Light-dependent phosphorylation of the gamma subunit of cGMP-phophodiesterase (PDE6γ) at residue threonine 22 in intact photoreceptor neurons

• Published in: Biochemical and biophysical research communications 2009-12, Vol.390 (4), p.1149-1153
• Main Authors: Janisch, Kerstin M., Kasanuki, J. Mie, Naumann, Matthew C., Davis, Richard J., Lin, Chyuan-Sheng, Semple-Rowland, Susan, Tsang, Stephen H.
• Format: Article
• Language: English
• Subjects: a mouse line carrying a targeted disruption of the Pde6g gene; Animals; Antibodies, Phospho-Specific - immunology; Cattle; cGMP-phosphodiesterase; Cyclic Nucleotide Phosphodiesterases, Type 6 - metabolism; electroretinogram; ERG; gene encoding the γ subunit of the cGMP phosphodiesterase; GNAT1; GTP-Binding Protein alpha Subunits - metabolism; Light; Light Signal Transduction; Mice; Mice, Inbred Strains; Mice, Transgenic; PDE; Pde6gtm1/ Pde6gtm1; Pdeg; Phosphorylation; Protein Processing, Post-Translational; Retinal Cone Photoreceptor Cells - enzymology; Retinal Cone Photoreceptor Cells - radiation effects; Retinal Rod Photoreceptor Cells - enzymology; Retinal Rod Photoreceptor Cells - radiation effects; rod outer segments; ROS; Signal Transduction; T22A; T22A–T35A; Threonine - immunology; Threonine - metabolism; Transducin - metabolism; transducin alpha; transgenic mice which T22 and T35 were replaced with alanines; transgenic mice which T22 was replaced with alanine; transgenic mice which T35 was replaced with alanine
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1016/j.bbrc.2009.10.106

The γ subunit of rod-specific cGMP phosphodiesterase 6 (PDE6γ), an effector of the G-protein GNAT1, is a key regulator of phototransduction. The results of several in vitro biochemical reconstitution experiments conducted to examine the effects of phosphorylation of PDE6γ on its ability to regulate the PDE6 catalytic core have been inconsistent, showing that phosphorylation of PDE6γ may increase or decrease the ability of PDE6γ to deactivate phototransduction. To resolve role of phosphorylation of PDE6γ in living photoreceptors, we generated transgenic mice in which either one or both Threonine (T) sites in PDE6γ (T22 and T35), which are candidates for putative regulatory phosphorylation, were substituted with alanine (A). Phosphorylation of these sites was examined as a function of light exposure. We found that phosphorylation of T22 increases with light exposure in intact mouse rods while constitutive phosphorylation of T35 is unaffected by light in intact mouse rods and cones. Phosphorylation of the cone isoform of PDE6γ, PDE6H, is constitutively phosphorylated at the T20 residue. Light-induced T22 phosphorylation was lost in T35A transgenic rods, and T35 phosphorylation was extinguished in T22A transgenic rods. The interdependency of phosphorylation of T22 and T35 suggests that light-induced, post-translational modification of PDE6γ is essential for the regulation of G-protein signaling.