches, the bound trigger protein on switch X rotates ~30° in a counterclockwise direction

around the axis of the dsRNA and is located ~2.6 Å more distant from the Dicer cleavage sites

than switch Y. (b) Dicer can access and process switch Y via trigger protein binding; the

switch then induces the knockdown of its target gene via RNAi (left). Dicer is inaccessible to

switch X in the presence of the trigger protein because the RNP interaction faces Dicer and

inhibits its access (right). The prevention of Dicer function causes the derepression of gene

knockdown.