RubyACRs, nonalgal anion channelrhodopsins with highly red-shifted absorption

Significance Our identification and characterization of two anion channelrhodopsins families, one from nonphotosynthetic microorganisms, shows that light-gated anion conductance is more widely spread among eukaryotic lineages than previously thought. The strongly red-shifted absorption spectra of the subset that we designate RubyACRs make them promising candidates for the long-sought inhibitory optogenetic tools producing large passive currents activated by red light, enabling deep tissue penetration. Previously only low-efficiency ion-pumping rhodopsins were available for neural inhibition at similar long wavelengths. The unusual residue composition of the retinal-binding pocket in RubyACRs expands our understanding of color tuning in rhodopsins. Finally, activation of chloride currents by energy transfer from a cytoplasmic fluorescent tag on RubyACRs introduces a potential dimension in molecular engineering of optogenetic tools.