Characterization of Human Cone Phosphodiesterase-6 Ectopically Expressed in Xenopus laevis Rods

Hakim Muradov; Kimberly K Boyd; Mohammad Haeri; Vasily Kerov; Barry E Knox; Nikolai O Artemyev

doi:10.1074/jbc.M109.049916

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Characterization of Human Cone Phosphodiesterase-6 Ectopically Expressed in Xenopus laevis Rods

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Characterization of Human Cone Phosphodiesterase-6 Ectopically Expressed in Xenopus laevis Rods

Hakim Muradov, Kimberly K Boyd, Mohammad Haeri, Vasily Kerov, Barry E Knox and Nikolai O Artemyev

The Journal of biological chemistry, Vol.284(47), pp.32662-32669

11/20/2009

DOI: 10.1074/jbc.M109.049916

PMCID: PMC2781681

PMID: 19801642

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Abstract

PDE6 (phosphodiesterase-6) is the effector molecule in the vertebrate phototransduction cascade. Progress in understanding the structure and function of PDE6 has been hindered by lack of an expression system of the enzyme. Here we report ectopic expression and analysis of compartmentalization and membrane dynamics of the enhanced green fluorescent protein (EGFP) fusion protein of human cone PDE6C in rods of transgenic Xenopus laevis . EGFP-PDE6C is correctly targeted to the rod outer segments in transgenic Xenopus , where it displayed a characteristic striated pattern of EGFP fluorescence. Immunofluorescence labeling indicated significant and light-independent co-localization of EGFP-PDE6C with the disc rim marker peripherin-2 and endogenous frog PDE6. The diffusion of EGFP-PDE6C on disc membranes investigated with fluorescence recovery after photobleaching was markedly slower than theoretically predicted. The enzymatic characteristics of immunoprecipitated recombinant PDE6C were similar to known properties of the native bovine PDE6C. PDE6C was potently inhibited by the cone- and rod-specific PDE6 γ-subunits. Thus, transgenic Xenopus laevis is a unique expression system for PDE6 well suited for analysis of the mechanisms of visual diseases linked to PDE6 mutations.

Mechanisms of Signal Transduction

Details

Title: Subtitle: Characterization of Human Cone Phosphodiesterase-6 Ectopically Expressed in Xenopus laevis Rods
Creators: Hakim Muradov - From the
Kimberly K Boyd - From the
Mohammad Haeri - the Departments of
Vasily Kerov - From the
Barry E Knox - the Departments of
Nikolai O Artemyev - From the
Resource Type: Journal article
Publication Details: The Journal of biological chemistry, Vol.284(47), pp.32662-32669
DOI: 10.1074/jbc.M109.049916
PMID: 19801642
PMCID: PMC2781681
NLM abbreviation: J Biol Chem
ISSN: 0021-9258
eISSN: 1083-351X
Publisher: American Society for Biochemistry and Molecular Biology; 9650 Rockville Pike, Bethesda, MD 20814, U.S.A
Grant note: EY-10843 / National Institutes of Health
Language: English
Date published: 11/20/2009
Academic Unit: Molecular Physiology and Biophysics; Iowa Neuroscience Institute; Ophthalmology and Visual Sciences
Record Identifier: 9984070368002771

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