title: Activity-dependent modulation of rod photoreceptor cyclic nucleotide-gated channels mediated by phosphorylation of a specific tyrosine residue.
abstract: Cyclic nucleotide-gated (CNG) channels are crucial for phototransduction in vertebrate rod photoreceptors. The cGMP sensitivity of these channels is modulated by diffusible intracellular messengers, including Ca2+/calmodulin, contributing to negative feedback during sensory adaptation. Membrane-associated protein tyrosine kinases and phosphatases also modulate rod CNG channels, but whether this results from direct changes in the phosphorylation state of the channel protein has been unclear. Here, we show that bovine rod CNG channel alpha-subunits (bRET) contain a tyrosine phosphorylation site crucial for modulation. bRET channels expressed in Xenopus oocytes exhibit modulation, whereas rat olfactory CNG channels (rOLF) do not. Chimeric channels reveal that differences in the C terminus, containing the cyclic nucleotide-binding domain, account for this difference. One specific tyrosine in bRET (Y498) appears to be crucial; replacement of this tyrosine in bRET curtails modulation, whereas installation into rOLF confers modulability. As the channel becomes dephosphorylated, there is an increase in the rate of spontaneous openings in the absence of ligand, indicating that changes in the phosphorylation state affect the allosteric gating equilibrium. Moreover, we find that dephosphorylation, which favors channel opening, requires open channels, whereas phosphorylation, which promotes channel closing, requires closed channels. Hence, modulation by changes in tyrosine phosphorylation is activity-dependent and may constitute a positive feedback mechanism, contrasting with negative feedback systems underlying adaptation.
authors.affiliation: Department of Molecular and Cellular Pharmacology, University of Miami School of Medicine, Miami, Florida 33101, USA.
authors.forename: E
authors.initials: E
authors.lastname: Molokanova
authors.forename: F
authors.initials: F
authors.lastname: Maddox
authors.forename: C W
authors.initials: CW
authors.lastname: Luetje
authors.forename: R H
authors.initials: RH
authors.lastname: Kramer
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D000494:Allosteric Regulation; D000818:Animals; D002417:Cattle; D000242:Cyclic AMP / Q000494:pharmacology; D006152:Cyclic GMP / Q000494:pharmacology; D054815:Cyclic Nucleotide-Gated Cation Channels; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology*; D007473:Ion Channels / Q000378:metabolism* / Q000502:physiology; D009865:Oocytes / Q000502:physiology; D018408:Patch-Clamp Techniques; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D017948:Retinal Rod Photoreceptor Cells / Q000737:chemistry / Q000201:enzymology*; D014443:Tyrosine / Q000378:metabolism; D014785:Vision, Ocular / Q000502:physiology*; D014981:Xenopus
chemical_list: D054815:Cyclic Nucleotide-Gated Cation Channels; D007473:Ion Channels; D014443:Tyrosine; D000255:Adenosine Triphosphate; D000242:Cyclic AMP; D011505:Protein-Tyrosine Kinases; D006152:Cyclic GMP