Journal Article

DZNE-2025-01325

http://join2-wiki.gsi.de/foswiki/pub/Main/Artwork/join2_logo100x88.png Optogenetic silencing by combining a rhodopsin cyclase with an engineered cGMP-gated potassium channel.

Spreen, A. ; Sadanandan, N. P. ; Schneider, M. W. ; Kuehn, E. ; Leemisa, A. N. ; De Zio, R. ; Meyer, N. ; Busse, W. ; Erlinghagen, B. ; Adenauer, L. ; Opitz, T. ; Bönigk, W. ; Schiewer, E. ; Heer, J. ; Schmitz, D.DZNE* ; Vierock, J. ; Beck, H.DZNE* ; Hegemann, P. ; Baier, H. ; Schneider-Warme, F. ; Sierra, Y. A. B. ; Seifert, R.

2025
Assoc. Washington, DC [u.a.]

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Please use a persistent id in citations: doi:10.1126/sciadv.adx1195

Abstract: Since the advent of optogenetics, great progress has been made in developing tools to modulate and detect cellular activity using light. We present a 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 guanosine 3',5'-monophosphate (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 summary, RoCK expands our optogenetic toolkit threefold for fast cGMP production, fast cGMP sensing, and K+-based cell silencing.

Keyword(s): Optogenetics: methods (MeSH) ; Animals (MeSH) ; Cyclic GMP: metabolism (MeSH) ; Mice (MeSH) ; Rhodopsin: metabolism (MeSH) ; Rhodopsin: genetics (MeSH) ; Humans (MeSH) ; Hippocampus: metabolism (MeSH) ; Hippocampus: cytology (MeSH) ; Guanylate Cyclase: metabolism (MeSH) ; Guanylate Cyclase: genetics (MeSH) ; Zebrafish (MeSH) ; Potassium Channels: metabolism (MeSH) ; Potassium Channels: genetics (MeSH) ; Protein Engineering (MeSH) ; Myocytes, Cardiac: metabolism (MeSH) ; Neurons: metabolism (MeSH) ; HEK293 Cells (MeSH) ; Cyclic GMP ; Rhodopsin ; Guanylate Cyclase ; Potassium Channels

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Contributing Institute(s):

1. Network Dysfunction (AG Schmitz)
2. Bonn common (Bonn common)

Research Program(s):

1. 351 - Brain Function (POF4-351) (POF4-351)
2. 899 - ohne Topic (POF4-899) (POF4-899)

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