Responses of the phototransduction cascade to dim light.
The biochemistry of visual excitation is kinetically explored by measuring the activity of the cGMP phosphodiesterase (PDE) at light levels that activate only a few tens of rhodopsin molecules per rod. At 23 degrees C and in the presence of ATP, the pulse of PDE activity lasts 4 s (full width at half maximum). Complementing the rod outer segments (ROS) with rhodopsin kinase (RK) and arrestin or its splice variant p44 does not significantly shorten the pulse. But when the ROS are washed, the duration of the signal doubles. Adding either arrestin or p44 back to washed ROS approximately restores the pulse width to its initial value, with p44 being 10 times more efficient than arrestin. This supports the idea that, in vivo, capping of phosphorylated R* is mostly done by p44. When myristoylated (14:0) recoverin is added to unwashed ROS, the pulse duration and amplitude increase by about 50% if the free calcium is 500 nM. This effect increases further if the calcium is raised to 1 microM. Whenever R* deactivation is changed--when RK is exogenously enriched or when ATP is omitted from the buffer--there is no impact on the rising slope of the PDE pulse but only on its amplitude and duration. We explain this effect as due to the unequal competition between transducin and RK for R*. The kinetic model issued from this idea fits the data well, and its prediction that enrichment with transducin should lengthen the PDE pulse is successfully validated.
Langlois
G
G
Institut de Pharmacologie Moléculaire et Cellulaire, Centre National de la Recherche Scientifique, Valbonne, France.
Chen
C K
CK
Palczewski
K
K
Hurley
J B
JB
Vuong
T M
TM
D015106:3',5'-Cyclic-GMP Phosphodiesterases / Q000378:metabolism; D000818:Animals; D000941:Antigens / Q000235:genetics / Q000378:metabolism; D019393:Arrestin; D001667:Binding, Competitive; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000378:metabolism; D002417:Cattle; D003623:Dark Adaptation / Q000502:physiology; D005136:Eye Proteins / Q000235:genetics / Q000378:metabolism; D051606:G-Protein-Coupled Receptor Kinase 1; D014644:Genetic Variation; D051597:Hippocalcin; D066298:In Vitro Techniques; D007700:Kinetics; D008027:Light; D008074:Lipoproteins; D008954:Models, Biological; D009419:Nerve Tissue Proteins; D018462:Photobiology; D011494:Protein Kinases / Q000378:metabolism; D012326:RNA Splicing; D051603:Recoverin; D017948:Retinal Rod Photoreceptor Cells / Q000502:physiology* / Q000528:radiation effects*; D012243:Rhodopsin / Q000378:metabolism / Q000528:radiation effects
D000941:Antigens; D019393:Arrestin; D002135:Calcium-Binding Proteins; D005136:Eye Proteins; D008074:Lipoproteins; D009419:Nerve Tissue Proteins; D051603:Recoverin; D051597:Hippocalcin; D012243:Rhodopsin; D011494:Protein Kinases; D051606:G-Protein-Coupled Receptor Kinase 1; D015106:3',5'-Cyclic-GMP Phosphodiesterases; D002118:Calcium