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Meghan Deanna Lysko, Philippe M D'Onofrio, Paulo D Koeberle; The role of telomere-associated proteins in retinal ganglion cell apoptosis after axotomy. Invest. Ophthalmol. Vis. Sci. 2016;57(12):752.
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© ARVO (1962-2015); The Authors (2016-present)
Telomeres are DNA-protein complexes that cap the ends of eukaryotic chromosomes. They have a dynamic secondary structure that binds and interacts with a large network of telomere-associated proteins that protect the genomic integrity and maintain the proliferative success of cells. When telomeres are destabilized, chromosome ends are misrecognized as DNA double stranded breaks, eliciting DNA damage responses that lead to cellular apoptosis. There is sparse evidence that telomere destabilization plays a role in the pathology of CNS diseases. In the present study we examined whether telomere dynamics play a role in retinal ganglion cell (RGC) degeneration after axotomy.
In order to evaluate the role of the Telomerase enzyme in RGC apoptosis after axotomy, rats received intraocular (IO)(4µL) or optic nerve (ON)(10µL) injections of either Trichostatin A (TSA: TERT up-regulator and Telomerase activator, 3 & 8 days post-axotomy), TAG-6, PIPER, or MST-312 (Telomerase Inhibitors III, IV, or IX, respectively), or a short hairpin RNA (shRNA) plasmid directed against TERT (catalytic subunit of Telomerase). In order to further examine the role of telomere-associated proteins in RGC apoptosis after axotomy, rats received ON injections (10µL) of Dicer short interfering RNAs (DsiRNAs) directed against TRF1, TRF2, TCAB1, DNA PKcs, Ku70, and EST1A. RGC survival was quantified in fixed flat-mounted retinas at 7 or 14 days post-axotomy.
IO delivery of TSA increased RGC survival by fourfold (p<0.001) at 14 days, while ON delivery of a TERT shRNA plasmid decreased RGC survival (p<0.001) at 7 days. However, ON delivery of Telomerase Inhibitors III, IV, and IX had no effect on RGC survival at 7 days. Furthermore, in vivo transfection of axotomized RGCs with TRF1 (p<0.01), TCAB1 (p<0.001), Ku70 (p<0.001), and EST1A (p<0.001) DsiRNAs decreased RGC survival at 7 days, while TRF2 or DNA PKcs DsiRNAs had no effect on RGC survival at 7 days.
Our results show that TERT gene expression promotes RGC survival while Telomerase enzymatic activity has no role in RGC survival after axotomy. Additionally, the TRF1, TCAB1, Ku70, and EST1A telomere-associated proteins play an important role in RGC survival after axotomy, while the TRF2 and DNA PKcs proteins do not.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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