title: Effect of the ILE86TER mutation in the γ subunit of cGMP phosphodiesterase (PDE6) on rod photoreceptor signaling.
abstract: The light-dependent decrease in cyclic guanosine monophosphate (cGMP) in the rod outer segment is produced by a phosphodiesterase (PDE6), consisting of catalytic α and β subunits and two inhibitory γ subunits. The molecular mechanism of PDE6γ regulation of the catalytic subunits is uncertain. To study this mechanism in vivo, we introduced a modified Pde6g gene for PDE6γ into a line of Pde6g(tm1)/Pde6g(tm1) mice that do not express PDE6γ. The resulting ILE86TER mice have a PDE6γ that lacks the two final carboxyl-terminal Ile(86) and Ile(87) residues, a mutation previously shown in vitro to reduce inhibition by PDE6γ. ILE86TER rods showed a decreased sensitivity and rate of activation, probably the result of a decreased level of expression of PDE6 in ILE86TER rods. More importantly, they showed a decreased rate of decay of the photoresponse, consistent with decreased inhibition of PDE6 α and β by PDE6γ. Furthermore, ILE86TER rods had a higher rate of spontaneous activation of PDE6 than WT rods. Circulating current in ILE86TER rods that also lacked both guanylyl cyclase activating proteins (GCAPs) could be increased several fold by perfusion with 100μM of the PDE6 inhibitor 3-isobutyl-1-methylxanthine (IBMX), consistent with a higher rate of dark PDE6 activity in the mutant photoreceptors. In contrast, IBMX had little effect on the circulating current of WT rods, unlike previous results from amphibians. Our results show for the first time that the Ile(86) and Ile(87) residues are necessary for normal inhibition of PDE6 catalytic activity in vivo, and that increased basal activity of PDE can be partially compensated by GCAP-dependent regulation of guanylyl cyclase.
authors.affiliation: Department of Pathology & Cell Biology, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA. sht2@columbia.edu
authors.forename: Stephen H
authors.initials: SH
authors.lastname: Tsang
authors.forename: Michael L
authors.initials: ML
authors.lastname: Woodruff
authors.forename: Chyuan-Sheng
authors.initials: CS
authors.lastname: Lin
authors.forename: Barry D
authors.initials: BD
authors.lastname: Jacobson
authors.forename: Matthew C
authors.initials: MC
authors.lastname: Naumann
authors.forename: Chun Wei
authors.initials: CW
authors.lastname: Hsu
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Davis
authors.forename: Marianne C
authors.initials: MC
authors.lastname: Cilluffo
authors.forename: Jeannie
authors.initials: J
authors.lastname: Chen
authors.forename: Gordon L
authors.initials: GL
authors.lastname: Fain
mesh_terms: D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000465:Algorithms; D000818:Animals; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6 / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D005136:Eye Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D005260:Female; D007700:Kinetics; D008027:Light; D008297:Male; D008564:Membrane Potentials / Q000528:radiation effects; D051379:Mice; D008810:Mice, Inbred C57BL; D008811:Mice, Inbred DBA; D008822:Mice, Transgenic; D012160:Retina / Q000378:metabolism / Q000473:pathology; D012162:Retinal Degeneration / Q000235:genetics / Q000473:pathology; D017948:Retinal Rod Photoreceptor Cells / Q000187:drug effects / Q000378:metabolism*; D017384:Sequence Deletion; D015398:Signal Transduction
chemical_list: D005136:Eye Proteins; D054707:Cyclic Nucleotide Phosphodiesterases, Type 6; C498747:Pde6a protein, mouse; C517438:Pde6b protein, mouse; C517444:Pde6g protein, mouse; D015056:1-Methyl-3-isobutylxanthine