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

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

doi:10.1016/j.cell.2022.01.007

Cited by 79 publications

(100 citation statements)

References 113 publications

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“…During preparation of this manuscript, an independent report of a retinal-bound ChRmine structure determined in detergent micelles and in complex with antibody fragments to 2.0 Å resolution was posted and published 37 . Two substantial differences are observed between that structure and the ones reported here.…”

Section: Resultsmentioning

confidence: 99%

Cryo-EM structures of the channelrhodopsin ChRmine in lipid nanodiscs

et al. 2022

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Microbial channelrhodopsins are light-gated ion channels widely used for optogenetic manipulation of neuronal activity. ChRmine is a bacteriorhodopsin-like cation channelrhodopsin (BCCR) more closely related to ion pump rhodopsins than other channelrhodopsins. ChRmine displays unique properties favorable for optogenetics including high light sensitivity, a broad, red-shifted activation spectrum, cation selectivity, and large photocurrents, while its slow closing kinetics impedes some applications. The structural basis for ChRmine function, or that of any other BCCR, is unknown. Here, we present cryo-EM structures of ChRmine in lipid nanodiscs in apo (opsin) and retinal-bound (rhodopsin) forms. The structures reveal an unprecedented trimeric architecture with a lipid filled central pore. Large electronegative cavities on either side of the membrane facilitate high conductance and selectivity for cations over protons. The retinal binding pocket structure suggests channel properties could be tuned with mutations and we identify ChRmine variants with ten-fold decreased and two-fold increased closing rates. A T119A mutant shows favorable properties relative to wild-type and previously reported ChRmine variants for optogenetics. These results provide insight into structural features that generate an ultra-potent microbial opsin and provide a platform for rational engineering of channelrhodopsins with improved properties that could expand the scale, depth, and precision of optogenetic experiments.

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Section: Resultsmentioning

confidence: 99%

Cryo-EM structures of the channelrhodopsin ChRmine in lipid nanodiscs

et al. 2022

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“…Meanwhile, quite a number of crystal structures have been resolved for various classes of type-1 rhodopsins ( Table 1 ). The most recent high resolution 3-D structures actually capitalized on the fantastic progress in cryo-EM ( Hirschi et al, 2021 ; Kishi et al, 2022 ).…”

Section: Spectral and Structural Properties And Solubilizationmentioning

confidence: 99%

“…For this purpose, mutagenesis of far-red absorbing rhodopsins like Crimson and CrimsonSA would be a good starting point ( Oda et al, 2018 ). Another option is the novel channelrhodopsin ChRmine, which has quite unusual properties, including a trimeric structure similar to BR ( Marshel et al, 2019 ; Kishi et al, 2022 ). A very fascinating example is NeoR, a subunit in the heterodimeric rhodopsin-cyclase from the fungus Rhizoclosmatium globosum.…”

Section: Bioengineeringmentioning

confidence: 99%

Rhodopsins: An Excitingly Versatile Protein Species for Research, Development and Creative Engineering

Grip

Ganapathy

2022

Front. Chem.

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The first member and eponym of the rhodopsin family was identified in the 1930s as the visual pigment of the rod photoreceptor cell in the animal retina. It was found to be a membrane protein, owing its photosensitivity to the presence of a covalently bound chromophoric group. This group, derived from vitamin A, was appropriately dubbed retinal. In the 1970s a microbial counterpart of this species was discovered in an archaeon, being a membrane protein also harbouring retinal as a chromophore, and named bacteriorhodopsin. Since their discovery a photogenic panorama unfolded, where up to date new members and subspecies with a variety of light-driven functionality have been added to this family. The animal branch, meanwhile categorized as type-2 rhodopsins, turned out to form a large subclass in the superfamily of G protein-coupled receptors and are essential to multiple elements of light-dependent animal sensory physiology. The microbial branch, the type-1 rhodopsins, largely function as light-driven ion pumps or channels, but also contain sensory-active and enzyme-sustaining subspecies. In this review we will follow the development of this exciting membrane protein panorama in a representative number of highlights and will present a prospect of their extraordinary future potential.

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“…Also relevant with this regard, more favourable wavelengths to minimize the crosstalk between the “classical couple” GCaMP6 and the C1 V1 have been recently reported. The possibility to further extend the palette of sensors and actuators is exemplified by works like the recent one characterizing variants of the fast, red-shifted opsin ChRmine that allowed to perform 3-colour all optical physiology (at one-photon though) ( Kishi et al, 2022 ). In relation to fact that in the future three-photon microscopy will be probably more used to perform optical measurements in deep brain structure, the problem of the spectral cross talk will possibly be worsen as the three-photon excitation spectra of fluorophores can be broader than the ones measured at two-photon (e.g., Liu et al, 2020 ).…”

Section: Physiologically Relevant Pro and Cons Of All-optical Approac...mentioning

confidence: 99%

Advantages, Pitfalls, and Developments of All Optical Interrogation Strategies of Microcircuits in vivo

Papaioannou

Medini

2022

Front. Neurosci.

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The holy grail for every neurophysiologist is to conclude a causal relationship between an elementary behaviour and the function of a specific brain area or circuit. Our effort to map elementary behaviours to specific brain loci and to further manipulate neural activity while observing the alterations in behaviour is in essence the goal for neuroscientists. Recent advancements in the area of experimental brain imaging in the form of longer wavelength near infrared (NIR) pulsed lasers with the development of highly efficient optogenetic actuators and reporters of neural activity, has endowed us with unprecedented resolution in spatiotemporal precision both in imaging neural activity as well as manipulating it with multiphoton microscopy. This readily available toolbox has introduced a so called all-optical physiology and interrogation of circuits and has opened new horizons when it comes to precisely, fast and non-invasively map and manipulate anatomically, molecularly or functionally identified mesoscopic brain circuits. The purpose of this review is to describe the advantages and possible pitfalls of all-optical approaches in system neuroscience, where by all-optical we mean use of multiphoton microscopy to image the functional response of neuron(s) in the network so to attain flexible choice of the cells to be also optogenetically photostimulated by holography, in absence of electrophysiology. Spatio-temporal constraints will be compared toward the classical reference of electrophysiology methods. When appropriate, in relation to current limitations of current optical approaches, we will make reference to latest works aimed to overcome these limitations, in order to highlight the most recent developments. We will also provide examples of types of experiments uniquely approachable all-optically. Finally, although mechanically non-invasive, all-optical electrophysiology exhibits potential off-target effects which can ambiguate and complicate the interpretation of the results. In summary, this review is an effort to exemplify how an all-optical experiment can be designed, conducted and interpreted from the point of view of the integrative neurophysiologist.

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

Abstract: Highlightsd The cryo-EM structure of a pump-like channelrhodopsin, at 2.0 A ˚resolution d Identification of key features distinguishing ChRmine from other channelrhodopsins d Identification of key features distinguishing ChRmine from ion pump rhodopsins

Cited by 79 publications

References 113 publications

Cryo-EM structures of the channelrhodopsin ChRmine in lipid nanodiscs

Cryo-EM structures of the channelrhodopsin ChRmine in lipid nanodiscs

Rhodopsins: An Excitingly Versatile Protein Species for Research, Development and Creative Engineering

Advantages, Pitfalls, and Developments of All Optical Interrogation Strategies of Microcircuits in vivo

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