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Christin Hanke-Gogokhia, Jeanne M Frederick, Houbin Zhang, Wolfgang Baehr; ARL3 regulates transport of prenylated and acylated proteins to photoreceptor outer segment in mouse retina. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):1321.
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© ARVO (1962-2015); The Authors (2016-present)
Based on in-vitro experiments, ARL3 (Arf-like protein 3) functions as a GDF (GDI displacement factor) for lipidated proteins. We generated photoreceptor- and retina-specific Arl3 deletions to investigate the function of ARL3 with focus on trafficking of lipidated proteins.
We generated Arl3+/GT mice containing a EUCOMM gene trap (GT) in intron 1 of the mouse Arl3 gene. Rod- and retina-specific Arl3 conditional knockout mice were obtained by crossing with Flp-mice, followed by iCre75+ and Six3Cre+ transgenics. Photoreceptor function was measured by ERG and progress of retinal degeneration among littermate mice was ascertained by optical coherence tomography, confocal immunohistochemistry and histology.
ERGs and retina histology of PN15 rod-specific knockout mice were indistinguishable from those of the WT littermates, suggesting normal photoreceptor development. OCT revealed rapid photoreceptor loss in retinas of Arl3flox/flox; iCre75+ mice after PN15. At PN20, scotopic ERG a-wave amplitudes were reduced 70-80% with the photopic ERG unaffected. In retinas of 1 month-old knockout mice, scotopic ERGs were extinguished and cone ERGs were attenuated; histology showed 4-5 rows of nuclei in the ONL. The 2 month-old Arl3flox/flox; iCre75+ retina was ~100 μm thinner and only one ONL row, presumably of cone nuclei, was present. Six3Cre+-mediated knockout of Arl3, deleting ARL3 during embryonic retina development, revealed enhanced photoreceptor degeneration: PN20 scotopic (↓80-90%) and photopic (↓70-80%) ERGs were reduced significantly, and OCT at PN15 confirmed faster retina degeneration with much thinner retina of Six3Cre+ knockout than iCre75+. Immunohistochemistry performed using retina sections of PN15 and 1 month-old rod- and retina-specific knockout mice revealed that rhodopsin, GC1 and CNGA1/3 trafficking appear normal. In contrast, prenylated PDE6 and GRK1, and acylated transducin-α mislocalize in the IS and ONL.
Rod- and retina-specific knockout of Arl3 revealed a rapidly-progressing rod degeneration followed by cone defects, resembling RP. Absence of ARL3 impaired the trafficking of peripheral membrane proteins, but not transmembrane proteins, consistent with its function as a GDF.
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