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Jennifer Caughey, Lauren Lee Daniele, Anuradha Dhingra, Kathleen Boesze-Battaglia; Changes in RPE peroxisome lipid metabolism in response to light onset. Invest. Ophthalmol. Vis. Sci. 2018;59(9):4033.
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© ARVO (1962-2015); The Authors (2016-present)
Healthy visual function relies on the symbiotic relationship between photoreceptors and the retinal pigment epithelium (RPE). The RPE phagocytoses the shed tips of photoreceptor outer segments, rich in very long chain fatty acids (VLCFA), with a diurnal peak at light onset. Despite the importance of peroxisomes for the breakdown of VLCFA, little is known of their role in RPE health. This study has two goals: first, to investigate whether peroxisome number and function depend on time of day and second, to determine if degradation of RPE peroxisomes relies on selective autophagy.
Peroxisome number was inferred from Western blot analysis (WB) of LC3B-/- and WT lysates made at different times of day and probed for PEX14, a peroxisome specific membrane protein. To gain insight into peroxisome function, we measured the activity of catalase at various times relative to light onset in WT and LC3B-/- mice using a fluorescence-based assay. Catalase activity was normalized to total protein or total catalase via dot blot. Immunofluorescence intensities were quantified for Pex14 & PMP70 in cryosections of eyes from LC3B-/- , ATG5ΔRPE and control mice. PMP70 is a peroxisome specific ATP binding cassette transporter for long chain fatty acyl CoA intermediates.
Catalase activity was higher in the early morning VS afternoon (p<.04). In contrast, catalase activity did not vary significantly in LC3B-/- mouse RPE with time of day. Expression of PEX14 did not vary over time in either LC3B-/- or WT mouse RPE. Autophagy deficient mice showed elevated expression of peroxisomal membrane proteins. PEX14 intensities showed a 2.3 fold increase in LC3B-/- mice (p<.0002) and a 1.6 fold increase in ATG5ΔRPE mice (p<.05). PMP70 intensities showed a 1.8 fold increase in LC3B-/- mice (p<.02) and a 2.5 fold increase in ATG5ΔRPE mice (p<.02).
RPE catalase activity varies diurnally in WT but not LC3B-/-, with the highest levels coinciding with the phagocytic peak. Regulation of catalase activity depends on LC3B-/-. Collectively, our studies suggest that peroxisome turnover is likely an autophagy dependent process.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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