Reproducible and Sustained Regulation of Gαs Signalling Using a Metazoan Opsin as an Optogenetic Tool

Bailes, Helena J.; Zhuang, Ling; Lucas, Robert J.

doi:10.1371/journal.pone.0030774

Cited by 84 publications

(99 citation statements)

References 40 publications

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“…Therefore, the Opn3 homologs, particularly the MosOpn3, would be highly useful optogenetic tools because of their abilities to form bistable pigments, which are bleach resistant and reusable, with the ubiquitously present retinal and to decrease cAMP efficiently. We have recently reported the jellyfish G s -coupled opsin, which can be used for up-regulating cAMP level in cultured cells (5,34). Together with the jellyfish opsin, the MosOpn3 and its derivatives could be used for regulating not only neural responses, but also nonneuronal responses that are mediated by cAMP-related GPCR signaling in any tissues by light.…”

Section: Discussionmentioning

confidence: 99%

Homologs of vertebrate Opn3 potentially serve as a light sensor in nonphotoreceptive tissue

Koyanagi

Takada

Nagata

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

146

214

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Most opsins selectively bind 11-cis retinal as a chromophore to form a photosensitive pigment, which underlies various physiological functions, such as vision and circadian photoentrainment. Recently, opsin 3 (Opn3), originally called encephalopsin or panopsin, and its homologs were identified in various tissues including brain, eye, and liver in both vertebrates and invertebrates, including human. Because Opn3s are mainly expressed in tissues that are not considered to contain sufficient amounts of 11-cis retinal to form pigments, the photopigment formation ability of Opn3 has been of interest. Here, we report the successful expression of Opn3 homologs, pufferfish teleost multiple tissue opsin (PufTMT) and mosquito Opn3 (MosOpn3) and show that these proteins formed functional photopigments with 11-cis and 9-cis retinals. The PufTMT-and MosOpn3-based pigments have absorption maxima in the blue-to-green region and exhibit a bistable nature. These Opn3 homolog-based pigments activate G i -type and G o -type G proteins light dependently, indicating that they potentially serve as light-sensitive G i /G o -coupled receptors. We also demonstrated that mammalian cultured cells transfected with the MosOpn3 or PufTMT became light sensitive without the addition of 11-cis retinal and the photosensitivity retained after the continuous light exposure, showing a reusable pigment formation with retinal endogenously contained in culture medium. Interestingly, we found that the MosOpn3 also acts as a light sensor when constituted with 13-cis retinal, a ubiquitously present retinal isomer. Our findings suggest that homologs of vertebrate Opn3 might function as photoreceptors in various tissues; furthermore, these Opn3s, particularly the mosquito homolog, could provide a promising optogenetic tool for regulating cAMP-related G protein-coupled receptor signalings.rhodopsin | phototransduction | nonvisual photoreception | opsin diversity M ost opsins bind 11-cis retinal as a chromophore to form a photosensitive pigment (opsin-based pigment) that serves as a light-sensitive G protein-coupled receptor (GPCR). The 11-cis to all-trans isomerization of the chromophore in an opsinbased pigment upon light absorption triggers G protein activation (1-4), and the photoreception of the opsin-based pigment initiates vision and nonvisual functions, including circadian entrainment and pupil responses. Several thousand opsins have been identified, and they are phylogenetically classified into eight groups based on the members that have existed early in animal evolution (5). The phylogenetic classification of each opsin also roughly corresponds to its molecular function. The G t -coupled opsin group contains vertebrate visual pigments, which activate transducin (G t )-type G proteins, and some nonvisual pigments, such as pinopsin, VA-opsin, parapinopsin, and parietopsin (1-4). The G q -coupled opsin group is composed of the visual pigments from many invertebrates, such as insects and cephalopods, and the vertebrate opsin 4 (Opn4) homologs (melanops...

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Section: Discussionmentioning

confidence: 99%

Homologs of vertebrate Opn3 potentially serve as a light sensor in nonphotoreceptive tissue

Koyanagi

Takada

Nagata

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

146

214

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“…The use of optically active rhodopsin chimeric receptors, however, has allowed us to manipulate neurocircuitry in vivo, with higher specificity to endogenous mechanisms (Airan et al, 2009). Currently, there are several chimeric receptors that upon photoactivation are able to signal intracellularly via G-protein-and arrestin-mediated cascades: α 1 -adrenergic, β 2 -adrenergic, adenosine A 2A , 5HT1A, μ-opioid receptor, and the D1 dopamine receptor (Airan et al, 2009;Bailes et al, 2012;Barish et al, 2013;Franke et al, 1992;Gunaydin et al, 2014;Kim et al, 2005;Li et al, 2015;Oh et al, 2010;Siuda et al, 2015a, b). Here we incorporate the chimeric rhodopsin/β 2 -adrenergic receptor (opto-β 2 AR) and show that activation of CaMKIIα+ neurons in the BLA via simulated β-adrenergic activation induces both acute and social anxiety-like states.…”

Section: Discussionmentioning

confidence: 99%

“…Thus, to gain better spatiotemporal isolation of adrenergic receptor pathways, we utilized the chimeric rhodopsin/β 2 -adrenergic receptor, opto-β 2 AR (Figure 3a). Opto-β 2 AR has been shown to activate cAMP, presumably through Gαs-mediated signaling, and to modulate neuronal excitability (Airan et al, 2009;Bailes et al, 2012;Kim et al, 2005;Siuda et al, 2015a), as such this chimeric receptor offers advantages over traditional pharmacological approaches in allowing cell-type specificity and greater spatiotemporal control over βAR signaling.…”

Section: Photoactivation Of β-Adrenergic Signaling In Vivo Within Thementioning

confidence: 99%

Chemogenetic and Optogenetic Activation of Gαs Signaling in the Basolateral Amygdala Induces Acute and Social Anxiety-Like States

Siuda

Al‐Hasani

McCall

et al. 2016

Neuropsychopharmacol

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Anxiety disorders are debilitating psychiatric illnesses with detrimental effects on human health. These heightened states of arousal are often in the absence of obvious threatening cues and are difficult to treat owing to a lack of understanding of the neural circuitry and cellular machinery mediating these conditions. Activation of noradrenergic circuitry in the basolateral amygdala is thought to have a role in stress, fear, and anxiety, and the specific cell and receptor types responsible is an active area of investigation. Here we take advantage of two novel cellular approaches to dissect the contributions of G-protein signaling in acute and social anxiety-like states. We used a chemogenetic approach utilizing the Gαs DREADD (rM3Ds) receptor and show that selective activation of generic Gαs signaling is sufficient to induce acute and social anxiety-like behavioral states in mice. Second, we use a recently characterized chimeric receptor composed of rhodopsin and the β 2 -adrenergic receptor (Opto-β 2 AR) with in vivo optogenetic techniques to selectively activate Gαs β-adrenergic signaling exclusively within excitatory neurons of the basolateral amygdala. We found that optogenetic induction of β-adrenergic signaling in the basolateral amygdala is sufficient to induce acute and social anxiety-like behavior. These findings support the conclusion that activation of Gαs signaling in the basolateral amygdala has a role in anxiety. These data also suggest that acute and social anxiety-like states may be mediated through signaling pathways identical to β-adrenergic receptors, thus providing support that inhibition of this system may be an effective anxiolytic therapy.

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“…The box jellyfish, Carybdea rastonii, expresses an opsin that is Gs coupled (λ-max ∼ 500 nm) (Fig. 1A) (9,22). While following the same strategy described in Fig.…”

Section: Reengineering Opsins To Obtain Specific Combinations Of Specmentioning

confidence: 99%

Optically triggering spatiotemporally confined GPCR activity in a cell and programming neurite initiation and extension

Karunarathne¹,

Giri²,

Kalyanaraman³

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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G-protein-coupled receptor (GPCR) activity gradients evoke important cell behavior but there is a dearth of methods to induce such asymmetric signaling in a cell. Here we achieved reversible, rapidly switchable patterns of spatiotemporally restricted GPCR activity in a single cell. We recruited properties of nonrhodopsin opsins-rapid deactivation, distinct spectral tuning, and resistance to bleachingto activate native Gi, Gq, or Gs signaling in selected regions of a cell. Optical inputs were designed to spatiotemporally control levels of second messengers, IP3, phosphatidylinositol (3,4,5)-triphosphate, and cAMP in a cell. Spectrally selective imaging was accomplished to simultaneously monitor optically evoked molecular and cellular response dynamics. We show that localized optical activation of an opsin-based trigger can induce neurite initiation, phosphatidylinositol (3,4,5)-triphosphate increase, and actin remodeling. Serial optical inputs to neurite tips can refashion early neuron differentiation. Methods here can be widely applied to program GPCRmediated cell behaviors.optogenetics | cell polarity G -protein-coupled receptors (GPCRs) initiate most of the signaling in metazoans and regulate a wide variety of cellular responses that include differentiation, migration, secretion, and contraction. Asymmetric activation of GPCR signaling activity in a cell is thought to play a critical role in varied processes such as cell polarization (1) and modulation of neuron function (2). There is still limited information about activation of signaling that is restricted in space and time across a single cell. An impediment is the lack of methods to continuously vary signal input to a single cell with high time resolution and precision to quantitate second messenger and cellular output from the same cell. A method that provides reversible, temporal control over GPCR activity in restricted regions of a single cell may help govern cell behavior and probe the cellular and molecular basis of single-cell responses.Here we used a set of light-triggered GPCRs, human color opsins, and related nonrhodopsin opsins, to achieve such confined GPCR activation in a single cell. In contrast to molecular gradients, an optical signal provides higher spatiotemporal control and it can be switched on or off or relocated almost instantaneously. We recruited the properties of color opsins to develop optical triggers that spatiotemporally confine signaling. These opsins deactivate rapidly and demonstrate relatively low sensitivity to light (3, 4). The deactivation characteristics curtail the diffusion of activated receptors across a cell and help localize receptor activation to selected regions of a cell. Low susceptibility to bleaching allows continuous reproducible activation. In addition, nonrhodopsin opsins selectively activate different G-protein types, suggesting that they can be individually used to regulate distinct second messengers (4, 5). Rhodopsin or its chimeric forms have been valuable for globally activating G proteins (6-8). Howev...

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Reproducible and Sustained Regulation of Gαs Signalling Using a Metazoan Opsin as an Optogenetic Tool

Cited by 84 publications

References 40 publications

Homologs of vertebrate Opn3 potentially serve as a light sensor in nonphotoreceptive tissue

Homologs of vertebrate Opn3 potentially serve as a light sensor in nonphotoreceptive tissue

Chemogenetic and Optogenetic Activation of Gαs Signaling in the Basolateral Amygdala Induces Acute and Social Anxiety-Like States

Optically triggering spatiotemporally confined GPCR activity in a cell and programming neurite initiation and extension

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