The delta subunit of type 6 phosphodiesterase reduces light-induced cGMP hydrolysis in rod outer segments.
The delta subunit of the rod photoreceptor PDE has previously been shown to copurify with the soluble form of the enzyme and to solubilize the membrane-bound form (). To determine the physiological effect of the delta subunit on the light response of bovine rod outer segments, we measured the real time accumulation of the products of cGMP hydrolysis in a preparation of permeablized rod outer segments. The addition of delta subunit GST fusion protein (delta-GST) to this preparation caused a reduction in the maximal rate of cGMP hydrolysis in response to light. The maximal reduction of the light response was about 80%, and the half-maximal effect occurred at 385 nm delta subunit. Several experiments suggest that this effect was not due to the effects of delta-GST on transducin or rhodopsin kinase. Immunoblots demonstrated that exogenous delta-GST solubilized the majority of the PDE in ROS but did not affect the solubility of transducin. Therefore, changes in the solubility of transducin cannot account for the effects of delta-GST in the pH assay. The reduction in cGMP hydrolysis was independent of ATP, which indicates that it was not due to effects of delta-GST on rhodopsin kinase. In addition to the effect on cGMP hydrolysis, the delta-GST fusion protein slowed the turn-off of the system. This is probably due, at least in part, to an observed reduction in the GTPase rate of transducin in the presence of delta-GST. These results demonstrate that delta-GST can modify the activity of the phototransduction cascade in preparations of broken rod outer segments, probably due to a functional uncoupling of the transducin to PDE step of the signal transduction cascade and suggest that the delta subunit may play a similar role in the intact outer segment.
Cook
T A
TA
Department of Pharmacology, University of Washington, Seattle, WA 98195, USA.
Ghomashchi
F
F
Gelb
M H
MH
Florio
S K
SK
Beavo
J A
JA
D015106:3',5'-Cyclic-GMP Phosphodiesterases / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002417:Cattle; D006152:Cyclic GMP / Q000378:metabolism*; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6; D005136:Eye Proteins; D007700:Kinetics; D008027:Light; D010455:Peptides / Q000494:pharmacology; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins / Q000494:pharmacology; D012374:Rod Cell Outer Segment / Q000187:drug effects / Q000201:enzymology*; D012995:Solubility; D015293:Transducin / Q000378:metabolism; D014785:Vision, Ocular
D005136:Eye Proteins; D010455:Peptides; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins; D000255:Adenosine Triphosphate; D015106:3',5'-Cyclic-GMP Phosphodiesterases; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6; D015293:Transducin; D006152:Cyclic GMP