title: Mutant rab8 Impairs docking and fusion of rhodopsin-bearing post-Golgi membranes and causes cell death of transgenic Xenopus rods.
abstract: Rab8 is a GTPase involved in membrane trafficking. In photoreceptor cells, rab8 is proposed to participate in the late stages of delivery of rhodopsin-containing post-Golgi membranes to the plasma membrane near the base of the connecting cilium. To test the function of rab8 in vivo, we generated transgenic Xenopus laevis expressing wild-type, constitutively active (Q67L), and dominant negative (T22N) forms of canine rab8 in their rod photoreceptors as green fluorescent protein (GFP) fusion proteins. Wild-type and constitutively active GFP-rab8 proteins were primarily associated with Golgi and post-Golgi membranes, whereas the dominant negative protein was primarily cytoplasmic. Expression of wild-type GFP-rab8 had minimal effects on cell survival and intracellular structures. In contrast, GFP-rab8T22N caused rapid retinal degeneration. In surviving peripheral rods, tubulo-vesicular structures accumulated at the base of the connecting cilium. Expression of GFP-rab8Q67L induced a slower retinal degeneration in some tadpoles. Transgene effects were transmitted to F1 offspring. Expression of the GFP-rab8 fusion proteins appears to decrease the levels of endogenous rab8 protein. Our results demonstrate a role for rab8 in docking of post-Golgi membranes in rods, and constitute the first report of a transgenic X. laevis model of retinal degenerative disease.
authors.affiliation: Department of Neuroscience, University of Connecticut Health Center, Farmington, Connecticut 06032-3705, USA. omoritz@neuron.uchc.edu
authors.forename: O L
authors.initials: OL
authors.lastname: Moritz
authors.forename: B M
authors.initials: BM
authors.lastname: Tam
authors.forename: L L
authors.initials: LL
authors.lastname: Hurd
authors.forename: J
authors.initials: J
authors.lastname: Peränen
authors.forename: D
authors.initials: D
authors.lastname: Deretic
authors.forename: D S
authors.initials: DS
authors.lastname: Papermaster
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D016923:Cell Death / Q000502:physiology*; D002462:Cell Membrane / Q000378:metabolism; D004195:Disease Models, Animal; D004285:Dogs; D017930:Genes, Reporter / Q000235:genetics; D006056:Golgi Apparatus / Q000502:physiology*; D015151:Immunoblotting; D008856:Microscopy, Fluorescence; D021381:Protein Transport / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012162:Retinal Degeneration / Q000235:genetics / Q000503:physiopathology; D017948:Retinal Rod Photoreceptor Cells / Q000166:cytology* / Q000378:metabolism* / Q000648:ultrastructure; D012243:Rhodopsin / Q000378:metabolism; D014982:Xenopus laevis; D020691:rab GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D011993:Recombinant Fusion Proteins; D012243:Rhodopsin; D020691:rab GTP-Binding Proteins