BACE1 Translation: At the Crossroads Between Alzheimer's Disease Neurodegeneration and Memory Consolidation

Abstract

Human life unfolds not only in time and space, but also in the recollection and interweaving of memories. Therefore, individual human identity depends fully on a proper access to the autobiographical memory. Such access is hindered under pathological conditions such as Alzheimer's disease, which affects millions of people worldwide. Unfortunately, no effective cure exists to prevent this disorder, the impact of which will rise alarmingly within the next decades. While Alzheimer's disease is largely considered to be the outcome of amyloid-β (Aβ) peptide accumulation in the brain, conceiving this complex disorder strictly as the result of Aβ-neurotoxicity is perhaps a too straight-line simplification. Instead, complementary to this view, the tableau of molecular disarrangements in the Alzheimer's disease brain may be reflecting, at least in part, a loss of function phenotype in memory processing. Here we take BACE1 translation and degradation as a gateway to study molecular mechanisms putatively involved in the transition between memory and neurodegeneration. BACE1 participates in the excision of Aβ-peptide from its precursor holoprotein, but plays a role in synaptic plasticity too. Its translation is governed by eIF2α phosphorylation: a hub integrating cellular responses to stress, but also a critical switch in memory consolidation. Paralleling these dualities, the eIF2α-kinase HRI has been shown to be a nitric oxide-dependent physiological activator of hippocampal BACE1 translation. Finally, beholding BACE1 as a representative protease active in the CNS, we venture a new perspective on the cellular basis of memory, which may incorporate neurodegeneration in itself as a drift in memory consolidating systems.

Keywords: Alzheimer’s disease; eIF2α; exosomes; heme-regulated eIF-2α kinase; memory; nitric oxide; physiological stress response; proteolysis; translation initiation; β-secretase.

Fig.1

BACE1 open reading frame flanked by its 5’ and 3’ untranslated regions (UTRs). Schematic representation depicting BACE1 transcript structure. Note how BACE1 3’UTR is almost 2.5 times larger than the protein coding region (ORF). In addition, BACE1 3’UTR contains two cytoplasmic polyadenylation (CPE) motifs (UUUUAU) and accessory sequences (AAUAA) that facilitate dendritic mRNA sorting by the CPE-binding protein (CPEB) [307]. BACE1 5’UTR, in turn, gates translation initiation combining the inhibitory effect of 1) upstream initiation codons (uAUGs) and 2) stable secondary structure deriving from a high GC-content (∼75 %).

Fig.2

Point mutations in AβPP and its processing by BACE1. A)“Framing β -amyloid”. Snapshot from an article by Prof. John Hardy [Framing β -amyloid, Nature Genetics 1, 233–234 (1992)] showing the initial AD-causative mutations (labelled as “AD” in the figure) that were discovered in AβPP. Today more than 15 disease-causing mutations are known to impact AβPP. For a recent personal account by Prof. Hardy on the story of how AD-causative mutations in AβPP were discovered, and how this led to the formulation of the amyloid hypothesis, you can see [84]. Image reproduced with permission. B) BACE1 participation in A β PP processing. Amyloid-β (Aβ) peptide is excised out of the amyloid precursor protein (AβPP) via sequential cleavages catalyzed by BACE1 and the γ-secretase complex, respectively. Inhibition (of both proteases) is an actively prosecuted goal as a pharmacological strategy in AD therapy. Nevertheless, all of the synthesized inhibitors to date have repeatedly failed in clinical trials. C) Scheme showing AD pathogenic mutations on AβPP affecting the β-secretase cleavage (in red), the β’-secretase cleavage (in blue), the α-secretase cleavage (in purple), and the γ-secretase cleavage (in orange) of AβPP. Those mutations affecting the aggregation of the peptide are highlighted with a “*” symbol, while those mutations affecting the degradation of the peptide are indicated with a “**” symbol. The symbol # is used to indicate the mutations that are pathogenic for cerebral amyloid angiopathy, while the symbol “?” indicates that the mechanisms of pathogenesis remains unknown for that particular mutation.

Fig.3

Cascade of neurotoxic effects deriving from Aβ peptide aggregation. Soluble monomers of Aβ peptide aggregate into different species (oligomers, protofibrils, larger fibrils). Neurotoxic effects attributed to each aggregative state are indicated in capital letters in the right column of the figure. Up to the present day, the relative contribution of each Aβ-species to the overall neurotoxicity-effect in AD remains a subject of intense debate. Little arrows indicate the presence of protofibrils in the middle transmission electron microscopy (TEM) panel. Image magnification: × 40,000, for oligomers and larger fibrils; for protofibrils, lower right scale bar equals 200 nM.

Fig.4

Role of ternary complexes in translation initiation. eIF2 recycling & ternary complexes regeneration. A) Eukaryotic initiation factor 2 (eIF2) is a heterotrimeric protein consisting in three subunits (α, β, γ). eIF2 binds a GTP molecule in its γ-subunit, while phosphorylation at serine 51 of the α-subunit modulates its interaction with eIF2B. B) eIF2 forms a ternary complex with the initiator transfer RNA coupled to the corresponding Methionine (Met-tRNAi), and a GTP molecule docked at the γ-subunit of eIF2. Note that tRNAi appears depicted in the diagram as a dark backbone finishing with three tips symbolizing the anticodon. C) Ternary complexes (the formation of which is thermodynamically favored by eIF2-GTP) are loaded in to the 40S ribosomal thanks to the intervention of other translation factors. D) Translation initiation is a GTP-dependent process. The energy resulting from the hydrolysis of the eIF2-bound GTP is required for initiation codon recognition and commitment of the ribosome to complete the initiation pathway. As a result, a low energy eIF2-GDP complex is released after each round of translation initiation. eIF2B is the guanine exchange factor (GEF) refueling eIF2 with new energy-rich GTP molecules. This process yields new active ternary complexes ready to engage in new cycles of translation initiation.

Fig.5

uAUGs contribution in gating BACE1 translation. BACE1 transcript leader, alternatively named 5’untranslated region (5’UTR), hinders BACE1 translation initiation under basal conditions. Upstream initiation codons (uAUGs) present in BACE1 5’UTR “seduce” the small ribosomal subunit and prevent it from reaching the main open reading frame (ORF). As a consequence, BACE1 translation will be kept at a low, insignificant rate. This situation will be reversed only following a drop in ternary complex availability.

Fig.6

BACE1 translation facilitation. BACE1 translational repression can be by-passed by ribosomal leaky scanning & reinitiation, a condition requiring a lowering in ternary complex availability. As depicted in the diagram, when ternary complex availability is high (A, basal conditions) translation initiation can occur at high frequency at the uAUGs, preventing translation of BACE1 from its main ORF. Conversely, when the availability of ternary complexes diminishes (B) there is a drop in the formation of active ribosomal complexes, leading to a decreased recognition of upstream AUGs (ribosome leaky scanning) and to a and more frequent recognition of the main ORF allowing BACE1 protein synthesis to start. s.r.s., small ribosomal subunit.

Fig.7

Intracellular signaling converging in eIF2B inhibition. The guanine-exchange activity of eIF2B can be modulated in response to stimuli conveyed by two alterative pathways. On the one hand, phosphorylation of the eukariotic initiation 2-alpha (eIF2α) is mediated by four different stress-activated eIF2α kinases (PERK, PKR, HRI, and GCN2), resulting in a competitive blockade of eIF2B. On the other hand, the otherwise constitutively inhibited Glycogen synthase kinase 3β (GSK3β), when released from its inhibition, catalyzes a direct inhibitory phosphorylation upon eIF2B. s.r.s., small ribosomal subunit.

Fig.8

Three-step pathway model for NO-induced BACE1 translation activation. The heme-regulated eIF2α kinase (HRI), a nitric oxide sensor, appeared to be a plausible node connecting nitric oxide (NO) stimulation with BACE1 expression, through phospho-eIF2α-mediated BACE1 translational de-repression. Other uAUG-bearing transcripts such as GluN2B [149] respond in a similar way to NO/HRI induced eIF2-phosphorylation, opening the possibility that NO behaves as a general translational facilitator for uAUG-bearing transcripts.

Fig.9

Proposed model for eIF2α-mediated translational control at the synapse. In a resting state exempt of cellular stress, nitric oxide (NO)-HRI signaling induces a peak in eIF2α phosphorylation levels, which are rapidly turned back to their basal state through the action of eIF2α-phosphatases. Such transient rise in eIF2α levels triggers the timely translation of synaptically polarized uAUG-bearing mRNAs, while temporarily arresting general protein translation. Importantly, such signaling model can only be effective when cellular stress does not interfere with eIF2α phosphorylation through the action of other eIF2-kinases (PERK, PKR, GCN2) or even an eventually stress-activated HRI.

Fig.10

Nitric oxide: a master physiological mediator hovering above the integrated stress response? The four existing eIF2α-kinases (PERK, PKR, HRI, GCN2) share a common catalytic-domain and harbor different activator-domains endowing each kinase with a differential sensitivity to stress. Following stress sensing, eIF2α-kinases phosphorylate eIF2α and shut down protein translation in a process known as integrated stress response. This process aims, for example, at securing metabolic resources under energy deprivation conditions, alleviating protein load in the endoplasmic reticulum under missfolding protein conditions, or at avoiding the translation of exogenous proteins from viral origin. In the absence of stress, nitric oxide (NO) signals through the Heme-regulated eIF2α-kinase (HRI), activating thereby BACE1 translation from its mRNA in a physiological setting.

Fig.11

Alteration of intracellular compartments in AD. Left panel: Intracellular compartments in a healthy neuron. AβPP is endocyted from the plasma membrane and cleaved to AβPP-βCTF by BACE in the membrane of early endosomes (EEs). EEs mature to multivesicular bodies (MVBs) when the rim of the endosome invaginates toward the lumen forming the intraluminal vesicles (ILVs). AβPP-βCTF can be cleaved to Aβ by γ-secretase on the membrane of the endosome or be incorporated to the ILVs where can also be processed by γ-secretase to Aβ. After ILVs are degraded after fusion of MVBs to lysosomes or secreted in form of exosomes. A utophagosomes are also degraded after fusion to lysosomes. Right panel: In AD, the loss of function of lysosomes decreases the degradation of autophagosomes and MVBs, which accumulated and become enlarged. In addition, accumulation of AβPP-βCTF in endosomes enhance the enlargement by recruiting the APPL1 effector. To compensate the lack of degradation of ILVs and their content (including Aβ), there is an increased secretion of ILVs in form of exosomes upon fusion of MVBs to the plasma membrane. Aβ is secreted from the lumen of MVBs or bound to exosomes. The secreted Aβ may work as a glue for proteins, lipids and nucleic acids destined to be degraded by the activated microglia. Exosomes can also be removed from the brain through the blood-brain barrier.

Fig.12

Heme-regulated eIF2α-kinase (HRI) activation by nitric oxide (NO) (A), HRI mRNA expression in different cellular types (B), and hypothetical model for NO/HRI-induced translational activation of cationic channels at the vascular wall (C). A) Diagram depicting NO-mediated signaling, including HRI kinase activation. HRI kinase activation by nitric oxide may constitute a third branch of NO-mediated intracellular signaling, by which translation initiation in uAUG-bearing transcripts becomes facilitated. B) Profile of HRI mRNA expression in different cellular types. HRI mRNA expression was detected in all the cellular types assayed, including human aortic vascular smooth cells (HA-VSMCs) and human cerebral vascular smooth cells (HC-VSMCs). (–), negative control without input RNA. C) Hypothetical model for NO/HRI-induced translational activation of cationic channels at the vascular wall. After NO is synthesized in the endothelial cells, it diffuses to the adjacent VSMC where, theoretically, it could activate the VSMC-resident HRI kinase; this would in turn facilitate the translation of uAUG-bearing transcripts such as those coding for the β1 subunit of the BK channel (KCNMB1), and the SK3 channel (KCNN3), respectively. Both cationic channels depicted in the figure (KCNMB1 and KCNN3) are calcium activated potassium channels (K_Ca). eNOS, endothelial nitric oxide synthase; nNOS, neuronal nitric oxide synthase; mtNOS, mitochondrial nitric oxide synthase; iNOS, inducible nitric oxide synthase; Cell lines: Raw, macrophages; BV2, microglia; SH-SY5Y: human neuroblastoma; LCFSN: traqueal epithelial. Primers used for semi-quantitative RT-PCR amplification: AGGAACAAGCGGAGCCG(mHRI_F); CCGACCAGTCCTTACGCC (mHRI_R). [446 bp amplicon].