Failure of Autophagy–Lysosomal Pathways in Rod Photoreceptors Causes the Early Retinal Degeneration Phenotype Observed inCln6^nclfMice

Abstract

PURPOSE. Vision loss caused by photoreceptor death represents one of the first symptoms in neuronal ceroid lipofuscinosis, a condition characterized by accumulation of intracellular waste. Cln6 nclf mice have a naturally occurring mutation in ceroid-lipofuscinosis neuronal (CLN) protein 6 and are a model of this disorder. In order to identify the effect intracellular waste (lipofuscin) accumulation plays in driving retinal degeneration, the time course of degeneration was carefully characterized functionally using the electroretinogram and structurally using histology. METHODS. Cln6 nclf and C57BL/6J, wild-type, mice were studied at postnatal day 18 (P18), P30, P60, P120, and P240, and retinal degeneration was correlated with changes in the retinal pigment epithelial (RPE) and neuronal autophagy-lysosomal pathways using super-resolution microscopy. RESULTS. In Cln6 nclf mice there was significant loss of rod photoreceptor function at P18, prior to photoreceptor nuclei loss at P60. In contrast, cone pathway function was not affected until P240. The loss of rod photoreceptor function correlated with significant disruption of the autophagy-lysosomal degradation pathways within photoreceptors, but not in the RPE or other retinal neurons. Additionally, there was cytosolic accumulation of P62 and undigested mitochondrial-derived, ATP synthase subunit C in the photoreceptor layers of Cln6 nclf mice at P30. CONCLUSIONS. These results suggest that rod photoreceptors have an increased sensitivity to disturbances in the autophagy-lysosomal pathway and the subsequent failure of mitochondrial turnover, relative to other retinal cells. It is likely that primary failure of the rod photoreceptors rather than the RPE or other retinal neurons underlies the early visual dysfunction that occurs in the Cln6 nclf mouse model.

Photoreceptor Degeneration in Pro23His Transgenic Rats (Line 3) Involves Autophagic and Necroptotic Mechanisms

“…We used the electroretinogram to measure retinal function of wild-type and P23H-3 rats (n ≥ 10) at P18, P30, P60, and P90, as described previously (Jobling et al, 2013;Vessey et al, 2015;Von Eisenhart-Rothe et al, 2018). There are a number of different ways of using the electroretinogram to measure rod and cone mediated responses (Kremers and Tanimoto, 2018).…”

“…After completion of ERG recording, OCT and fundus images were captured on a Micron III rodent fundus camera using Micron III software (Phoenix Technology Group, Pleasanton, CA, United States) as described previously (Von Eisenhart-Rothe et al, 2018).…”

Mice deficient in the lysosomal enzyme palmitoyl-protein thioesterase 1 (PPT1) display a complex retinal phenotype

“…Alternatively, accumulation of lysosomal storage material may impair the fusion of autophagosomes with lysosomes and thus the processing of autophagic material, resulting in accumulation of SQSTM1/p62-positive aggregates 72 . Increased levels of the autophagic markers SQSTM1/p62 and/or microtubule-associated protein 1 light chain 3-II (LC3-II) as pathological markers for impaired constitutive autophagy have also been reported in mouse models of various other NCLs, both in the brain 41,73–75 and in the retina 62,76,77 .…”

Moving towards a new era of genomics in the neuronal ceroid lipofuscinoses

“…For example, in addition to displaying progressively accumulating NCL-type lysosomal storage material [159], the naturally occurring CLN6 mouse model, Cln6 nclf , also exhibits markers of autophagy impairment, including an accumulation of autophagic vacuoles, p62 aggregates, and an increase in LC3-II puncta, while ER stress or unfolded protein response (UPR) activation were not immediately evident [156]. An involvement of CLN6 in autophagic clearance is further supported by two more recent studies showing alterations in autophagy markers in primary neural cultures from naturally occurring CLN6-deficient sheep [160], and in Cln6 nclf mice, in the photoreceptor layer of the retina [161]. Interestingly, a recent paper has identified CLN6 as an interactor with the heat shock protein αB-crystallin, which suppressed the accumulation of a disease mutant version that is prone to aggregration, an effect that was abolished upon lysosomal inhibition; the authors suggest this implies that CLN6 plays a protective role, via an autophagylysosome dependent process, in the cellular response to proteins that are prone to aggregation [162].…”