APOE4 affects basal and NMDAR mediated protein synthesis in neurons by perturbing calcium homeostasis

Abstract

Apolipoprotein E (APOE), one of the primary lipoproteins in the brain has three isoforms in humans – APOE2, APOE3, and APOE4. APOE4 is the most well-established risk factor increasing the pre-disposition for Alzheimer’s disease. The presence of the APOE4 allele alone is shown to cause synaptic defects in neurons and recent studies have identified multiple pathways directly influenced by APOE4. However, the mechanisms underlying APOE4 induced synaptic dysfunction remain elusive. Here, we report that the acute exposure of primary cortical neurons to APOE4 leads to a significant decrease in global protein synthesis. APOE4 treatment also abrogates the NMDA mediated translation response indicating an impairment of synaptic signaling. Importantly, we demonstrate that both APOE3 and APOE4 generate a distinct translation response which is closely linked to their respective calcium signature. Acute exposure to APOE3 causes a short burst of calcium through NMDARs in neurons leading to an initial decrease in protein synthesis which quickly recovers. Contrarily, APOE4 leads to a sustained increase in calcium levels by activating both NMDARs and L-VGCCs, thereby causing sustained translation inhibition through eEF2 phosphorylation, which in turn disrupts NMDAR response. Thus, we show that APOE4 affects basal and activity mediated protein synthesis response in neurons by affecting calcium homeostasis. We propose this as a possible mechanism to explain the synaptic dysfunction caused by APOE4.Highlights / SummaryAPOE3 treatment causes a short burst of calcium through NMDARs, leading to an acute increase in eEF2 phosphorylation which eventually recovers to basal levels.Global translation follows a similar temporal profile of initial inhibition followed by recovery in APOE3 treated neurons, thus unaffecting the NMDA mediated translation response.APOE4 treatment activates both NMDARs and L-VGCCs leading to a marked elevation in calcium levels, thus causing sustained increase in eEF2 phosphorylation as well as global translation inhibition.Hence, the NMDA mediated response is perturbed, potentially causing a stress-related phenotype in APOE4 treated neurons.Thus, different calcium signatures and sources lead to distinct temporal profiles of translation.

Early and lifelong effects of APOE4 on neuronal gene expression networks relevant to Alzheimer’s disease

“…Our results, in concert with previous data, suggest that calcium signaling disruptions may play an important role in APOE4 effects and start early in the brain. Prior studies have found that APOE4 triggers greater intracellular calcium levels than APOE3, and impacts NMDA-receptor-mediated signaling in cultured neurons to disrupt calcium signaling and protein translation 52 .…”

Does the imbalance in the apolipoprotein E isoforms underlie the pathophysiological process of sporadic Alzheimer's disease?

“…Thus, the dysregulation of calcium homeostasis by APOE ε4 leads to its impaired protein synthesis response (Figure 2A). 48 …”

A multi‐hit hypothesis for an APOE4‐dependent pathophysiological state

“…The UPR can reduce ER protein folding load by inhibiting global protein translation, but chronic activation can stimulate pro‐inflammatory signalling converging on NF‐ĸB and apoptosis (Dose et al, 2016). Interestingly, UPR activation has been linked to ApoE4 expression in E4‐TR mouse brain (Zalocusky et al, 2021; N. Zhao et al, 2020) and ApoE4 fragment accumulation in vitro (Zhong et al, 2009), while in parallel ApoE4 may suppress global translation via prolonged neuronal calcium influx (Ramakrishna et al, 2021). Speculatively, ApoE4 may converge on ER stress and mitochondrial dysfunction via direct ApoE4 fragment accumulation and indirectly via chronically elevated intracellular calcium levels (Hit 7).…”

“…Early observations demonstrated that the direct application of full length or truncated ApoE to neuronal cells resulted in an increase in intracellular calcium (Ohkubo et al, 2001; Qiu et al, 2004; Tolar et al, 1999; Wang & Gruenstein, 1997) and calcium bursting responses (Konings et al, 2021). Further, ApoE4 specifically elevated intracellular calcium levels relative to ApoE3 and was shown to be likely mediated by ApoE receptor activation of L‐type voltage gated calcium channels (VGCCs) and NMDARs, which may involve prolonged calcium influx dynamics (Ohkubo et al, 2001; Qiu et al, 2003; Ramakrishna et al, 2021; Veinbergs et al, 2002; Figure 7, Path A). Further, NMDAR‐dependent accumulation of intracellular calcium and induced cell death was also shown to be exacerbated by injury in the presence of APOE4 or APOE ‐KO but was rescued by APOE3 in vitro (Jiang et al, 2015).…”

“…The mechanisms underpinning these perturbations in signalling may fall downstream of elevated intracellular calcium influx by ApoE4 (Hit 7). Indeed, ApoE4 application to cultured neurons led to sustained calcium influx both basally and with stimulation, followed by a greater p‐eEF2‐mediated inhibition of global translation but preferential upregulation of multiple synaptic gene targets (Ramakrishna et al, 2021). As discussed in ‘Hit 1’, it is unclear what may mediate acute synaptogenic to chronic synaptotoxic signalling involving APOE4 , although speculatively this may involve gradual compensatory changes in kinase‐phosphatase activation.…”