Astrocytes have a high impact on the structure of the central nervous system, as they control neural activity, development, and plasticity. Heterogeneity of astrocytes has been shown before, but so far only a few studies have demonstrated heterogeneous morphology of astrocytes concerning aging. In this study, we examined morphologic differences of astrocyte subpopulations in adult mice and the progression of these differences with age. We surveyed astrocytes in olfactory bulb slices of mice aged 3 months, 1 year and 2 years (three animals each age group), based on their appearance in anti-GFAP immunostaining. Based on this data we established three different types of astrocytes: type I (stellate), type II (elliptic), and type III (squid-like). We found that with the advanced age of the mice, astrocytes grow in size and complexity. Major changes occurred between the ages of 3 months and 1 year, while between 1 and 2 years no significant development in cell size and complexity could be detected. Our results show that astrocytes in the olfactory bulb are heterogeneous and undergo morphological transformation until late adolescence but not upon senescence. Structural plasticity is further substantiated by the expression of vimentin in some astrocyte processes in all age groups.

PSENEN/PEN2 is the smallest subunit of the γ-secretase complex, an intramembrane protease that cleaves proteins within their transmembrane domains. Mutations in components of the γ-secretase underlie familial Alzheimer disease. In addition to its proteolytic activity, supplementary, γ-secretase independent, functions in the macroautophagy/autophagy-lysosome system have been proposed. Here, we screened for PSENEN-interacting proteins and identified CLN3. Mutations in CLN3 are causative for juvenile neuronal ceroid lipofuscinosis, a rare lysosomal storage disorder considered the most common neurodegenerative disease in children. As mutations in the PSENEN and CLN3 genes cause different neurodegenerative diseases, understanding shared cellular functions of both proteins might be pertinent for understanding general cellular mechanisms underlying neurodegeneration. We hypothesized that CLN3 modulates γ-secretase activity and that PSENEN and CLN3 play associated roles in the autophagy-lysosome system. We applied CRISPR gene-editing and obtained independent isogenic HeLa knockout cell lines for PSENEN and CLN3 . Following previous studies, we demonstrate that PSENEN is essential for forming a functional γ-secretase complex and is indispensable for γ-secretase activity. In contrast, CLN3 does not modulate γ-secretase activity to a significant degree. We observed in PSENEN - and CLN3 -knockout cells corresponding alterations in the autophagy-lysosome system. These include reduced activity of lysosomal enzymes and lysosome number, an increased number of autophagosomes, increased lysosome-autophagosome fusion, and elevated levels of TFEB (transcription factor EB). Our study strongly suggests converging roles of PSENEN and CLN3 in the autophagy-lysosome system in a γ-secretase activity-independent manner, supporting the idea of common cytopathological processes underlying different neurodegenerative diseases. Abbreviations: Aβ, amyloid-beta; AD, Alzheimer disease; APP, amyloid precursor protein; ATP5MC, ATP synthase membrane subunit c; DQ-BSA, dye-quenched bovine serum albumin; ER, endoplasmic reticulum; GFP, green fluorescent protein; ICC, immunocytochemistry; ICD, intracellular domain; JNCL, juvenile neuronal ceroid lipofuscinosis; KO, knockout; LC3, microtubule associated protein 1 light chain 3; NCL, neuronal ceroid lipofuscinoses; PSEN, presenilin; PSENEN/PEN2: presenilin enhancer, gamma-secretase subunit; TAP, tandem affinity purification; TEV, tobacco etch virus; TF, transferrin; WB, Western blot; WT, wild type.

SorLA is a member of the Vps10p-domain (Vps10p-D) receptor family of type-I transmembrane proteins conveying neuronal endosomal sorting. The extracellular/luminal moiety of SorLA has a unique mosaic domain composition and interacts with a large number of different and partially unrelated ligands, including the amyloid precursor protein as well as amyloid-β. Several studies support a strong association of SorLA with sporadic and familial forms of Alzheimer’s disease (AD). Although SorLA seems to be an important factor in AD, the large number of different ligands suggests a role as a neuronal multifunctional receptor with additional intracellular sorting capacities. Therefore, understanding the determinants of SorLA’s subcellular targeting might be pertinent for understanding neuronal endosomal sorting mechanisms in general. A number of cytosolic adaptor proteins have already been demonstrated to determine intracellular trafficking of SorLA. Most of these adaptors and several ligands of the extracellular/luminal moiety are shared with the Vps10p-D receptor Sortilin. Although SorLA and Sortilin show both a predominant intracellular and endosomal localization, they are targeted to different endosomal compartments. Thus, independent adaptor proteins may convey their differential endosomal targeting. Here, we hypothesized that Sortilin and SorLA interact with the cytosolic adaptors PSD95 and PICK1 which have been shown to bind the Vps10p-D receptor SorCS3. We observed only an interaction for SorLA and PICK1 in mammalian-two-hybrid, pull-down and cellular recruitment experiments. We demonstrate by mutational analysis that the C-terminal minimal PDZ domain binding motif VIA of SorLA mediates the interaction. Moreover, we show co-localization of SorLA and PICK1 at vesicular structures in primary neurons. Although the physiological role of the interaction between PICK1 and SorLA remains unsolved, our study suggests that PICK1 partakes in regulating SorLA’s intracellular itinerary.

Evidence from genetics and from analyzing cellular and animal models have converged to suggest links between neurodegenerative disorders of early and late life. Here, we summarize emerging links between the most common late life neurodegenerative disease, Alzheimer’s disease, and the most common early life neurodegenerative diseases, neuronal ceroid lipofuscinoses. Genetic studies reported an overlap of clinically diagnosed Alzheimer’s disease and mutations in genes known to cause neuronal ceroid lipofuscinoses. Accumulating data strongly suggest dysfunction of intracellular trafficking mechanisms and the autophagy-endolysosome system in both types of neurodegenerative disorders. This suggests shared cytopathological processes underlying these different types of neurodegenerative diseases. A better understanding of the common mechanisms underlying the different diseases is important as this might lead to the identification of novel targets for therapeutic concepts, the transfer of therapeutic strategies from one disease to the other and therapeutic approaches tailored to patients with specific mutations. Here, we review dysfunctions of the endolysosomal autophagy pathway in Alzheimer’s disease and neuronal ceroid lipofuscinoses and summarize emerging etiologic and genetic overlaps.