Molecular mechanism for ligand discrimination of cyclic nucleotide-gated channels

Varnum, Michael D.; Black, Kevin D.; Zagotta, William N.

doi:10.1016/0896-6273(95)90150-7

Cited by 187 publications

(230 citation statements)

References 24 publications

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“…The ligand-induced movement of the C-helix is widely thought to initiate the conformational changes that lead to opening of the channel pore, but the structural evidence in support of this hypothesis is equivocal (10,(24)(25)(26)(27)(28)(29). The crystal structure of the HCN2 carboxyl-terminal region in the absence of ligand shows little difference from the cyclic nucleotide-bound structure (12).…”

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confidence: 93%

Double electron–electron resonance reveals cAMP-induced conformational change in HCN channels

Puljung¹,

DeBerg

Zagotta³

et al. 2014

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

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Binding of 3′,5′-cyclic adenosine monophosphate (cAMP) to hyperpolarization-activated cyclic nucleotide-gated (HCN) ion channels regulates their gating. cAMP binds to a conserved intracellular cyclic nucleotide-binding domain (CNBD) in the channel, increasing the rate and extent of activation of the channel and shifting activation to less hyperpolarized voltages. The structural mechanism underlying this regulation, however, is unknown. We used double electron-electron resonance (DEER) spectroscopy to directly map the conformational ensembles of the CNBD in the absence and presence of cAMP. Site-directed, double-cysteine mutants in a soluble CNBD fragment were spin-labeled, and interspin label distance distributions were determined using DEER. We found motions of up to 10 Å induced by the binding of cAMP. In addition, the distributions were narrower in the presence of cAMP. Continuouswave electron paramagnetic resonance studies revealed changes in mobility associated with cAMP binding, indicating less conformational heterogeneity in the cAMP-bound state. From the measured DEER distributions, we constructed a coarse-grained elastic-network structural model of the cAMP-induced conformational transition. We find that binding of cAMP triggers a reorientation of several helices within the CNBD, including the C-helix closest to the cAMP-binding site. These results provide a basis for understanding how the binding of cAMP is coupled to channel opening in HCN and related channels.hyperpolarization-activated ion channels | allosteric regulation

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confidence: 93%

Double electron–electron resonance reveals cAMP-induced conformational change in HCN channels

Puljung¹,

DeBerg

Zagotta³

et al. 2014

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

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“…Stage IV oocytes were isolated as described previously (34). The animal use protocols were consistent with the recommendations of the American Veterinary Medical Association and were approved by the Institutional Animal Care and Use Committee of Washington State University.…”

Section: Methodsmentioning

confidence: 99%

Alternative Splicing Governs Cone Cyclic Nucleotide-gated (CNG) Channel Sensitivity to Regulation by Phosphoinositides

Dai

Lama-Sherpa

Varnum

2014

Journal of Biological Chemistry

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Background: Variants of photoreceptor CNG channels are produced by alternative splicing of precursor mRNA. Results: Inclusion of an optional exon in human CNGA3 transcripts produces channel isoforms with enhanced sensitivity to phosphoinositides via an allosteric mechanism. Conclusion: Alternative splicing of CNGA3 transcripts controls phosphoinositide regulation of cone CNG channels. Significance: These findings reveal the functional importance of CNGA3 alternative splicing.

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“…Protons have been shown to modulate ion channels through effects on ligand binding, channel gating, or ion conduction (16)(17)(18)(19)(20)(21). In the case of the capsaicin receptor, extracellular protons are believed to act primarily by increasing the probability of channel opening (7,22), rather than by altering unitary conductance or interacting directly with a vanilloid-binding site (which may be intracellular) (6,23).…”

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confidence: 99%

Acid potentiation of the capsaicin receptor determined by a key extracellular site

Jordt

Tominaga

Julius

2000

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

581

415

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The capsaicin (vanilloid) receptor, VR1, is a sensory neuron-specific ion channel that serves as a polymodal detector of pain-producing chemical and physical stimuli. The response of VR1 to capsaicin or noxious heat is dynamically potentiated by extracellular protons within a pH range encountered during tissue acidosis, such as that associated with arthritis, infarction, tumor growth, and other forms of injury. A molecular determinant for this important physiological activity was localized to an extracellular Glu residue (E600) in the region linking the fifth transmembrane domain with the putative pore-forming region of the channel. We suggest that this residue serves as a key regulatory site of the receptor by setting sensitivity to other noxious stimuli in response to changes in extracellular proton concentration. We also demonstrate that protons, vanilloids, and heat promote channel opening through distinct pathways, because mutations at a second site (E648) selectively abrogate proton-evoked channel activation without diminishing responses to other noxious stimuli. Our findings provide molecular evidence for stimulus-specific steps in VR1 activation and offer strategies for the development of novel analgesic agents. P ain is initiated when noxious thermal, mechanical, or chemical stimuli excite the peripheral terminals of specialized primary afferent neurons called nociceptors (1). Tissue damage, such as that associated with infection, inflammation, or ischemia, produces an array of chemical mediators that activate or sensitize nociceptor terminals to elicit pain and promote tenderness at the site of injury. An important component of this proalgesic response is local acidosis, namely, a reduction in extracellular pH to levels below the physiological norm of Ϸ7.6 (2). Protons are capable of modulating the activity of a number of receptors and ion channels expressed by primary afferent nociceptors, including acid-sensitive channels of the degenerin family (3), ATPgated channels (4, 5), and vanilloid receptors (6, 7). Analyses of native and cloned vanilloid receptors suggest that they play a role in mediating sustained proton responses in vivo (8)(9)(10)(11)(12). This is further supported by the profoundly reduced responses to acid in cultured dorsal root ganglion neurons and isolated skin nerve preparations of mice with a targeted disruption of the gene encoding for the cloned capsaicin (vanilloid) receptor VR1 (13).Vanilloid receptors are nociceptor-specific cation channels that serve as the molecular target for capsaicin, the main pungent ingredient in ''hot'' chili peppers. VR1 is homologous to members of the transient receptor potential family of ion channels first identified in the Drosophila phototransduction pathway (6,14). We have shown that, when expressed in heterologous systems, VR1 is also activated by noxious heat (6, 7) with a threshold (Ͼ43°C) resembling that of native heat-activated currents in cultured sensory neurons (15). Acidification of the extracellular milieu has two primary effects on V...

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Molecular mechanism for ligand discrimination of cyclic nucleotide-gated channels

Cited by 187 publications

References 24 publications

Double electron–electron resonance reveals cAMP-induced conformational change in HCN channels

Double electron–electron resonance reveals cAMP-induced conformational change in HCN channels

Alternative Splicing Governs Cone Cyclic Nucleotide-gated (CNG) Channel Sensitivity to Regulation by Phosphoinositides

Acid potentiation of the capsaicin receptor determined by a key extracellular site

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