title: Ectopic transgene expression in the retina of four transgenic mouse lines.
abstract: Retinal expression of transgenes was examined in four mouse lines. Two constructs were driven by the choline acetyltransferase (ChAT) promoter: green fluorescent protein conjugated to tau protein (tau-GFP) or cytosolic yellow fluorescent protein (YFP) generated through CRE recombinase-induced expression of Rosa26 (ChAT-CRE/Rosa26YFP). Two other constructs targeted inhibitory interneurons: GABAergic horizontal and amacrine cells identified by glutamic acid decarboxylase (GAD65-GFP) or parvalbumin (PV) cells (PV-CRE/Rosa26YFP). Animals were transcardially perfused and retinal sections prepared. Antibodies against PV, calretinin (CALR), calbindin (CALB), and tyrosine hydroxylase (TH) were used to counterstain transgene-expressing cells. In PVxRosa and ChAT-tauGFP constructs, staining appeared in vertically oriented row of processes resembling Müller cells. In the ChATxRosa construct, populations of amacrine cells and neurons in the ganglion cell layer were labeled. Some cones also exhibited GFP fluorescence. CALR, PV and TH were found in none of these cells. Occasionally, we found GFP/CALR and GFP/PV double-stained cells in the ganglion cell layer (GCL). In the GAD65-GFP construct, all layers of the neuroretina were labeled, except photoreceptors. Not all horizontal cells expressed GFP. We did not find GFP/TH double-labeled cells and GFP was rarely present in CALR- and CALB-containing cells. Many PV-positive neurons were also labeled for GFP, including small diameter amacrines. In the GCL, single labeling for GFP and PV was ascertained, as well as several CALR/PV double-stained neurons. In the GCL, cells triple labeled with GFP/CALR/CALB were sparse. In conclusion, only one of the four transgenic constructs exhibited an expression pattern consistent with endogenous retinal protein expression, while the others strongly suggested ectopic gene expression.
authors.affiliation: Department of Experimental Zoology and Neurobiology, University of Pécs, Pécs, Hungary.
authors.forename: Robert
authors.initials: R
authors.lastname: Gábriel
authors.affiliation: Laboratory of Molecular Biology and Genetics, Institute of Experimental Medicine, Hungarian Academy of Sciences, 1450, Budapest, Hungary.
authors.forename: Ferenc
authors.initials: F
authors.lastname: Erdélyi
authors.affiliation: Laboratory of Molecular Biology and Genetics, Institute of Experimental Medicine, Hungarian Academy of Sciences, 1450, Budapest, Hungary.
authors.forename: Gábor
authors.initials: G
authors.lastname: Szabó
authors.affiliation: Center for Structural and Functional Neuroscience, University of Montana, Missoula, MT, USA.|Department of Biomedical and Pharmaceutical Sciences, University of Montana, Missoula, MT, USA.|Department of Pharmacology and Neuroscience, Texas Tech University Health Sciences Center, School of Medicine, Lubbock, TX, 79430, USA.
authors.forename: J Josh
authors.initials: JJ
authors.lastname: Lawrence
authors.affiliation: Institute of Sport Sciences and Physical Education, University of Pécs, Ifjúság u. 6., 7624, Pécs, Hungary. mwilhelm@gamma.ttk.pte.hu.
authors.forename: Márta
authors.initials: M
authors.lastname: Wilhelm
mesh_terms: D025042:Amacrine Cells / Q000166:cytology / Q000378:metabolism; D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism; D064092:Calbindin 1 / Q000378:metabolism; D064032:Calbindin 2 / Q000378:metabolism; D002795:Choline O-Acetyltransferase / Q000378:metabolism*; D059329:Cholinergic Neurons / Q000166:cytology / Q000378:metabolism; D063928:Ependymoglial Cells / Q000166:cytology / Q000378:metabolism; D059330:GABAergic Neurons / Q000166:cytology / Q000378:metabolism; D015870:Gene Expression; D049452:Green Fluorescent Proteins / Q000378:metabolism; D007150:Immunohistochemistry / Q000379:methods*; D008164:Luminescent Proteins / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D010320:Parvalbumins / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012160:Retina / Q000378:metabolism*; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism; D016875:tau Proteins / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; C576499:Calb1 protein, mouse; C576511:Calb2 protein, mouse; D064092:Calbindin 1; D064032:Calbindin 2; D008164:Luminescent Proteins; C558298:Mapt protein, mouse; D010320:Parvalbumins; D016875:tau Proteins; C054562:yellow fluorescent protein, Bacteria; D049452:Green Fluorescent Proteins; D014446:Tyrosine 3-Monooxygenase; D002795:Choline O-Acetyltransferase
keywords: ChAT-CRE/Rosa26YFP; ChAT-tauGFP; Expression mismatch; GAD65-GFP; PV-CRE/Rosa26YFP