title: Submembrane assembly and renewal of rod photoreceptor cGMP-gated channel: insight into the actin-dependent process of outer segment morphogenesis.
abstract: The photoreceptor outer segment (OS) is comprised of two compartments: plasma membrane (PM) and disk membranes. It is unknown how the PM renewal is coordinated with that of the disk membranes. Here we visualized the localization and trafficking process of rod cyclic nucleotide-gated channel α-subunit (CNGA1), a PM component essential for phototransduction. The localization was visualized by fusing CNGA1 to a fluorescent protein Dendra2 and expressing in Xenopus laevis rod photoreceptors. Dendra2 allowed us to label CNGA1 in a spatiotemporal manner and therefore discriminate between old and newly trafficked CNGA1-Dendra2 in the OS PM. Newly synthesized CNGA1 was preferentially trafficked to the basal region of the lateral OS PM where newly formed and matured disks are also added. Unique trafficking pattern and diffusion barrier excluded CNGA1 from the PM domains, which are the proposed site of disk membrane maturation. Such distinct compartmentalization allows the confinement of cyclic nucleotide-gated channel in the PM, while preventing the disk membrane incorporation. Cytochalasin D and latrunculin A treatments, which are known to disrupt F-actin-dependent disk membrane morphogenesis, prevented the entrance of newly synthesized CNGA1 to the OS PM, but did not prevent the entrance of rhodopsin and peripherin/rds to the membrane evaginations believed to be disk membrane precursors. Uptake of rhodopsin and peripherin/rds coincided with the overgrowth of the evaginations at the base of the OS. Thus F-actin is essential for the trafficking of CNGA1 to the ciliary PM, and coordinates the formations of disk membrane rim region and OS PM.
authors.affiliation: Department of Pharmacology, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44106-4965.
authors.forename: Ina
authors.initials: I
authors.lastname: Nemet
authors.affiliation: Department of Pharmacology, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44106-4965.
authors.forename: Guilian
authors.initials: G
authors.lastname: Tian
authors.affiliation: Department of Pharmacology, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44106-4965 yxi19@case.edu.
authors.forename: Yoshikazu
authors.identifier: http://orcid.org/0000-0003-4696-4836
authors.initials: Y
authors.lastname: Imanishi
mesh_terms: D000199:Actins / Q000378:metabolism*; D000818:Animals; D030801:Animals, Genetically Modified; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure; D002614:Chelating Agents / Q000494:pharmacology; D054815:Cyclic Nucleotide-Gated Cation Channels / Q000235:genetics; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D010450:Endopeptidases / Q000494:pharmacology; D066298:In Vitro Techniques; D007814:Larva; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008954:Models, Biological; D009024:Morphogenesis / Q000235:genetics / Q000502:physiology*; D038761:Photobleaching; D021381:Protein Transport / Q000502:physiology; D012160:Retina / Q000166:cytology*; D017948:Retinal Rod Photoreceptor Cells / Q000166:cytology* / Q000648:ultrastructure; D012243:Rhodopsin / Q000235:genetics / Q000378:metabolism; D014981:Xenopus
chemical_list: D000199:Actins; C475741:CNGA1 protein, bovine; D002614:Chelating Agents; D054815:Cyclic Nucleotide-Gated Cation Channels; D008164:Luminescent Proteins; D004533:Egtazic Acid; D012243:Rhodopsin; D010450:Endopeptidases; C023139:dispase; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid
keywords: CNGA1; cyclic nucleotide-gated channel; morphogenesis; photoreceptor; retina; rhodopsin