Unraveling axonal traffic jam
The protease BACE1 participates in Aβ production and has been shown to accumulate in dystrophic neurons and contribute to axonal pathology in patients with Alzheimer’s disease (AD) and in animal models. The mechanisms mediating BACE1 accumulation are unclear. Here, Lomoio et al. showed that the Golgi-localized γ-ear-containing ARF binding protein 3 (GGA3) plays a main role in BACE1 localization. Deletion of Gga3 resulted in BACE1 accumulation and axonal swelling that was prevented by BACE inhibition. The authors identified a loss-of-function mutation in GGA3 in patients with AD, and Gga3 deletion worsened AD pathology in a mouse model. The results contribute to elucidate the mechanisms mediating axonal damage in AD.
Abstract
Axonal dystrophy, indicative of perturbed axonal transport, occurs early during Alzheimer’s disease (AD) pathogenesis. Little is known about the mechanisms underlying this initial sign of the pathology. This study proves that Golgi-localized γ-ear-containing ARF binding protein 3 (GGA3) loss of function, due to Gga3 genetic deletion or a GGA3 rare variant that cosegregates with late-onset AD, disrupts the axonal trafficking of the β-site APP-cleaving enzyme 1 (BACE1) resulting in its accumulation in axonal swellings in cultured neurons and in vivo. We show that BACE pharmacological inhibition ameliorates BACE1 axonal trafficking and diminishes axonal dystrophies in Gga3 null neurons in vitro and in vivo. These data indicate that axonal accumulation of BACE1 engendered by GGA3 loss of function results in local toxicity leading to axonopathy. Gga3 deletion exacerbates axonal dystrophies in a mouse model of AD before β-amyloid (Aβ) deposition. Our study strongly supports a role for GGA3 in AD pathogenesis, where GGA3 loss of function triggers BACE1 axonal accumulation independently of extracellular Aβ, and initiates a cascade of events leading to the axonal damage distinctive of the early stage of AD.
INTRODUCTION
Dysfunction of axonal transport has been implicated in the pathogenesis of Alzheimer’s disease (AD) (1–3). Axonal swellings indicative of axonal transport disruption have been found in postmortem brains from patients with early stage AD (4). It has also been shown that in vivo impairment of axonal transport and decreased axonal transport rates might have an impact on AD pathogenesis early in the disease process (5–7). Moreover, increased plasma concentration of neurofilament light chain (NfL) indicative of axonal damage has been detected in AD presymptomatic stages (8, 9). However, the mechanisms leading to axonal damage in the early phase of pathology remain unclear.
Dystrophic neurites surround β-amyloid (Aβ) deposits forming neuritic plaques (10). The aspartyl protease β-site amyloid precursor protein (APP)–cleaving enzyme 1 (BACE1) is the β-secretase that catalyzes the rate limiting step in Aβ generation (11). Under normal conditions, BACE1 localizes to presynaptic terminals, whereas, in AD brains, it accumulates in dystrophic neurites (12–15). Likewise, defects in axon transport leading to endogenous BACE1 buildup in axonal swellings have been reported (16). Although we do not know precisely when BACE1 pathology occurs, other studies suggest it to occur early in AD mouse models (17). Nevertheless, the specific mechanism(s) of BACE1 accumulation remains unknown.
We previously demonstrated that the Golgi-localized γ-ear-containing ADP-ribosylation factor (ARF) binding protein 3 (GGA3), a monomeric clathrin adaptor highly expressed in the brain (18), regulates BACE1 lysosomal degradation (19). GGA3 consists of four segments: a VPS-27/Hrs/STAM (VHS) domain that binds the acidic di-leucine sorting signal, DXXLL; a GGA/TOM1 (GAT) domain that binds Arf:guanosine 5′-triphosphate and ubiquitin; a hinge region that recruits clathrin; and a gamma-adaptin ear (GAE) domain that exhibits sequence similarity to the ear region of γ-adaptin and recruits a number of accessory proteins (20). We have shown that, in nonneuronal cells, GGA3 depletion induces BACE1 accumulation in early endosomes by preventing its delivery to lysosomes (19, 21, 22) and that Gga3 deletion increases BACE1 amount in vivo (22) and exacerbates Aβ pathology in a mouse model of familial AD (23). Moreover, we established that GGA3 is decreased and inversely correlated with BACE1 in the temporal cortices of patients with AD (19) and in a mouse model of familial AD (23). Despite compelling evidence indicating that GGA3 regulates BACE1 trafficking, the role of GGA3 in BACE1 neuronal polarized sorting is unknown.
Here, we characterized the neuronal localization of GGA3 in murine hippocampal neurons and found that the protein is distributed, along with BACE1, in both dendrites and axons. We determined that Gga3 deletion triggers BACE1 elevation and axonal trafficking disruption resulting in BACE1 accumulation in axonal swellings. BACE pharmacological inhibition prevented axonal swelling formation, in vitro and in vivo, and improved BACE1 axonal motility. These data indicate that axonal accumulation of BACE1 triggered by Gga3 deletion results in localized toxicity leading to the disruption of the trafficking of other presynaptic proteins and axonopathy.
In support of a role of GGA3 in AD pathogenesis, we identified a GGA3 rare variant, Ins545T, that cosegregated with late-onset AD (LOAD). Ins545T results in a loss of function as demonstrated by its inability to rescue BACE1 axonal trafficking and accumulation in axonal swellings in Gga3−/− neurons. Last, we demonstrated that in 2-month-old 5XFAD mice, Gga3 deletion exacerbates axonal pathology and triggers BACE1 accumulation in axonal swellings before senile plaques formation. Our findings demonstrate that GGA3 loss of function triggers BACE1 axonal buildup and initiates a cascade of events leading to the axonal damage characteristic of the early stage of AD in the absence of extracellular Aβ deposition in mice….
