Skip to main content
The EMBO Journal logoLink to The EMBO Journal
. 1996 Dec 16;15(24):7036–7045.

The transient receptor potential protein (Trp), a putative store-operated Ca2+ channel essential for phosphoinositide-mediated photoreception, forms a signaling complex with NorpA, InaC and InaD.

A Huber 1, P Sander 1, A Gobert 1, M Bähner 1, R Hermann 1, R Paulsen 1
PMCID: PMC452529  PMID: 9003779

Abstract

The transient receptor potential protein (Trp) is a putative capacitative Ca2+ entry channel present in fly photoreceptors, which use the inositol 1,4,5-trisphosphate (InsP3) signaling pathway for phototransduction. By immunoprecipitation studies, we find that Trp is associated into a multiprotein complex with the norpA-encoded phospholipase C, an eye-specific protein kinase C (InaC) and with the InaD protein (InaD). InaD is a putative substrate of InaC and contains two PDZ repeats, putative protein-protein interaction domains. These proteins are present in the photoreceptor membrane at about equimolar ratios. The Trp homolog analyzed here is isolated together with NorpA, InaC and InaD from blowfly (Calliphora) photoreceptors. Compared to Drosophila Trp, the Calliphora Trp homolog displays 77% amino acid identity. The highest sequence conservation is found in the region that contains the putative transmembrane domains S1-S6 (91% amino acid identity). As investigated by immunogold labeling with specific antibodies directed against Trp and InaD, the Trp signaling complex is located in the microvillar membranes of the photoreceptor cells. The spatial distribution of the signaling complex argues against a direct conformational coupling of Trp to an InsP3 receptor supposed to be present in the membrane of internal photoreceptor Ca2+ stores. It is suggested that the organization of signal transducing proteins into a multiprotein complex provides the structural basis for an efficient and fast activation and regulation of Ca2+ entry through the Trp channel.

Full text

PDF
7036

Images in this article

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bennett D. L., Petersen C. C., Cheek T. R. Calcium signalling. Cracking ICRAC in the eye. Curr Biol. 1995 Nov 1;5(11):1225–1228. doi: 10.1016/s0960-9822(95)00243-0. [DOI] [PubMed] [Google Scholar]
  2. Bennett V. Ankyrins. Adaptors between diverse plasma membrane proteins and the cytoplasm. J Biol Chem. 1992 May 5;267(13):8703–8706. [PubMed] [Google Scholar]
  3. Bentrop J., Paulsen R. Light-modulated ADP-ribosylation, protein phosphorylation and protein binding in isolated fly photoreceptor membranes. Eur J Biochem. 1986 Nov 17;161(1):61–67. doi: 10.1111/j.1432-1033.1986.tb10124.x. [DOI] [PubMed] [Google Scholar]
  4. Berridge M. J. Capacitative calcium entry. Biochem J. 1995 Nov 15;312(Pt 1):1–11. doi: 10.1042/bj3120001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Berridge M. J. Inositol trisphosphate and calcium signalling. Nature. 1993 Jan 28;361(6410):315–325. doi: 10.1038/361315a0. [DOI] [PubMed] [Google Scholar]
  6. Bird G. S., Bian X., Putney J. W., Jr Calcium entry signal? Nature. 1995 Feb 9;373(6514):481–482. doi: 10.1038/373481b0. [DOI] [PubMed] [Google Scholar]
  7. Bloomquist B. T., Shortridge R. D., Schneuwly S., Perdew M., Montell C., Steller H., Rubin G., Pak W. L. Isolation of a putative phospholipase C gene of Drosophila, norpA, and its role in phototransduction. Cell. 1988 Aug 26;54(5):723–733. doi: 10.1016/s0092-8674(88)80017-5. [DOI] [PubMed] [Google Scholar]
  8. Cosens D. J., Manning A. Abnormal electroretinogram from a Drosophila mutant. Nature. 1969 Oct 18;224(5216):285–287. doi: 10.1038/224285a0. [DOI] [PubMed] [Google Scholar]
  9. Danscher G. Localization of gold in biological tissue. A photochemical method for light and electronmicroscopy. Histochemistry. 1981;71(1):81–88. doi: 10.1007/BF00592572. [DOI] [PubMed] [Google Scholar]
  10. Friel D. D. TRP: its role in phototransduction and store-operated Ca2+ entry. Cell. 1996 May 31;85(5):617–619. doi: 10.1016/s0092-8674(00)81021-1. [DOI] [PubMed] [Google Scholar]
  11. Hardie R. C., Minke B. Novel Ca2+ channels underlying transduction in Drosophila photoreceptors: implications for phosphoinositide-mediated Ca2+ mobilization. Trends Neurosci. 1993 Sep;16(9):371–376. doi: 10.1016/0166-2236(93)90095-4. [DOI] [PubMed] [Google Scholar]
  12. Hardie R. C., Minke B. Phosphoinositide-mediated phototransduction in Drosophila photoreceptors: the role of Ca2+ and trp. Cell Calcium. 1995 Oct;18(4):256–274. doi: 10.1016/0143-4160(95)90023-3. [DOI] [PubMed] [Google Scholar]
  13. Hardie R. C., Minke B. The trp gene is essential for a light-activated Ca2+ channel in Drosophila photoreceptors. Neuron. 1992 Apr;8(4):643–651. doi: 10.1016/0896-6273(92)90086-s. [DOI] [PubMed] [Google Scholar]
  14. Harteneck C., Obukhov A. G., Zobel A., Kalkbrenner F., Schultz G. The Drosophila cation channel trpl expressed in insect Sf9 cells is stimulated by agonists of G-protein-coupled receptors. FEBS Lett. 1995 Jan 30;358(3):297–300. doi: 10.1016/0014-5793(94)01455-a. [DOI] [PubMed] [Google Scholar]
  15. Heinemann S. H., Terlau H., Stühmer W., Imoto K., Numa S. Calcium channel characteristics conferred on the sodium channel by single mutations. Nature. 1992 Apr 2;356(6368):441–443. doi: 10.1038/356441a0. [DOI] [PubMed] [Google Scholar]
  16. Heukeshoven J., Dernick R. Improved silver staining procedure for fast staining in PhastSystem Development Unit. I. Staining of sodium dodecyl sulfate gels. Electrophoresis. 1988 Jan;9(1):28–32. doi: 10.1002/elps.1150090106. [DOI] [PubMed] [Google Scholar]
  17. Hochstrate P. Lanthanum mimicks the trp photoreceptor mutant of Drosophila in the blowfly Calliphora. J Comp Physiol A. 1989 Dec;166(2):179–187. doi: 10.1007/BF00193462. [DOI] [PubMed] [Google Scholar]
  18. Hotta Y., Benzer S. Genetic dissection of the Drosophila nervous system by means of mosaics. Proc Natl Acad Sci U S A. 1970 Nov;67(3):1156–1163. doi: 10.1073/pnas.67.3.1156. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hu Y., Schilling W. P. Receptor-mediated activation of recombinant Trpl expressed in Sf9 insect cells. Biochem J. 1995 Jan 15;305(Pt 2):605–611. doi: 10.1042/bj3050605. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hu Y., Vaca L., Zhu X., Birnbaumer L., Kunze D. L., Schilling W. P. Appearance of a novel Ca2+ influx pathway in Sf9 insect cells following expression of the transient receptor potential-like (trpl) protein of Drosophila. Biochem Biophys Res Commun. 1994 Jun 15;201(2):1050–1056. doi: 10.1006/bbrc.1994.1808. [DOI] [PubMed] [Google Scholar]
  21. Huber A., Sander P., Paulsen R. Phosphorylation of the InaD gene product, a photoreceptor membrane protein required for recovery of visual excitation. J Biol Chem. 1996 May 17;271(20):11710–11717. doi: 10.1074/jbc.271.20.11710. [DOI] [PubMed] [Google Scholar]
  22. Huber A., Sander P., Wolfrum U., Groell C., Gerdon G., Paulsen R. Isolation of genes encoding photoreceptor-specific proteins by immunoscreening with antibodies directed against purified blowfly rhabdoms. J Photochem Photobiol B. 1996 Aug;35(1-2):69–76. doi: 10.1016/1011-1344(96)07310-1. [DOI] [PubMed] [Google Scholar]
  23. Huber A., Smith D. P., Zuker C. S., Paulsen R. Opsin of Calliphora peripheral photoreceptors R1-6. Homology with Drosophila Rh1 and posttranslational processing. J Biol Chem. 1990 Oct 15;265(29):17906–17910. [PubMed] [Google Scholar]
  24. Irvine R. F. Inositol phosphates and Ca2+ entry: toward a proliferation or a simplification? FASEB J. 1992 Sep;6(12):3085–3091. doi: 10.1096/fasebj.6.12.1325932. [DOI] [PubMed] [Google Scholar]
  25. Jan L. Y., Jan Y. N. Potassium channels and their evolving gates. Nature. 1994 Sep 8;371(6493):119–122. doi: 10.1038/371119a0. [DOI] [PubMed] [Google Scholar]
  26. Joseph S. K., Samanta S. Detergent solubility of the inositol trisphosphate receptor in rat brain membranes. Evidence for association of the receptor with ankyrin. J Biol Chem. 1993 Mar 25;268(9):6477–6486. [PubMed] [Google Scholar]
  27. Kleuss C., Hescheler J., Ewel C., Rosenthal W., Schultz G., Wittig B. Assignment of G-protein subtypes to specific receptors inducing inhibition of calcium currents. Nature. 1991 Sep 5;353(6339):43–48. doi: 10.1038/353043a0. [DOI] [PubMed] [Google Scholar]
  28. Kleuss C., Scherübl H., Hescheler J., Schultz G., Wittig B. Different beta-subunits determine G-protein interaction with transmembrane receptors. Nature. 1992 Jul 30;358(6385):424–426. doi: 10.1038/358424a0. [DOI] [PubMed] [Google Scholar]
  29. Kleuss C., Scherübl H., Hescheler J., Schultz G., Wittig B. Selectivity in signal transduction determined by gamma subunits of heterotrimeric G proteins. Science. 1993 Feb 5;259(5096):832–834. doi: 10.1126/science.8094261. [DOI] [PubMed] [Google Scholar]
  30. Kurose H., Regan J. W., Caron M. G., Lefkowitz R. J. Functional interactions of recombinant alpha 2 adrenergic receptor subtypes and G proteins in reconstituted phospholipid vesicles. Biochemistry. 1991 Apr 2;30(13):3335–3341. doi: 10.1021/bi00227a024. [DOI] [PubMed] [Google Scholar]
  31. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  32. Masai I., Okazaki A., Hosoya T., Hotta Y. Drosophila retinal degeneration A gene encodes an eye-specific diacylglycerol kinase with cysteine-rich zinc-finger motifs and ankyrin repeats. Proc Natl Acad Sci U S A. 1993 Dec 1;90(23):11157–11161. doi: 10.1073/pnas.90.23.11157. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. McKay R. R., Miller K., Weckström M., Torkkeli P., Järvilehto M., Shortridge R. D. The rpa (receptor potential absent) visual mutant of the blowfly (Calliphora erythrocephala) is deficient in phospholipase C in the eye. J Neurogenet. 1994 Jul;9(3):177–187. doi: 10.3109/01677069409167278. [DOI] [PubMed] [Google Scholar]
  34. Montell C., Rubin G. M. Molecular characterization of the Drosophila trp locus: a putative integral membrane protein required for phototransduction. Neuron. 1989 Apr;2(4):1313–1323. doi: 10.1016/0896-6273(89)90069-x. [DOI] [PubMed] [Google Scholar]
  35. Neubig R. R. Membrane organization in G-protein mechanisms. FASEB J. 1994 Sep;8(12):939–946. doi: 10.1096/fasebj.8.12.8088459. [DOI] [PubMed] [Google Scholar]
  36. Niemeyer B. A., Suzuki E., Scott K., Jalink K., Zuker C. S. The Drosophila light-activated conductance is composed of the two channels TRP and TRPL. Cell. 1996 May 31;85(5):651–659. doi: 10.1016/s0092-8674(00)81232-5. [DOI] [PubMed] [Google Scholar]
  37. Pak W. L., Grossfield J., Arnold K. S. Mutants of the visual pathway of Drosophila melanogaster. Nature. 1970 Aug 1;227(5257):518–520. doi: 10.1038/227518b0. [DOI] [PubMed] [Google Scholar]
  38. Petersen C. C., Berridge M. J., Borgese M. F., Bennett D. L. Putative capacitative calcium entry channels: expression of Drosophila trp and evidence for the existence of vertebrate homologues. Biochem J. 1995 Oct 1;311(Pt 1):41–44. doi: 10.1042/bj3110041. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Phillips A. M., Bull A., Kelly L. E. Identification of a Drosophila gene encoding a calmodulin-binding protein with homology to the trp phototransduction gene. Neuron. 1992 Apr;8(4):631–642. doi: 10.1016/0896-6273(92)90085-r. [DOI] [PubMed] [Google Scholar]
  40. Plangger A., Malicki D., Whitney M., Paulsen R. Mechanism of arrestin 2 function in rhabdomeric photoreceptors. J Biol Chem. 1994 Oct 28;269(43):26969–26975. [PubMed] [Google Scholar]
  41. Pollock J. A., Assaf A., Peretz A., Nichols C. D., Mojet M. H., Hardie R. C., Minke B. TRP, a protein essential for inositide-mediated Ca2+ influx is localized adjacent to the calcium stores in Drosophila photoreceptors. J Neurosci. 1995 May;15(5 Pt 2):3747–3760. doi: 10.1523/JNEUROSCI.15-05-03747.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Randriamampita C., Tsien R. Y. Emptying of intracellular Ca2+ stores releases a novel small messenger that stimulates Ca2+ influx. Nature. 1993 Aug 26;364(6440):809–814. doi: 10.1038/364809a0. [DOI] [PubMed] [Google Scholar]
  43. Ranganathan R., Bacskai B. J., Tsien R. Y., Zuker C. S. Cytosolic calcium transients: spatial localization and role in Drosophila photoreceptor cell function. Neuron. 1994 Oct;13(4):837–848. doi: 10.1016/0896-6273(94)90250-x. [DOI] [PubMed] [Google Scholar]
  44. Ranganathan R., Harris G. L., Stevens C. F., Zuker C. S. A Drosophila mutant defective in extracellular calcium-dependent photoreceptor deactivation and rapid desensitization. Nature. 1991 Nov 21;354(6350):230–232. doi: 10.1038/354230a0. [DOI] [PubMed] [Google Scholar]
  45. Ranganathan R., Malicki D. M., Zuker C. S. Signal transduction in Drosophila photoreceptors. Annu Rev Neurosci. 1995;18:283–317. doi: 10.1146/annurev.ne.18.030195.001435. [DOI] [PubMed] [Google Scholar]
  46. Rodbell M. The role of GTP-binding proteins in signal transduction: from the sublimely simple to the conceptually complex. Curr Top Cell Regul. 1992;32:1–47. doi: 10.1016/b978-0-12-152832-4.50003-3. [DOI] [PubMed] [Google Scholar]
  47. Rubenstein R. C., Linder M. E., Ross E. M. Selectivity of the beta-adrenergic receptor among Gs, Gi's, and Go: assay using recombinant alpha subunits in reconstituted phospholipid vesicles. Biochemistry. 1991 Nov 5;30(44):10769–10777. doi: 10.1021/bi00108a023. [DOI] [PubMed] [Google Scholar]
  48. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Schaeffer E., Smith D., Mardon G., Quinn W., Zuker C. Isolation and characterization of two new drosophila protein kinase C genes, including one specifically expressed in photoreceptor cells. Cell. 1989 May 5;57(3):403–412. doi: 10.1016/0092-8674(89)90915-x. [DOI] [PubMed] [Google Scholar]
  50. Schneuwly S., Burg M. G., Lending C., Perdew M. H., Pak W. L. Properties of photoreceptor-specific phospholipase C encoded by the norpA gene of Drosophila melanogaster. J Biol Chem. 1991 Dec 25;266(36):24314–24319. [PubMed] [Google Scholar]
  51. Shieh B. H., Niemeyer B. A novel protein encoded by the InaD gene regulates recovery of visual transduction in Drosophila. Neuron. 1995 Jan;14(1):201–210. doi: 10.1016/0896-6273(95)90255-4. [DOI] [PubMed] [Google Scholar]
  52. Shieh B. H., Zhu M. Y. Regulation of the TRP Ca2+ channel by INAD in Drosophila photoreceptors. Neuron. 1996 May;16(5):991–998. doi: 10.1016/s0896-6273(00)80122-1. [DOI] [PubMed] [Google Scholar]
  53. Sinkins W. G., Vaca L., Hu Y., Kunze D. L., Schilling W. P. The COOH-terminal domain of Drosophila TRP channels confers thapsigargin sensitivity. J Biol Chem. 1996 Feb 9;271(6):2955–2960. doi: 10.1074/jbc.271.6.2955. [DOI] [PubMed] [Google Scholar]
  54. Smith D. P., Ranganathan R., Hardy R. W., Marx J., Tsuchida T., Zuker C. S. Photoreceptor deactivation and retinal degeneration mediated by a photoreceptor-specific protein kinase C. Science. 1991 Dec 6;254(5037):1478–1484. doi: 10.1126/science.1962207. [DOI] [PubMed] [Google Scholar]
  55. Studer D., Michel M., Müller M. High pressure freezing comes of age. Scanning Microsc Suppl. 1989;3:253–269. [PubMed] [Google Scholar]
  56. Vaca L., Sinkins W. G., Hu Y., Kunze D. L., Schilling W. P. Activation of recombinant trp by thapsigargin in Sf9 insect cells. Am J Physiol. 1994 Nov;267(5 Pt 1):C1501–C1505. doi: 10.1152/ajpcell.1994.267.5.C1501. [DOI] [PubMed] [Google Scholar]
  57. Wes P. D., Chevesich J., Jeromin A., Rosenberg C., Stetten G., Montell C. TRPC1, a human homolog of a Drosophila store-operated channel. Proc Natl Acad Sci U S A. 1995 Oct 10;92(21):9652–9656. doi: 10.1073/pnas.92.21.9652. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Wolfrum U. Centrin in the photoreceptor cells of mammalian retinae. Cell Motil Cytoskeleton. 1995;32(1):55–64. doi: 10.1002/cm.970320107. [DOI] [PubMed] [Google Scholar]
  59. Wong F., Schaefer E. L., Roop B. C., LaMendola J. N., Johnson-Seaton D., Shao D. Proper function of the Drosophila trp gene product during pupal development is important for normal visual transduction in the adult. Neuron. 1989 Jul;3(1):81–94. doi: 10.1016/0896-6273(89)90117-7. [DOI] [PubMed] [Google Scholar]
  60. Zhu L., McKay R. R., Shortridge R. D. Tissue-specific expression of phospholipase C encoded by the norpA gene of Drosophila melanogaster. J Biol Chem. 1993 Jul 25;268(21):15994–16001. [PubMed] [Google Scholar]
  61. Zhu X., Chu P. B., Peyton M., Birnbaumer L. Molecular cloning of a widely expressed human homologue for the Drosophila trp gene. FEBS Lett. 1995 Oct 16;373(3):193–198. doi: 10.1016/0014-5793(95)01038-g. [DOI] [PubMed] [Google Scholar]
  62. Zhu X., Jiang M., Peyton M., Boulay G., Hurst R., Stefani E., Birnbaumer L. trp, a novel mammalian gene family essential for agonist-activated capacitative Ca2+ entry. Cell. 1996 May 31;85(5):661–671. doi: 10.1016/s0092-8674(00)81233-7. [DOI] [PubMed] [Google Scholar]
  63. Zitt C., Zobel A., Obukhov A. G., Harteneck C., Kalkbrenner F., Lückhoff A., Schultz G. Cloning and functional expression of a human Ca2+-permeable cation channel activated by calcium store depletion. Neuron. 1996 Jun;16(6):1189–1196. doi: 10.1016/s0896-6273(00)80145-2. [DOI] [PubMed] [Google Scholar]
  64. Zuker C. S. The biology of vision of Drosophila. Proc Natl Acad Sci U S A. 1996 Jan 23;93(2):571–576. doi: 10.1073/pnas.93.2.571. [DOI] [PMC free article] [PubMed] [Google Scholar]
  65. Zweifach A., Lewis R. S. Rapid inactivation of depletion-activated calcium current (ICRAC) due to local calcium feedback. J Gen Physiol. 1995 Feb;105(2):209–226. doi: 10.1085/jgp.105.2.209. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES