Dynamics of G protein effector interactions and their impact on timing and sensitivity of G protein-mediated signal transduction

Bodmann, Eva-Lisa; Wolters, Valerie; Bünemann, Moritz

doi:10.1016/j.ejcb.2015.06.004

Cited by 12 publications

(7 citation statements)

References 37 publications

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“…Moreover, LWS cone opsin and RL3Am could be valuable optogenetic tools for selectively controlling Gi and Go, respectively. Gi is ubiquitously expressed in human cells and regulates a wide variety of processes through a common mechanism of suppressing cAMP [ 47 ]. Go is especially interesting because it is the most abundant Gα in the central nervous system, being 5–10 times more abundant than Gi and comprising up to 1% of membrane proteins in the brain [ 15 ].…”

Section: Discussionmentioning

confidence: 99%

“…One important distinction is that the GsX assay utilizes a live-cell reporter, which allows continuous real-time monitoring, thus facilitating kinetic analysis, whereas the TGF-β shedding assay relies on immunoassays as an end-point readout, so kinetic information must be inferred by collecting end points across a time series, which decreases throughput. As with all assays that monitor second messenger levels, response kinetics in the GsX assay reflect the balance of competing enzymatic activities, in this case adenylyl cyclase and phosphodiesterases, and must be interpreted with this in mind [ 47 ].…”

Section: Discussionmentioning

confidence: 99%

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A live cell assay of GPCR coupling allows identification of optogenetic tools for controlling Go and Gi signaling

et al. 2018

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BackgroundAnimal opsins are light-sensitive G-protein-coupled receptors (GPCRs) that enable optogenetic control over the major heterotrimeric G-protein signaling pathways in animal cells. As such, opsins have potential applications in both biomedical research and therapy. Selecting the opsin with the best balance of activity and selectivity for a given application requires knowing their ability to couple to a full range of relevant Gα subunits. We present the GsX assay, a set of tools based on chimeric Gs subunits that transduce coupling of opsins to diverse G proteins into increases in cAMP levels, measured with a real-time reporter in living cells. We use this assay to compare coupling to Gi/o/t across a panel of natural and chimeric opsins selected for potential application in gene therapy for retinal degeneration.ResultsOf the opsins tested, wild-type human rod opsin had the highest activity for chimeric Gs proxies for Gi and Gt (Gsi and Gst) and was matched in Go proxy (Gso) activity only by a human rod opsin/scallop opsin chimera. Rod opsin drove roughly equivalent responses via Gsi, Gso, and Gst, while cone opsins showed much lower activities with Gso than Gsi or Gst, and a human rod opsin/amphioxus opsin chimera demonstrated higher activity with Gso than with Gsi or Gst. We failed to detect activity for opsin chimeras bearing three intracellular fragments of mGluR6, and observed unexpectedly complex response profiles for scallop and amphioxus opsins thought to be specialized for Go.ConclusionsThese results identify rod opsin as the most potent non-selective Gi/o/t-coupled opsin, long-wave sensitive cone opsin as the best for selectively activating Gi/t over Go, and a rod opsin/amphioxus opsin chimera as the best choice for selectively activating Go over Gi/t.Electronic supplementary materialThe online version of this article (doi:10.1186/s12915-017-0475-2) contains supplementary material, which is available to authorized users.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

A live cell assay of GPCR coupling allows identification of optogenetic tools for controlling Go and Gi signaling

et al. 2018

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“…In general, protein-carbohydrate interactions are usually weak and low-affine (micromolar-millimolar range) ( Holgersson et al, 2005 ) and signalling therefore, emerges as being controlled by ligand multivalency and/or by receptor multiplicity ( Kiessling and Pohl, 1996 ; Rabinovich, 2002 ; Vasta et al, 2012 ). In line with this notion studies of receptors present at the plant and mammalian plasma membrane revealed a conserved strategy to ensure specific, instantaneous, switchable and evolvable downstream signalling; namely, increased responsiveness and specificity via combinatorial systems ( Ostrom et al, 2001 ; Piñeyro, 2009 ; Bodmann et al, 2015 ; Bücherl et al, 2017 ). The signalling properties of NFRe remain to be determined, but our findings based on the properties of this LysM receptor kinase, together with the symbiotic phenotypes of nfre mutants unveil a more complex signalling operating in the epidermal cells of L. japonicus than anticipated from studies of the basic and essential receptor-components.…”

Section: Discussionmentioning

confidence: 81%

Epidermal LysM receptor ensures robust symbiotic signalling in Lotus japonicus

Murakami

Cheng

Gysel

et al. 2018

eLife

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Recognition of Nod factors by LysM receptors is crucial for nitrogen-fixing symbiosis in most legumes. The large families of LysM receptors in legumes suggest concerted functions, yet only NFR1 and NFR5 and their closest homologs are known to be required. Here we show that an epidermal LysM receptor (NFRe), ensures robust signalling in L. japonicus. Mutants of Nfre react to Nod factors with increased calcium spiking interval, reduced transcriptional response and fewer nodules in the presence of rhizobia. NFRe has an active kinase capable of phosphorylating NFR5, which in turn, controls NFRe downstream signalling. Our findings provide evidence for a more complex Nod factor signalling mechanism than previously anticipated. The spatio-temporal interplay between Nfre and Nfr1, and their divergent signalling through distinct kinases suggests the presence of an NFRe-mediated idling state keeping the epidermal cells of the expanding root system attuned to rhizobia.

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“…Hydrolysis of GTP to GDP and re-association of the trimeric G protein with the GPRC results in inactivation of these pathways. 4 The R183Q mutation in GNAQ is predicted to result in a protein with impaired auto-hydrolysis of activated Gαq, and therefore impaired inactivation of Gαq. The current understanding and data suggests that the mutation results in hyper-activation of downstream pathways, which include RAS-MEK-ERK, HIPPO-YAP, 5 and, indirectly, mTOR (Figure 1).…”

Section: Introductionmentioning

confidence: 99%