Abstract
Cyclic GMP (cGMP) signalling regulates multiple biological functions through activation of protein kinase G and cyclic nucleotide-gated (CNG) channels. In sensory neurons, cGMP permits signal modulation, amplification and encoding, before depolarization. Here we implement a guanylyl cyclase rhodopsin from Blastocladiella emersonii as a new optogenetic tool (BeCyclOp), enabling rapid light-triggered cGMP increase in heterologous cells (Xenopus oocytes, HEK293T cells) and in Caenorhabditis elegans. Among five different fungal CyclOps, exhibiting unusual eight transmembrane topologies and cytosolic N-termini, BeCyclOp is the superior optogenetic tool (light/dark activity ratio: 5,000; no cAMP production; turnover (20 °C) ∼17 cGMP s−1). Via co-expressed CNG channels (OLF in oocytes, TAX-2/4 in C. elegans muscle), BeCyclOp photoactivation induces a rapid conductance increase and depolarization at very low light intensities. In O2/CO2 sensory neurons of C. elegans, BeCyclOp activation evokes behavioural responses consistent with their normal sensory function. BeCyclOp therefore enables precise and rapid optogenetic manipulation of cGMP levels in cells and animals.
Highlights
Cyclic GMP signalling regulates multiple biological functions through activation of protein kinase G and cyclic nucleotide-gated (CNG) channels
An optogenetic tool affecting Cyclic GMP (cGMP) levels in such cells would enable control over intracellular encoding events in cGMP mediated signalling
The full-length cDNA and protein product of BeCyclOp has been confirmed in B. emersonii[26]
Summary
Cyclic GMP (cGMP) signalling regulates multiple biological functions through activation of protein kinase G and cyclic nucleotide-gated (CNG) channels. Optogenetic tools for light-driven cAMP generation exist, for example, photoactivated adenylyl cyclases (PACs), which permit rapid production of cAMP in cells and animals[21,22,23], and can affect behaviour through modulation of specific cellular processes or signalling in the nervous system[24]. Another second messenger widely utilized is cGMP, activating protein kinase G, or CNG channels, especially in sensory neurons in various animals. Naturally evolved light-triggered guanylyl cyclases may be more efficient optogenetic tools
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