Optogenetic silencing by combining a rhodopsin cyclase with an engineered cGMP-gated potassium channel

Since the advent of optogenetics, significant progress has been made in developing tools to modulate and detect cellular activity using light. We present a novel two-component optogenetic silencing tool, RoCK (Rhodopsin Cyclase/K+ channel), which pairs the rhodopsin guanylyl cyclase CaRhGC with customized SthK K⁺ channels that are engineered to open selectively upon cGMP binding. By enhancing the cGMP sensitivity and open probability of SthK mutants, we obtained four channel variants suited for different levels of cGMP concentration. CaRhGC’s membrane-bound nature enables localized cGMP production, and the lack of dark activity reduces the risk for off-target effects. Optimized RoCK effectively modulated cellular activity in mouse hippocampal neurons, in acute hippocampal slices, and in rabbit cardiomyocytes. In zebrafish, RoCK silenced motor neurons in vivo, suppressing the characteristic coiling behavior of embryos, thus highlighting its potential for behavioral studies. In summa..., , , # Optogenetic silencing by combining a rhodopsin cyclase with an engineered cGMP-gated potassium channel

Dataset DOI: [10.5061/dryad.k98sf7mm2](10.5061/dryad.k98sf7mm2)

## Description of the data and file structure

The CSV files contain multiple tables, each corresponding to one figure of the manuscript and labeled with the respective figure number. Each table provides the underlying data required to reproduce and replot the corresponding figure.

### Files and variables

#### File: Figure_1i.csv

**Description:** The ratio of maximal ion channel current I(cGMP)/I(cAMP) for different ion channel mutants of the SthK K+-channel is given for different experiments.

##### Variables

* Figure 1i: Figure identifier
* SthK1.0: I(cGMP)/I(cAMP)
* SthK1.1: I(cGMP)/I(cAMP)
* SthK2.0: I(cGMP)/I(cAMP)
* SthK2.1: I(cGMP)/I(cAMP)

#### File: Figure_4b.csv

**Description:** This file gives the diastolic resting membrane potential in between spikes of cardiomyocytes during different phases of an opto...