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. 2000 May;78(5):2321–2333. doi: 10.1016/S0006-3495(00)76778-6

Mutating three residues in the bovine rod cyclic nucleotide-activated channel can switch a nucleotide from inactive to active.

S P Scott 1, J Cummings 1, J C Joe 1, J C Tanaka 1
PMCID: PMC1300823  PMID: 10777730

Abstract

Cyclic nucleotide-gated (CNG) channels, which were initially studied in retina and olfactory neurons, are activated by cytoplasmic cGMP or cAMP. Detailed comparisons of nucleotide-activated currents using nucleotide analogs and mutagenesis revealed channel-specific residues in the nucleotide-binding domain that regulate the binding and channel-activation properties. Of particular interest are N(1)-oxide cAMP, which does not activate bovine rod channels, and Rp-cGMPS, which activates bovine rod, but not catfish, olfactory channels. Previously, we showed that four residues coordinate the purine interactions in the binding domain and that three of these residues vary in the alpha subunits of the bovine rod, catfish, and rat olfactory channels. Here we show that both N(1)-oxide cAMP and Rp-cGMPS activate rat olfactory channels. A mutant of the bovine rod alpha subunit, substituted with residues from the rat olfactory channel at the three variable positions, was weakly activated by N(1)-oxide cAMP, and a catfish olfactory-like bovine rod mutant lost activation by Rp-cGMPS. These experiments underscore the functional importance of purine contacts with three residues in the cyclic nucleotide-binding domain. Molecular models of nucleotide analogs in the binding domains, constructed with AMMP, showed differences in the purine contacts among the channels that might account for activation differences.

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Selected References

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  1. Ahmad I., Korbmacher C., Segal A. S., Cheung P., Boulpaep E. L., Barnstable C. J. Mouse cortical collecting duct cells show nonselective cation channel activity and express a gene related to the cGMP-gated rod photoreceptor channel. Proc Natl Acad Sci U S A. 1992 Nov 1;89(21):10262–10266. doi: 10.1073/pnas.89.21.10262. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Ahmad I., Leinders-Zufall T., Kocsis J. D., Shepherd G. M., Zufall F., Barnstable C. J. Retinal ganglion cells express a cGMP-gated cation conductance activatable by nitric oxide donors. Neuron. 1994 Jan;12(1):155–165. doi: 10.1016/0896-6273(94)90160-0. [DOI] [PubMed] [Google Scholar]
  3. Altenhofen W., Ludwig J., Eismann E., Kraus W., Bönigk W., Kaupp U. B. Control of ligand specificity in cyclic nucleotide-gated channels from rod photoreceptors and olfactory epithelium. Proc Natl Acad Sci U S A. 1991 Nov 1;88(21):9868–9872. doi: 10.1073/pnas.88.21.9868. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Arancio O., Kandel E. R., Hawkins R. D. Activity-dependent long-term enhancement of transmitter release by presynaptic 3',5'-cyclic GMP in cultured hippocampal neurons. Nature. 1995 Jul 6;376(6535):74–80. doi: 10.1038/376074a0. [DOI] [PubMed] [Google Scholar]
  5. Arancio O., Kiebler M., Lee C. J., Lev-Ram V., Tsien R. Y., Kandel E. R., Hawkins R. D. Nitric oxide acts directly in the presynaptic neuron to produce long-term potentiation in cultured hippocampal neurons. Cell. 1996 Dec 13;87(6):1025–1035. doi: 10.1016/s0092-8674(00)81797-3. [DOI] [PubMed] [Google Scholar]
  6. Baumann A., Frings S., Godde M., Seifert R., Kaupp U. B. Primary structure and functional expression of a Drosophila cyclic nucleotide-gated channel present in eyes and antennae. EMBO J. 1994 Nov 1;13(21):5040–5050. doi: 10.1002/j.1460-2075.1994.tb06833.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Biel M., Altenhofen W., Hullin R., Ludwig J., Freichel M., Flockerzi V., Dascal N., Kaupp U. B., Hofmann F. Primary structure and functional expression of a cyclic nucleotide-gated channel from rabbit aorta. FEBS Lett. 1993 Aug 23;329(1-2):134–138. doi: 10.1016/0014-5793(93)80209-d. [DOI] [PubMed] [Google Scholar]
  8. Bradley J., Zhang Y., Bakin R., Lester H. A., Ronnett G. V., Zinn K. Functional expression of the heteromeric "olfactory" cyclic nucleotide-gated channel in the hippocampus: a potential effector of synaptic plasticity in brain neurons. J Neurosci. 1997 Mar 15;17(6):1993–2005. doi: 10.1523/JNEUROSCI.17-06-01993.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Butt E., Pöhler D., Genieser H. G., Huggins J. P., Bucher B. Inhibition of cyclic GMP-dependent protein kinase-mediated effects by (Rp)-8-bromo-PET-cyclic GMPS. Br J Pharmacol. 1995 Dec;116(8):3110–3116. doi: 10.1111/j.1476-5381.1995.tb15112.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Butt E., van Bemmelen M., Fischer L., Walter U., Jastorff B. Inhibition of cGMP-dependent protein kinase by (Rp)-guanosine 3',5'-monophosphorothioates. FEBS Lett. 1990 Apr 9;263(1):47–50. doi: 10.1016/0014-5793(90)80702-k. [DOI] [PubMed] [Google Scholar]
  11. Bönigk W., Altenhofen W., Müller F., Dose A., Illing M., Molday R. S., Kaupp U. B. Rod and cone photoreceptor cells express distinct genes for cGMP-gated channels. Neuron. 1993 May;10(5):865–877. doi: 10.1016/0896-6273(93)90202-3. [DOI] [PubMed] [Google Scholar]
  12. Bönigk W., Müller F., Middendorff R., Weyand I., Kaupp U. B. Two alternatively spliced forms of the cGMP-gated channel alpha-subunit from cone photoreceptor are expressed in the chick pineal organ. J Neurosci. 1996 Dec 1;16(23):7458–7468. doi: 10.1523/JNEUROSCI.16-23-07458.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Dhallan R. S., Macke J. P., Eddy R. L., Shows T. B., Reed R. R., Yau K. W., Nathans J. Human rod photoreceptor cGMP-gated channel: amino acid sequence, gene structure, and functional expression. J Neurosci. 1992 Aug;12(8):3248–3256. doi: 10.1523/JNEUROSCI.12-08-03248.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Dhallan R. S., Yau K. W., Schrader K. A., Reed R. R. Primary structure and functional expression of a cyclic nucleotide-activated channel from olfactory neurons. Nature. 1990 Sep 13;347(6289):184–187. doi: 10.1038/347184a0. [DOI] [PubMed] [Google Scholar]
  15. Ding C., Potter E. D., Qiu W., Coon S. L., Levine M. A., Guggino S. E. Cloning and widespread distribution of the rat rod-type cyclic nucleotide-gated cation channel. Am J Physiol. 1997 Apr;272(4 Pt 1):C1335–C1344. doi: 10.1152/ajpcell.1997.272.4.C1335. [DOI] [PubMed] [Google Scholar]
  16. Distler M., Biel M., Flockerzi V., Hofmann F. Expression of cyclic nucleotide-gated cation channels in non-sensory tissues and cells. Neuropharmacology. 1994 Nov;33(11):1275–1282. doi: 10.1016/0028-3908(94)90027-2. [DOI] [PubMed] [Google Scholar]
  17. Dryer S. E., Henderson D. A cyclic GMP-activated channel in dissociated cells of the chick pineal gland. Nature. 1991 Oct 24;353(6346):756–758. doi: 10.1038/353756a0. [DOI] [PubMed] [Google Scholar]
  18. Fesenko E. E., Kolesnikov S. S., Lyubarsky A. L. Induction by cyclic GMP of cationic conductance in plasma membrane of retinal rod outer segment. Nature. 1985 Jan 24;313(6000):310–313. doi: 10.1038/313310a0. [DOI] [PubMed] [Google Scholar]
  19. Frings S., Lynch J. W., Lindemann B. Properties of cyclic nucleotide-gated channels mediating olfactory transduction. Activation, selectivity, and blockage. J Gen Physiol. 1992 Jul;100(1):45–67. doi: 10.1085/jgp.100.1.45. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Gordon S. E., Zagotta W. N. Localization of regions affecting an allosteric transition in cyclic nucleotide-activated channels. Neuron. 1995 Apr;14(4):857–864. doi: 10.1016/0896-6273(95)90229-5. [DOI] [PubMed] [Google Scholar]
  21. Goulding E. H., Ngai J., Kramer R. H., Colicos S., Axel R., Siegelbaum S. A., Chess A. Molecular cloning and single-channel properties of the cyclic nucleotide-gated channel from catfish olfactory neurons. Neuron. 1992 Jan;8(1):45–58. doi: 10.1016/0896-6273(92)90107-o. [DOI] [PubMed] [Google Scholar]
  22. Goulding E. H., Tibbs G. R., Siegelbaum S. A. Molecular mechanism of cyclic-nucleotide-gated channel activation. Nature. 1994 Nov 24;372(6504):369–374. doi: 10.1038/372369a0. [DOI] [PubMed] [Google Scholar]
  23. Karlson K. H., Ciampolillo-Bates F., McCoy D. E., Kizer N. L., Stanton B. A. Corrigendum to "Cloning of a cGMP-gated cation channel from mouse kidney inner medullary collecting duct" [Biochim. Biophys. Acta 1236 (1995) 197-200]. Biochim Biophys Acta. 1995 Sep 13;1238(2):197–197. doi: 10.1016/0005-2736(95)00131-l. [DOI] [PubMed] [Google Scholar]
  24. Kaupp U. B., Niidome T., Tanabe T., Terada S., Bönigk W., Stühmer W., Cook N. J., Kangawa K., Matsuo H., Hirose T. Primary structure and functional expression from complementary DNA of the rod photoreceptor cyclic GMP-gated channel. Nature. 1989 Dec 14;342(6251):762–766. doi: 10.1038/342762a0. [DOI] [PubMed] [Google Scholar]
  25. Kaupp U. B. The cyclic nucleotide-gated channels of vertebrate photoreceptors and olfactory epithelium. Trends Neurosci. 1991 Apr;14(4):150–157. doi: 10.1016/0166-2236(91)90087-b. [DOI] [PubMed] [Google Scholar]
  26. Kawai F., Sterling P. AMPA receptor activates a G-protein that suppresses a cGMP-gated current. J Neurosci. 1999 Apr 15;19(8):2954–2959. doi: 10.1523/JNEUROSCI.19-08-02954.1999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Kingston P. A., Zufall F., Barnstable C. J. Rat hippocampal neurons express genes for both rod retinal and olfactory cyclic nucleotide-gated channels: novel targets for cAMP/cGMP function. Proc Natl Acad Sci U S A. 1996 Sep 17;93(19):10440–10445. doi: 10.1073/pnas.93.19.10440. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Komatsu H., Mori I., Rhee J. S., Akaike N., Ohshima Y. Mutations in a cyclic nucleotide-gated channel lead to abnormal thermosensation and chemosensation in C. elegans. Neuron. 1996 Oct;17(4):707–718. doi: 10.1016/s0896-6273(00)80202-0. [DOI] [PubMed] [Google Scholar]
  29. Kramer R. H., Tibbs G. R. Antagonists of cyclic nucleotide-gated channels and molecular mapping of their site of action. J Neurosci. 1996 Feb 15;16(4):1285–1293. doi: 10.1523/JNEUROSCI.16-04-01285.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Kumar V. D., Weber I. T. Molecular model of the cyclic GMP-binding domain of the cyclic GMP-gated ion channel. Biochemistry. 1992 May 19;31(19):4643–4649. doi: 10.1021/bi00134a015. [DOI] [PubMed] [Google Scholar]
  31. Leinders-Zufall T., Rosenboom H., Barnstable C. J., Shepherd G. M., Zufall F. A calcium-permeable cGMP-activated cation conductance in hippocampal neurons. Neuroreport. 1995 Sep 11;6(13):1761–1765. doi: 10.1097/00001756-199509000-00013. [DOI] [PubMed] [Google Scholar]
  32. Ludwig J., Margalit T., Eismann E., Lancet D., Kaupp U. B. Primary structure of cAMP-gated channel from bovine olfactory epithelium. FEBS Lett. 1990 Sep 17;270(1-2):24–29. doi: 10.1016/0014-5793(90)81226-e. [DOI] [PubMed] [Google Scholar]
  33. Mancuso C., Preziosi P., Grossman A. B., Navarra P. The role of carbon monoxide in the regulation of neuroendocrine function. Neuroimmunomodulation. 1997 Sep-Dec;4(5-6):225–229. doi: 10.1159/000097340. [DOI] [PubMed] [Google Scholar]
  34. McCoy D. E., Guggino S. E., Stanton B. A. The renal cGMP-gated cation channel: its molecular structure and physiological role. Kidney Int. 1995 Oct;48(4):1125–1133. doi: 10.1038/ki.1995.396. [DOI] [PubMed] [Google Scholar]
  35. McKay D. B., Weber I. T., Steitz T. A. Structure of catabolite gene activator protein at 2.9-A resolution. Incorporation of amino acid sequence and interactions with cyclic AMP. J Biol Chem. 1982 Aug 25;257(16):9518–9524. [PubMed] [Google Scholar]
  36. Nakamura T., Gold G. H. A cyclic nucleotide-gated conductance in olfactory receptor cilia. 1987 Jan 29-Feb 4Nature. 325(6103):442–444. doi: 10.1038/325442a0. [DOI] [PubMed] [Google Scholar]
  37. Nawy S., Jahr C. E. cGMP-gated conductance in retinal bipolar cells is suppressed by the photoreceptor transmitter. Neuron. 1991 Oct;7(4):677–683. doi: 10.1016/0896-6273(91)90380-i. [DOI] [PubMed] [Google Scholar]
  38. Nawy S. The metabotropic receptor mGluR6 may signal through G(o), but not phosphodiesterase, in retinal bipolar cells. J Neurosci. 1999 Apr 15;19(8):2938–2944. doi: 10.1523/JNEUROSCI.19-08-02938.1999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Parent A., Schrader K., Munger S. D., Reed R. R., Linden D. J., Ronnett G. V. Synaptic transmission and hippocampal long-term potentiation in olfactory cyclic nucleotide-gated channel type 1 null mouse. J Neurophysiol. 1998 Jun;79(6):3295–3301. doi: 10.1152/jn.1998.79.6.3295. [DOI] [PubMed] [Google Scholar]
  40. Park C. S., MacKinnon R. Divalent cation selectivity in a cyclic nucleotide-gated ion channel. Biochemistry. 1995 Oct 17;34(41):13328–13333. doi: 10.1021/bi00041a008. [DOI] [PubMed] [Google Scholar]
  41. Pittler S. J., Lee A. K., Altherr M. R., Howard T. A., Seldin M. F., Hurwitz R. L., Wasmuth J. J., Baehr W. Primary structure and chromosomal localization of human and mouse rod photoreceptor cGMP-gated cation channel. J Biol Chem. 1992 Mar 25;267(9):6257–6262. [PubMed] [Google Scholar]
  42. Santy L. C., Guidotti G. Reconstitution and characterization of two forms of cyclic nucleotide-gated channels from skeletal muscle. Am J Physiol. 1996 Dec;271(6 Pt 1):E1051–E1060. doi: 10.1152/ajpendo.1996.271.6.E1051. [DOI] [PubMed] [Google Scholar]
  43. Sautter A., Biel M., Hofmann F. Molecular cloning of cyclic nucleotide-gated cation channel subunits from rat pineal gland. Brain Res Mol Brain Res. 1997 Aug;48(1):171–175. doi: 10.1016/s0169-328x(97)00155-1. [DOI] [PubMed] [Google Scholar]
  44. Savchenko A., Barnes S., Kramer R. H. Cyclic-nucleotide-gated channels mediate synaptic feedback by nitric oxide. Nature. 1997 Dec 18;390(6661):694–698. doi: 10.1038/37803. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Scott S. P., Harrison R. W., Weber I. T., Tanaka J. C. Predicted ligand interactions of 3'5'-cyclic nucleotide-gated channel binding sites: comparison of retina and olfactory binding site models. Protein Eng. 1996 Apr;9(4):333–344. doi: 10.1093/protein/9.4.333. [DOI] [PubMed] [Google Scholar]
  46. Scott S. P., Tanaka J. C. Molecular interactions of 3',5'-cyclic purine analogues with the binding site of retinal rod ion channels. Biochemistry. 1995 Feb 21;34(7):2338–2347. doi: 10.1021/bi00007a030. [DOI] [PubMed] [Google Scholar]
  47. Scott S. P., Tanaka J. C. Three residues predicted by molecular modeling to interact with the purine moiety alter ligand binding and channel gating in cyclic nucleotide-gated channels. Biochemistry. 1998 Dec 8;37(49):17239–17252. doi: 10.1021/bi981185d. [DOI] [PubMed] [Google Scholar]
  48. Tanaka J. C., Eccleston J. F., Furman R. E. Photoreceptor channel activation by nucleotide derivatives. Biochemistry. 1989 Apr 4;28(7):2776–2784. doi: 10.1021/bi00433a006. [DOI] [PubMed] [Google Scholar]
  49. Varnum M. D., Black K. D., Zagotta W. N. Molecular mechanism for ligand discrimination of cyclic nucleotide-gated channels. Neuron. 1995 Sep;15(3):619–625. doi: 10.1016/0896-6273(95)90150-7. [DOI] [PubMed] [Google Scholar]
  50. Veske A., Nilsson S. E., Gal A. Characterization of canine rod photoreceptor cGMP-gated cation channel alpha-subunit gene and exclusion of its involvement in the hereditary retinal dystrophy of Swedish Briards. Gene. 1997 Nov 20;202(1-2):115–119. doi: 10.1016/s0378-1119(97)00461-7. [DOI] [PubMed] [Google Scholar]
  51. Weber I. T., Harrison R. W. Molecular mechanics calculations on HIV-1 protease with peptide substrates correlate with experimental data. Protein Eng. 1996 Aug;9(8):679–690. doi: 10.1093/protein/9.8.679. [DOI] [PubMed] [Google Scholar]
  52. Weber I. T., Steitz T. A. Structure of a complex of catabolite gene activator protein and cyclic AMP refined at 2.5 A resolution. J Mol Biol. 1987 Nov 20;198(2):311–326. doi: 10.1016/0022-2836(87)90315-9. [DOI] [PubMed] [Google Scholar]
  53. Wei J. Y., Cohen E. D., Yan Y. Y., Genieser H. G., Barnstable C. J. Identification of competitive antagonists of the rod photoreceptor cGMP-gated cation channel: beta-phenyl-1,N2-etheno-substituted cGMP analogues as probes of the cGMP-binding site. Biochemistry. 1996 Dec 24;35(51):16815–16823. doi: 10.1021/bi961763v. [DOI] [PubMed] [Google Scholar]
  54. Wei J. Y., Roy D. S., Leconte L., Barnstable C. J. Molecular and pharmacological analysis of cyclic nucleotide-gated channel function in the central nervous system. Prog Neurobiol. 1998 Oct;56(1):37–64. doi: 10.1016/s0301-0082(98)00029-x. [DOI] [PubMed] [Google Scholar]
  55. Weyand I., Godde M., Frings S., Weiner J., Müller F., Altenhofen W., Hatt H., Kaupp U. B. Cloning and functional expression of a cyclic-nucleotide-gated channel from mammalian sperm. Nature. 1994 Apr 28;368(6474):859–863. doi: 10.1038/368859a0. [DOI] [PubMed] [Google Scholar]
  56. Yu W. P., Grunwald M. E., Yau K. W. Molecular cloning, functional expression and chromosomal localization of a human homolog of the cyclic nucleotide-gated ion channel of retinal cone photoreceptors. FEBS Lett. 1996 Sep 16;393(2-3):211–215. doi: 10.1016/0014-5793(96)00889-7. [DOI] [PubMed] [Google Scholar]
  57. Zagotta W. N., Siegelbaum S. A. Structure and function of cyclic nucleotide-gated channels. Annu Rev Neurosci. 1996;19:235–263. doi: 10.1146/annurev.ne.19.030196.001315. [DOI] [PubMed] [Google Scholar]
  58. Zimmerman A. L., Yamanaka G., Eckstein F., Baylor D. A., Stryer L. Interaction of hydrolysis-resistant analogs of cyclic GMP with the phosphodiesterase and light-sensitive channel of retinal rod outer segments. Proc Natl Acad Sci U S A. 1985 Dec;82(24):8813–8817. doi: 10.1073/pnas.82.24.8813. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Zong X., Zucker H., Hofmann F., Biel M. Three amino acids in the C-linker are major determinants of gating in cyclic nucleotide-gated channels. EMBO J. 1998 Jan 15;17(2):353–362. doi: 10.1093/emboj/17.2.353. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. Zufall F., Firestein S., Shepherd G. M. Cyclic nucleotide-gated ion channels and sensory transduction in olfactory receptor neurons. Annu Rev Biophys Biomol Struct. 1994;23:577–607. doi: 10.1146/annurev.bb.23.060194.003045. [DOI] [PubMed] [Google Scholar]
  61. Zufall F., Shepherd G. M., Barnstable C. J. Cyclic nucleotide gated channels as regulators of CNS development and plasticity. Curr Opin Neurobiol. 1997 Jun;7(3):404–412. doi: 10.1016/s0959-4388(97)80070-0. [DOI] [PubMed] [Google Scholar]
  62. de la Villa P., Kurahashi T., Kaneko A. L-glutamate-induced responses and cGMP-activated channels in three subtypes of retinal bipolar cells dissociated from the cat. J Neurosci. 1995 May;15(5 Pt 1):3571–3582. doi: 10.1523/JNEUROSCI.15-05-03571.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]

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