Light-induced translocation of cyclic-GMP phosphodiesterase on rod disc membranes in rat retina.
OBJECTIVE
Cyclic GMP phosphodiesterase (PDE) is the light-regulated effector enzyme of vertebrate rods. Upon photo-activation of rhodopsin followed by activation of transducin/GTP, PDE rapidly hydrolyzes 3', 5'-cyclic GMP (cGMP) to 5'-GMP, which results in closure of cGMP-dependent ion channels and generation of a nerve signal. In the rod photoreceptors, PDE is entirely localized within the rod outer segment (ROS), a specialized compartment consisting of thousands of disc stacks. This study investigated the effects of light on the subcellular localization of PDE in ROS.


METHODS
Adult rats were either dark- or light-adapted for various durations before eyes were isolated and processed for transmission electron microscopy. Immunogold electron microscopy was performed with antibodies against PDE. Lateral displacement of PDE on ROS disc membrane was analyzed from electron micrographs. PDE enzymatic activities were measured with thin layer chromatography.


RESULTS
Light exposure induced translocation of PDE away from the edges of the dark-adapted disc membranes adjacent to the ROS plasma membrane. In dark-adapted ROS, a substantial portion (19+/-2%) of total PDE was localized near the edges of the disc membranes. Within 1 min of light exposure in the presence of GTP, over half of such PDE molecules (10+/-1% of total PDE) had moved away from the edges of the discs toward disc center. This light induced translocation of PDE was GTP dependent, as the effect was abolished when hydrolysis-resistant GTPgammaS was used in place of GTP. The percentage of PDE found near the disc edge corresponds to the fraction of PDE activity relative to maximal PDE activity revealed by limited trypsin proteolysis.


CONCLUSIONS
These results suggest that light and GTP modulates lateral displacement of PDE, which might contribute to light-induced reduction of rod photoreceptor sensitivity.
Chen
Jing
J
Department of Biomedical Engineering, College of Engineering, Boston University, Boston, MA, USA. jing.chen@childrens.harvard.edu
Yoshida
Tatsuro
T
Bitensky
Mark W
MW
D015106:3',5'-Cyclic-GMP Phosphodiesterases / Q000378:metabolism*; D000818:Animals; D000906:Antibodies; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology* / Q000528:radiation effects / Q000648:ultrastructure; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6 / Q000378:metabolism; D005260:Female; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D007150:Immunohistochemistry; D008027:Light; D055537:Light Signal Transduction / Q000187:drug effects / Q000528:radiation effects*; D010539:Permeability / Q000187:drug effects / Q000528:radiation effects; D021381:Protein Transport / Q000187:drug effects / Q000528:radiation effects; D051381:Rats; D020318:Rats, Long-Evans; D017382:Reactive Oxygen Species / Q000302:isolation & purification; D015203:Reproducibility of Results; D017948:Retinal Rod Photoreceptor Cells / Q000166:cytology* / Q000201:enzymology* / Q000528:radiation effects / Q000648:ultrastructure; D013997:Time Factors
D000906:Antibodies; D017382:Reactive Oxygen Species; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D015106:3',5'-Cyclic-GMP Phosphodiesterases; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6