# Structure/Function of Channelrhodopsins and Related Retinylidene Proteins

> **NIH NIH R35** · UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON · 2022 · $627,370

## Abstract

My laboratory focuses on the structure, function, and mechanisms of microbial rhodopsins, widespread visual
pigment-like proteins with diverse functions. Over the past decade, a subfamily, light-gated ion channels
(channelrhodopsins), have had exceptional impact because of their central role in the transformative technology
of optogenetics. We originally found them in the chlorophyte alga Chlamydomonas reinhardtii as phototaxis
receptors that depolarize the cell membrane by producing cation currents in response to light. Subsequently
neuroscientists found that these light-gated cation channelrhodopsins (CCRs) expressed in neurons produce
depolarizing currents that enable light to trigger action potentials. Targeted photoactivation of neurons enabled
by expression of CCRs in neural circuits has proven to be a powerful technique transforming many aspects of
neuroscience research. Nevertheless, their light-gated channel activity is one of the least understood rhodopsin
functions in terms of molecular mechanisms. Several advances in our work over the past 5 years, coupled to our
knowledge and expertise over decades of research on microbial rhodopsins, guide our current research strategy.
In 2015 we discovered exclusively anion-conducting (physiologically Cl-) channelrhodopsins (ACRs) in the
distant phylum of cryptophyte algae. A breakthrough for optogenetics, ACRs enable efficient light-induced
hyperpolarization and therefore are potent inhibitors of neuron firing. Also seminal to our research plans, our
recent crystal structure of the most used ACR in optogenetics (GtACR1 from Guillardia theta) revealed a
preexisting tunnel in the closed dark state that we propose is the channel closed by 3 well-defined constrictions.
The GtACR1 tunnel is the only candidate ion pathway imaged in a channelrhodopsin, and provides a valuable
resource for elucidating the mystery of channel gating by light. Principles learned from our study will likely
enhance our understanding also of other microbial rhodopsins. Our current research investigates the diversity
and molecular mechanisms of channelrhodopsins by: (i) ongoing genome mining to expand our knowledge and
also advance optogenetics, focused on ACRs, but including CCRs (e.g. possible K+ and Ca++ channels).
Recently we identified two new ACR families and long-sought red-shifted ACRs (“RubyACRs”) activated by
tissue-penetrating long wavelengths, valuable for optogenetics and opening the way to elucidating color tuning
mechanisms of channelrhodopsins; (ii) unraveling the relationship of electrical steps in channel function to
photochemical transitions by structure-based mutagenesis, photo-electrophysiology in vivo, and kinetic optical
and vibrational spectroscopy in vitro; and (iii) determination of atomic structures by X-ray crystallography and
cryoEM, including innovative approaches to image the transient open-channel conformation. Elucidating
mechanisms of channelrhodopsins will advance basic science and also facili...

## Key facts

- **NIH application ID:** 10380871
- **Project number:** 5R35GM140838-02
- **Recipient organization:** UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
- **Principal Investigator:** JOHN LEE SPUDICH
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $627,370
- **Award type:** 5
- **Project period:** 2021-05-01 → 2026-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10380871

## Citation

> US National Institutes of Health, RePORTER application 10380871, Structure/Function of Channelrhodopsins and Related Retinylidene Proteins (5R35GM140838-02). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10380871. Licensed CC0.

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