Poly-ADP-glycohydrolase (PARG) expression in wild-type and PARG-110 knock-out retina

Abstract

Purpose: Poly-ADP-ribosylation is a post-translational modification involved in many cellular pathways such as transcription, DNA repair and cell death. PARylation is performed by poly-ADP-ribose-polymerase using NAD+. Excessive PARP activity causes photoreceptor degeneration in the rd1 mouse and this activity is antagonized by poly-ADP-ribose-glycohydrolase (PARG) which hydrolyzes PAR polymers from acceptor proteins. Here, we investigated the impact of partial PARG loss using PARG110 KO knockout retinae and analysed the expression of lower molecular weight PARG isoforms in PARG110 KO and wt retina.

Methods: Mice carrying a specific deletion of the 110 kDa isoform of the PARG protein and corresponding wt retinae were analysed at P11-P30 using TUNEL staining and PAR immunohistochemistry. Retinal function and morphology were assessed using ERG and OCT at P30. PARG antibody which detects PARG110 and PARG-56 kDa proteins was used to detect PARG expression in PARG110 KO and wt retinae. Co-staining performed with antibodies for calbindin and PKCα to detect PARG expressing horizontal cells, amacrine cells, ganglion cells, and bipolar cells, respectively.

Results: TUNEL assay and PAR accumulation showed very few dying cells for PARG KO and wt retinae. ERG and OCT analysis showed that rod and cone photoreceptor signalling were normal in PARG KO retina. PARG immunofluorescence showed positive staining in the outer plexiform layer, in the inner nuclear layer, ganglion cell layer and optical fibres. PARG staining in the OPL was colocalized with calbindin suggesting prominent expression in horizontal cells. PARG staining in the INL and GCL showed colocalisation also for amacrine and ganglion cells. Most PKCα stained bipolar cells in the INL also showed high PARG expression.

Conclusions: PARG110 KO retinae appeared morphologically and functionally normal. In PARG110 KO retina, there was no sign of photoreceptor degeneration. These results suggest that neuroprotective strategies aimed at inhibiting PARG would not negatively affect photoreceptor retinal cell survival.

Keywords: 615 neuroprotection • 688 retina • 696 retinal degenerations: hereditary