Endosomal trafficking is required for glycosylation and normal maturation of the Alzheimer’s-associated protein sorLA

Abstract

The sorting receptor sorLA encoded by the SORL1 gene is implicated in Alzheimer’s disease (AD) pathogenesis. Genetic studies have identified AD-associated SORL1 mutations and the expression of sorLA in AD brains is reported to be reduced. SorLA is a receptor of the retromer trafficking complex and functions at the endosome, and deficiency in sorLA phenocopies the endosomal pathologies found in AD. SorLA undergoes posttranslational modifications and maturation with ultimate ectodomain shedding, however knowledge of these processes remains limited. Here we demonstrate that sorLA exists at the cell membrane in two forms, an immature and a mature form, characterized by distinct N-glycosylation profiles. The mature sorLA form has acquired complex type N-glycans and is shed from the cell surface by the TACE juxtmembrane cleavage. The immature form of sorLA present at the cell surface is shown to have immature ER-type N-glycans (high-mannose type susceptible to endo H) and does not undergo shedding, however, upon endocytosis and recycling to the cell surface via endosomal trafficking pathways the immature sorLA form acquires complex-type N-glycans. These results suggest an unusual secretion model for sorLA whereby that immature sorLA first traffics to the cell membrane without acquiring Golgi processing of N-glycans, and only upon retrograde trafficking does sorLA acquire normal Golgi maturation of N-glycans and become susceptible to TACE regulated shedding. Supportive evidence for this model include a sorLA mutant with deficient endosomal trafficking and in vivo studies demonstrating requirement of retromer for sorLA trafficking in the brain of retromer VPS26 deficient mice. Collectively, our study establishes the role endosomal trafficking plays in sorLA’s normal maturation, and point to impaired maturation as a signature of AD-associated sorLA dysfunction.

Impaired SorLA maturation and trafficking as a new mechanism for SORL1 missense variants in Alzheimer disease

“…These data are fully consistent with our results obtained in HEK293 cells in which we detect a disproportionate accumulation of immature forms, and the presence of both immature and mature forms at the plasma membrane. The presence of both species at the plasma membrane in HEK293 cells, which is consistent with the data previously reported by other groups (Christensen et al , 2020; Jacobsen et al , 2001), could therefore be related to the overexpression of SorLA and the saturation of the ER quality control system.…”

“…When overexpressed in HEK293 cells, WT-SorLA protein consisted of two distinct forms on immunoblot analysis (Figure 1B, Figure 2) that were hypothesized to be different glycoforms of the protein. As previously shown (Jacobsen et al , 2001; Christensen et al , 2020), removal of all N -linked glycans by treatment with PNGase F resulted in a single band that migrates at ∼250kDa, which corresponds to the predicted molecular weight of the full-length SorLA (SorLA FL ) protein (Figure 2A). Treatment with neuraminidase altered the mobility of only the upper band that subsequently migrated as the lower form, indicating that the upper species correspond to complex type N -glycans capped with sialic acid residues (Figure 2A).…”

“…Newly synthesized SorLA molecules follow the constitutive secretory pathway from the endoplasmic reticulum (ER) to the cell surface, through the Golgi. During their transport through the secretory pathway, SorLA proteins are subjected to several post-translational modifications, including both N- and O-glycosylation, and maturation processes (Jacobsen et al , 2001; Fiete et al , 2007; Schmidt et al , 2007; Fjorback et al , 2012; Steentoft et al , 2013; Hampe et al , 2000; Christensen et al , 2020), resulting in the presence of immature and mature species. At the cell surface, a minority of SorLA molecules are subject to proteolytic shedding, releasing the soluble ectodomain of the receptor, termed soluble SorLA (sSorLA).…”

A genetically modified minipig model for Alzheimer’s disease with SORL1 haploinsufficiency

“…We recently showed how deficiency of the neuronal retromer core unit VPS26B leads to a decrease in functional SORLA maturation, 16 , 42 so here we wanted to determine whether the reverse is also true: whether SORL1 deficiency affects the expression level of VPS26 proteins. WB analysis of Cx homogenates ( Figure 2 E) showed strong and significantly reduced expression of both isoforms: VPS26B ( Figure 2 H) and VPS26A ( Figure 2 I).…”

The neuronal retromer can regulate both neuronal and microglial phenotypes of Alzheimer's disease

“…A reliable effect was, however, observed within CA1 neurons, with larger SORL1-positive puncta in VPS35-depleted neurons, an effect that was partially rescued upon VPS35 repletion ( Figures 5F and 5G ). This likely reflects the fact that VPS35 depletion is known to cause enlarged endosomes ( Bhalla et al, 2012 ), a cytopathogenic hallmark of AD ( Cataldo et al, 2000 ), and that retromer plays a role in transporting SORL1 from the endosome to the cell surface ( Christensen et al, 2020 ).…”