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J. M. Tonne, K. A. Linberg, D. Eisenschink, M. K. Traub, T. Rytz, T. Thomas, G. P. Lewis, D. K. Vaughan, S. K. Fisher; SUMOylation of Retinal Proteins in Aroused vs. Torpid Ground Squirrels. Invest. Ophthalmol. Vis. Sci. 2008;49(13):2060.
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© ARVO (1962-2015); The Authors (2016-present)
The Small Ubiquitin-Like Modifier (SUMO) is a 100 amino acid protein that is covalently bound to target proteins by the Ubc9 enzyme. Thus, "SUMOylation" is a post-translational modification that draws upon a pre-existing pool of free SUMO. SUMOylation is hypothesized to spare protein conjugates from proteasomal degradation. SUMOylation of brain proteins is up-regulated during hibernation torpor in the 13-lined ground squirrel (GS), which naturally tolerates a repeated ischemia-reperfusion cycle over several months (Lee et al. J. Cereb. Blood Flow Metab. 2007). Protein sparing by SUMOylation might thus be a key neuroprotective strategy in the CNS. We tested the hypothesis that SUMOylation of retinal proteins would be similarly up-regulated during torpor in the cone-dominant GS retina.
Young of the year GSs from the UW Oshkosh captive colony were used. Hibernation-naïve (aroused) GSs were sacrificed in August, October and November, whereas first-hibernation (torpid) GSs were sacrificed in October and December. Retinas were processed for in situ immunolabelling, Western blotting, and reverse transcriptase-PCR. Immunoprobes (gifts of Dr. Lee) and primers that distinguished SUMO-1 from SUMO-2/3 were used.
Messenger RNAs for SUMO-1 and SUMO-2/3 were detected in retinas from aroused and torpid GSs. Compared to blots from aroused GSs, blots from torpid GSs demonstrated down-regulated free SUMO-2/3 (Mr 16 kD) alongside up-regulated SUMO-2/3 protein conjugates (Mr 60-200 kD). A similar but less-robust response was seen on blots probed with anti-SUMO-1. Retinal SUMO conjugates were not identical to those identified in brain (Lee et al. 2007). In situ immunoreactivity (IR) for both types of SUMO in August/aroused GS retina was observed in the RPE and all retinal layers except for two regions in photoreceptors: their outer segments and ellipsoid mitochondria. SUMO-IR in November/aroused and December/torpid GS retinas was reduced, relative to that seen in August/aroused retinas, particularly sclerad of the inner plexiform layer.
Our Western blot data support the hypothesis that SUMOylation of retinal proteins is upregulated during torpor when the GS brain exhibits resilience to repeated ischemia-reperfusion cycles. Reduced in situ SUMO-IR during torpor may reflect epitope masking resulting from this upregulated SUMOylation of protein targets, combined with their cross-linking by aldehyde fixation.
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