Structural basis for channel conduction in the pump-like channelrhodopsin ChRmine

Koichiro E Kishi; Yoon Seok Kim; Masahiro Fukuda; Masatoshi Inoue; Tsukasa Kusakizako; Peter Y Wang; Charu Ramakrishnan; Eamon F X Byrne; Elina Thadhani; Joseph M Paggi; Toshiki E Matsui; Keitaro Yamashita; Takashi Nagata; Masae Konno; Sean Quirin; Maisie Lo; Tyler Benster; Tomoko Uemura; Kehong Liu; Mikihiro Shibata; Norimichi Nomura; So Iwata; Osamu Nureki; Ron O Dror; Keiichi Inoue; Karl Deisseroth; Hideaki E Kato

doi:10.1016/j.cell.2022.01.007

Structural basis for channel conduction in the pump-like channelrhodopsin ChRmine

Cell. 2022 Feb 17;185(4):672-689.e23. doi: 10.1016/j.cell.2022.01.007. Epub 2022 Feb 2.

Authors

Koichiro E Kishi¹, Yoon Seok Kim², Masahiro Fukuda¹, Masatoshi Inoue², Tsukasa Kusakizako³, Peter Y Wang², Charu Ramakrishnan⁴, Eamon F X Byrne², Elina Thadhani⁵, Joseph M Paggi⁶, Toshiki E Matsui¹, Keitaro Yamashita⁷, Takashi Nagata⁸, Masae Konno⁸, Sean Quirin², Maisie Lo², Tyler Benster², Tomoko Uemura⁹, Kehong Liu⁹, Mikihiro Shibata¹⁰, Norimichi Nomura⁹, So Iwata¹¹, Osamu Nureki³, Ron O Dror¹², Keiichi Inoue¹³, Karl Deisseroth¹⁴, Hideaki E Kato¹⁵

Affiliations

¹ Komaba Institute for Science, The University of Tokyo, Meguro, Tokyo, Japan.
² Department of Bioengineering, Stanford University, Stanford, CA, USA.
³ Department of Biological Sciences, Graduate School of Science, The University of Tokyo, Bunkyo, Tokyo, Japan.
⁴ CNC Program, Stanford University, Palo Alto, CA, USA.
⁵ Department of Bioengineering, Stanford University, Stanford, CA, USA; Department of Computer Science, Stanford University, Stanford, CA, USA.
⁶ Department of Computer Science, Stanford University, Stanford, CA, USA.
⁷ MRC Laboratory of Molecular Biology, Cambridge Biomedical Campus, Cambridge, UK.
⁸ Institute for Solid State Physics, The University of Tokyo, Kashiwa, Japan; PRESTO, Japan Science and Technology Agency, Kawaguchi, Saitama, Japan.
⁹ Department of Cell Biology, Graduate School of Medicine, Kyoto University, Kyoto, Sakyo, Japan.
¹⁰ WPI Nano Life Science Institute (WPI-NanoLSI), Kanazawa University, Kakuma, Kanazawa, Japan; High-Speed AFM for Biological Application Unit, Institute for Frontier Science Initiative, Kanazawa University, Kakuma, Kanazawa, Japan.
¹¹ Department of Cell Biology, Graduate School of Medicine, Kyoto University, Kyoto, Sakyo, Japan; RIKEN SPring-8 Center, Kouto, Sayo-cho, Sayo-gun, Hyogo, Japan.
¹² Department of Computer Science, Stanford University, Stanford, CA, USA; Institute for Computational and Mathematical Engineering, Stanford University, Stanford, CA, USA.
¹³ Institute for Solid State Physics, The University of Tokyo, Kashiwa, Japan.
¹⁴ Department of Bioengineering, Stanford University, Stanford, CA, USA; CNC Program, Stanford University, Palo Alto, CA, USA; Howard Hughes Medical Institute, Stanford University, Stanford, CA, USA; Department of Psychiatry and Behavioral Sciences, Stanford University, Stanford, CA, USA. Electronic address: deissero@stanford.edu.
¹⁵ Komaba Institute for Science, The University of Tokyo, Meguro, Tokyo, Japan; Department of Biological Sciences, Graduate School of Science, The University of Tokyo, Bunkyo, Tokyo, Japan; PRESTO, Japan Science and Technology Agency, Kawaguchi, Saitama, Japan; FOREST, Japan Science and Technology Agency, Kawaguchi, Saitama, Japan. Electronic address: hekato@bio.c.u-tokyo.ac.jp.

Abstract

ChRmine, a recently discovered pump-like cation-conducting channelrhodopsin, exhibits puzzling properties (large photocurrents, red-shifted spectrum, and extreme light sensitivity) that have created new opportunities in optogenetics. ChRmine and its homologs function as ion channels but, by primary sequence, more closely resemble ion pump rhodopsins; mechanisms for passive channel conduction in this family have remained mysterious. Here, we present the 2.0 Å resolution cryo-EM structure of ChRmine, revealing architectural features atypical for channelrhodopsins: trimeric assembly, a short transmembrane-helix 3, a twisting extracellular-loop 1, large vestibules within the monomer, and an opening at the trimer interface. We applied this structure to design three proteins (rsChRmine and hsChRmine, conferring further red-shifted and high-speed properties, respectively, and frChRmine, combining faster and more red-shifted performance) suitable for fundamental neuroscience opportunities. These results illuminate the conduction and gating of pump-like channelrhodopsins and point the way toward further structure-guided creation of channelrhodopsins for applications across biology.

Keywords: ChRmine; PLCR; all-optical; channelrhodopsin; cryo-EM; microbial opsin; optogenetics; pump-like channelrhodopsin; structure-guided engineering.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Animals
Channelrhodopsins / chemistry*
Channelrhodopsins / metabolism*
Channelrhodopsins / ultrastructure
Cryoelectron Microscopy
Female
HEK293 Cells
Humans
Ion Channel Gating*
Male
Mice
Mice, Inbred C57BL
Models, Molecular
Optogenetics
Phylogeny
Rats
Rats, Sprague-Dawley
Schiff Bases / chemistry
Sf9 Cells
Structure-Activity Relationship

Substances

Channelrhodopsins
Schiff Bases

Abstract

Publication types

MeSH terms

Substances

Grants and funding