Figure 4: Functional testing of the biosensor in Hank’s buffer
containing 1 mM Ca2+ confirms dose-dependent
activation of Orco by VUAA1. The response is analogous to membrane
fluctuations observed following activation of a TRP cationic channel.
The p-values for the difference in normalized fluorescence between
PP-Orco and GS115 strains were lower than 0.01 for the 0.5 mM, 1 mM and
2 mM runs and 0.001 when the concentration of VUAA1 was 1.5 mM.
The normalized fluorescence emitted by these cultures reaches a maximum
1 minute after the compound has been injected into the medium. The
normalized fluorescence then gradually drops over the next 2-3 minutes,
after which it equilibrates for the remainder of the run. This response
closely mirrors the fluctuations in membrane potentials that are induced
followed activation of transient receptor potential (TRP) channels such
as TRPA1 in D. melanogaster and A. gambiae [45–47].
TRP channels are ion channels that transduce a range of stimuli,
including heat, light, taste, pain and pressure; and the functional
similarity between a seven-transmembrane-helix protein such as Orco and
six-transmembrane-helix TRP channels is notable.