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Yuhong Wang, Ammaji Rajala, Michelle Ranjo-Bishop, Robert E Anderson, Raju V S Rajala; Characterization of Autophagy-linked FYVE (Alfy) Protein in the Retina. Invest. Ophthalmol. Vis. Sci. 2014;55(13):398.
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© ARVO (1962-2015); The Authors (2016-present)
In the central nervous system, autophagy is a normal cellular process that prevents the formation of protein deposits from large protein aggregates that are inefficiently degraded by proteosomes. Autophagy-linked FYVE protein (Alfy) is a recently discovered, ubiquitously expressed protein that has an FYVE domain that binds to phosphoinositide 3-kinase (PI3K) -generated PI-3-P with high affinity. Overexpression of the C-terminal fragment of Alfy has been shown to increase aggregate clearance and neuroprotection in a Drosophila eye model of polyglutamine toxicity clearance. There are no studies available on the expression and role of Alfy in the retina, so we characterized Alfy in this study.
Localization of Alfy was examined by immunohistochemistry performed on wild type and cone-dominant Nrl-/- retinal sections. The C-terminal fragments of Alfy and PI-3-P binding mutant were tested both in vitro (mammalian cells) and in vivo (lipoplex-mediated subretinal injection into mice). The outer nuclear thickness was used as a function of rod photoreceptor degeneration, and cone cell death was assessed by peanut agglutinin and cone-transducin labeling.
We found that Alfy was expressed in inner segments and the outer and inner plexiform layers, in both rod- and cone-dominant retina. The expression of Alfy-WT in HEK-293T cells did not change the morphology of the cells. However, the PI-3-P-binding mutant induced autophagosome-like vacuoles and increased cell death. We also found increased rod and cone cell death in mice injected with PI-3-P-binding mutant, but not in WT-injected mice.
Our study shows that Alfy is expressed in the retina. Our research also suggests that the interaction between PI3K-generated PI-3-P and the FYVE domain of Alfy is indispensable for normal functions of autophagy in the retina.
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