Emerging Diversity of Channelrhodopsins and Their Structure-Function Relationships

Abstract

Cation and anion channelrhodopsins (CCRs and ACRs, respectively) from phototactic algae have become widely used as genetically encoded molecular tools to control cell membrane potential with light. Recent advances in polynucleotide sequencing, especially in environmental samples, have led to identification of hundreds of channelrhodopsin homologs in many phylogenetic lineages, including non-photosynthetic protists. Only a few CCRs and ACRs have been characterized in detail, but there are indications that ion channel function has evolved within the rhodopsin superfamily by convergent routes. The diversity of channelrhodopsins provides an exceptional platform for the study of structure-function evolution in membrane proteins. Here we review the current state of channelrhodopsin research and outline perspectives for its further development.

Keywords: algae; ion channels; microbial rhodopsins; optogenetics; phototaxis.

Figure 1

A schematic of the major lineages in the eukaryotic tree of life adapted from Keeling et al. (2014) distributed under the terms of the Creative Commons Public Domain declaration. The red circles indicate the lineages in which ChR homologs have been identified.

Figure 2

Crystal structures of CrChR2 (6EID; Volkov et al., 2017) and GtACR1 (6EDQ; Li et al., 2019). The side chains of six glutamates conserved in most CCRs and the chromophore are shown as spheres. The intramolecular tunnel is shown in green. SB⁺, protonated Schiff base.

Figure 3

Residue motifs and ionic selectivity of representative ChRs. The residue numbers are according to the CrChR2 sequence. CrChR2, Chlamydomonas reinhardtii channelrhodopsin 2; DsChR1, Dunaliella salina channelrhodopsin 1; MvChR1, Mesostigma viride channelrhodopsin 1; CsCCR, Crustomastix stigmatica cation channelrhodopsin; MsCCR, Mantoniella squamata cation channelrhodopsin; GtACR1, Guillardia theta anion channelrhodopsin 1; RapACR, rapid anion channelrhodopsin from Rhodomonas salina; PgACR1, Phaeocystis globosa anion channelrhodopsin 1; CarACR1, Cafeteria roenbergensis anion channelrhodopsin 1; sTACR1, Stramenopiles sp. TOSAG23–3 anion channelrhodopsin 1; PymeACR1, Pyramimonas melkonianii anion channelrhodopsin 1; Py2087ACR1, Pyramimonas sp. CCMP2087 anion channelrhodopsin 1; HfACR1, Hondaea fermentalgiana anion channelrhodopsin 1; CarACR2, Cafeteria roenbergensis anion channelrhodopsin 2.

Figure 4

The action spectra of photocurrents generated by representative ChRs tagged with EYFP. The magenta line shows the absorption spectrum of EYFP. Note the additional band in the HfACR1 spectrum that reflects FRET from EYFP to rhodopsin. PsChR2, Platymonas subcordiformis channelrhodopsin 2; GtACR1, Guillardia theta anion channelrhodopsin 1; HfACR1, Hondaea fermentalgiana anion channelrhodopsin 1.