Expanding the color palette of channelrhodopsins

Fig. 1. Action spectra of four color-variants: blue C2-T159C, cyan C2-T159C-E123T, green C1V1-S220G and orange C1V1-E122T. In grey VChR1 and C1V1 are shown for comparison. From Prigge et al., 2012

In a recent paper published in the Journal of Biological Chemistry, the Hegemann and Deisseroth labs introduced several color-shifted Channelrhodopsins (ChRs) with different absorption and kinetic properties. Prigge et al. mutated several key amino acids in ChR2 and C1V1 to further separate action spectra of those two existing ChRs. The resulting colour-variants are separated by 30 nm from each other and show peak absorption at 460 ,490, 520 and 550 nm respectively (Fig. 1). All color-variants exhibit two times larger photocurrents in HEK-cells then the wild type ChR2. Further engineering yielded off-kinetics spanning the range from ms to s for each colour variant. The two most spectrally separated variants (ChR2 T159C and C1V1-triple) were used to show the feasibility of a separate, wavelength-dependent activation of a HEK cell population expressing on those variants (Fig. 2). In addition the blue absorbing mutant ChR2 T159C-L132C exhibits 3 times larger photocurrents then ChR2 H134R, has a small inactivation and a reduced proton permeation making this variant the most efficient ChR for blue activation so far.

Fig. 2. Using a Fura2-based calcium channel assay, Prigge et al. tested the possibility to separately activate different ChRs in one preparation. (A) Left, HEK cells expressing either C1V1-triple (cyan) or C2-TC (red). Right, Fura2 emission at 515 nm. (B) Fura2 traces while the whole objective plane is illuminated with different wavelength. From Prigge et al., 2012.

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