May 2004
Volume 45, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2004
Role of Farnesylation of Transducin in Rod Photoreceptor Cells
Author Affiliations & Notes
  • Y. Fukada
    Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, Tokyo, Japan
  • K. Hagiwara
    Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, Tokyo, Japan
  • M. Katadae
    Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, Tokyo, Japan
  • P.J. Casey
    Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, NC
  • A. Wada
    Department of Analytical Chemistry, Kobe Pharmaceutical University, Kobe, Japan
  • M. Ito
    Department of Analytical Chemistry, Kobe Pharmaceutical University, Kobe, Japan
  • Footnotes
    Commercial Relationships  Y. Fukada, None; K. Hagiwara, None; M. Katadae, None; P.J. Casey, None; A. Wada, None; M. Ito, None.
  • Footnotes
    Support  HFSP Grant RGP0003/2003; Grant–in–Aid for Scientific Research (S) from JSPS
Investigative Ophthalmology & Visual Science May 2004, Vol.45, 1272. doi:
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      Y. Fukada, K. Hagiwara, M. Katadae, P.J. Casey, A. Wada, M. Ito; Role of Farnesylation of Transducin in Rod Photoreceptor Cells . Invest. Ophthalmol. Vis. Sci. 2004;45(13):1272.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Abstract: : Purpose: Farnesylation of the γ–subunit of the retinal G–protein, transducin (Tα/Tßγ), is indispensable for light–initiated signaling in rod photoreceptor cells. However, the farnesyl–mediated molecular interactions important for signaling are not well understood. The present study aimed at deciphering the role of the farnesyl in protein–protein and protein–membrane interactions in visual transduction process in rod photoreceptor cells. Methods: We created a functional Tßγ analog in which the farnesyl was replaced by (3–azidophenoxy)geranyl (POG), a novel farnesyl analog with a distal photoreactive azido group that is activatable with UV–irradiation. Results: In the presence of Tα–GDP and/or lipid membranes, UV–irradiation of POG–modified Tßγ (POG–Tßγ) invariably yielded a cross–linked product of Tγ–Tß, reflecting a constitutive interaction of the Tγ C–terminal lipid with Tß. In addition to Tγ–Tß adduct, a Tγ–Tα cross–link was detected in the aqueous fraction. Reconstitution of POG–Tßγ with Tα and light–activated rhodopsin (Rh*) in photoreceptor membranes resulted in cross–linking of Tγ with a glycerophospholipid, indicating molecular interaction of the farnesyl with cellular membranes. The Tγ–phospholipid cross–link was observed only in the presence of both Tα–GDP and Rh*, and was abolished by the addition of GTPγS or by replacing Rh* with opsin. These findings suggest a transient farnesyl–membrane interaction occurs only in a signaling state formed in a transducin–Rh* ternary complex. On the other hand, UV–irradiation of POG–Tßγ in a soluble complex with phosducin, a negative regulator of G–protein, yielded a Tγ –phosducin adduct in addition to the Tγ–Tß cross–link. Conclusions: The results obtained in the present study illustrate that, rather than being a static membrane anchor, the farnesyl moiety plays an active role in the dynamics of protein–protein and protein–membrane interactions at defined steps in the signal transduction process.

Keywords: photoreceptors • signal transduction • lipids 
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