Cation and Anion Channelrhodopsins: Sequence Motifs and Taxonomic Distribution
[2];
- University of Texas Health Science Center, Houston, TX (United States); OSTI
- University of Texas Health Science Center, Houston, TX (United States)
- University of Guelph, ON (Canada)
- Max Planck Institute for Plant Breeding Research, Integrative Bioinformatics, Cologne (Germany)
- Chinese Academy of Agricultural Sciences, Shenzhen (China)
- University of Alberta, Edmonton, AB (Canada)
- University of Alberta, Edmonton, AB (Canada); Beijing Genomics Institute—Shenzhen, Shenzhen (China)
Cation and anion channelrhodopsins (CCRs and ACRs, respectively) primarily from two algal species, Chlamydomonas reinhardtii and Guillardia theta, have become widely used as optogenetic tools to control cell membrane potential with light. We mined algal and other protist polynucleotide sequencing projects and metagenomic samples to identify 75 channelrhodopsin homologs from four channelrhodopsin families, including one revealed in dinoflagellates in this study. We carried out electrophysiological analysis of 33 natural channelrhodopsin variants from different phylogenetic lineages and 10 metagenomic homologs in search of sequence determinants of ion selectivity, photocurrent desensitization, and spectral tuning in channelrhodopsins. Our results show that association of a reduced number of glutamates near the conductance path with anion selectivity depends on a wider protein context, because prasinophyte homologs with a glutamate pattern identical to that in cryptophyte ACRs are cation selective. Desensitization is also broadly context dependent, as in one branch of stramenopile ACRs and their metagenomic homologs, its extent roughly correlates with phylogenetic relationship of their sequences. Regarding spectral tuning, we identified two prasinophyte CCRs with red-shifted spectra to 585 nm. They exhibit a third residue pattern in their retinal-binding pockets distinctly different from those of the only two types of red-shifted channelrhodopsins known (i.e., the CCR Chrimson and RubyACRs). In cryptophyte ACRs we identified three specific residue positions in the retinal-binding pocket that define the wavelength of their spectral maxima. Lastly, we found that dinoflagellate rhodopsins with a TCP motif in the third transmembrane helix and a metagenomic homolog exhibit channel activity. Channelrhodopsins are widely used in neuroscience and cardiology as research tools and are considered prospective therapeutics, but their natural diversity and mechanisms remain poorly characterized. Genomic and metagenomic sequencing projects are producing an ever-increasing wealth of data, whereas biophysical characterization of the encoded proteins lags behind. In this study, we used manual and automated patch clamp recording of representative members of four channelrhodopsin families, including a family in dinoflagellates that we report in this study. Our results contribute to a better understanding of molecular determinants of ionic selectivity, photocurrent desensitization, and spectral tuning in channelrhodopsins.
- Research Organization:
- USDOE Joint Genome Institute (JGI), Berkeley, CA (United States)
- Sponsoring Organization:
- National Institutes of Health (NIH); Natural Sciences and Engineering Research Council of Canada (NSERC); Robert A. Welch Foundation; USDOE Office of Science (SC)
- Grant/Contract Number:
- AC02-05CH11231
- OSTI ID:
- 1904117
- Journal Information:
- mBio (Online), Journal Name: mBio (Online) Journal Issue: 4 Vol. 12; ISSN 2150-7511
- Publisher:
- American Society for Microbiology (ASM)Copyright Statement
- Country of Publication:
- United States
- Language:
- English
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