Exploring the role of mitochondrial uncoupling protein 4 in brain metabolism: implications for Alzheimer's disease

Abstract

The brain's high demand for energy necessitates tightly regulated metabolic pathways to sustain physiological activity. Glucose, the primary energy substrate, undergoes complex metabolic transformations, with mitochondria playing a central role in ATP production via oxidative phosphorylation. Dysregulation of this metabolic interplay is implicated in Alzheimer's disease (AD), where compromised glucose metabolism, oxidative stress, and mitochondrial dysfunction contribute to disease progression. This review explores the intricate bioenergetic crosstalk between astrocytes and neurons, highlighting the function of mitochondrial uncoupling proteins (UCPs), particularly UCP4, as important regulators of brain metabolism and neuronal function. Predominantly expressed in the brain, UCP4 reduces the membrane potential in the inner mitochondrial membrane, thereby potentially decreasing the generation of reactive oxygen species. Furthermore, UCP4 mitigates mitochondrial calcium overload and sustains cellular ATP levels through a metabolic shift from mitochondrial respiration to glycolysis. Interestingly, the levels of the neuronal UCPs, UCP2, 4 and 5 are significantly reduced in AD brain tissue and a specific UCP4 variant has been associated to an increased risk of developing AD. Few studies modulating the expression of UCP4 in astrocytes or neurons have highlighted protective effects against neurodegeneration and aging, suggesting that pharmacological strategies aimed at activating UCPs, such as protonophoric uncouplers, hold promise for therapeutic interventions in AD and other neurodegenerative diseases. Despite significant advances, our understanding of UCPs in brain metabolism remains in its early stages, emphasizing the need for further research to unravel their biological functions in the brain and their therapeutic potential.

Keywords: Alzheimer’s disease; UCP4; astrocytes; mitochondria; neurons; uncoupling agent; uncoupling protein.

Figure 1

The metabolic crosstalk between astrocytes and neurons. (A) Lactate is produced from pyruvate by lactate dehydrogenase (LDH) an enzyme that operates in both directions, using NADH. Lactate produced by astrocytes can be used as energy currency in neurons, by entering the TCA cycle or as a substrate for the synthesis of glutamate. During intense neuronal activity extracellular glutamate rises leading to increased glutamate uptake by astrocytes through EAATs. This activates the Na+/K+ ATPase causing a consequential drop of ATP levels, which is counteracted by increasing glucose uptake from the blood stream and glycolysis. From glycolysis lactate is formed and released via MCTs to sustain and aid neurons. (B) Astrocyte-mediated redox adaptation to neurotransmission involves glutamate (Glu) released into the synaptic cleft, stimulating Glutamate receptors (Glu-R) in both post-synaptic neurons and astrocytes. Mitochondrial uptake of intracellular Ca²⁺ in post-synaptic neurons leads to reactive oxygen species (ROS) production. Glutamate binding to astrocytic receptors triggers a cascade via Cdk5-mediated phosphorylation of Nrf2, promoting its nuclear translocation and binding to antioxidant responsive elements (ARE). This prompts the expression of antioxidant genes, facilitating the biosynthesis and release of glutathione (GSH). Neurons uptake GSH precursors from astrocytes to detoxify activity-induced mitochondrial ROS, ensuring neuronal redox balance during synaptic activity. (C) The activation of CD38 (cluster of differentiation 38) by cyclic ADP-ribose (cADPR) induces the release of mitovesicles (extracellular vesicle containing mitochondria) by astrocytes which functionally support neurons. In turn mitochondria which require recycling are released by neurons and taken-up by astrocytes/glia.

Figure 2

Sequence alignment of UCPs proteins highlighting percentage of identity and mapping the functional residues. The human protein sequences of UCP1 (Uniprot P25874), UCP2 (Uniprot Uniprot P55851), UCP3 (Uniprot P55916), UCP4 Uniprot (O95847) and UCP5 (Uniprot O95258) were aligned using Jalview software 2.11.3.2 with T-Coffee server, visualizing similarity among sequences with percentage of identity option which colors the residues according to the ( > 80%, >60%,  > 40%,  < 40%). Residues which have been implicated by site-directed mutagenesis in transport of H⁺, sensing pH shifts, transport of Cl⁻ or interacting with nucleotide are highlighted across sequences.

Figure 3

In silico modeling of UCP4 binding site for purine nucleotide triphosphates. (A) AlphaFold protein structure prediction for UPC4 (AF-O95847-F1-model, Uniprot ID O95847) colored by confidence score (pLDDT) and showing membrane position calculated using PPM 3.0 web server. IS, intermembrane space; MM, mitochondria matrix. (B) The superimposition of UCP1 (RCSB_ID 8HBW) with UPC4 (AF-O95847-F1-model) using UCSF ChimeraX version: 1.7 matchmaker which was then rendered by Root Mean Square Deviation (RMSD) attribute (left), the residues of UCP4 with high correspondences with UCP1 are represented in blue (RMSD <1Amstrong) (right). (C) Cytoplasmic view of the GTP-UCP4 interaction with the helix colored. (D) 3D side view of GTP interaction with binding residues of UCP4 (AF-O95847-F1-model) predicted with AutoDock4 version 4.2.6. (E) 2D interaction diagram of GTP-UCP4 interaction using LigPlot⁺ version 2.2 results representing 2D.

Figure 4

Sequence alignment of UCP4 in different species. (A) The UCP4 protein sequences from humans (Homo sapiens, UniProt ID: O95847), monkeys (Macaca mulatta, UniProt ID: F6QG76), mice (Mus musculus, UniProt ID: Q9D6D0), rats (Rattus norvegicus, UniProt ID: A6JJ30), nematodes (Caenorhabditis elegans), and flies (Drosophila melanogaster, UniProt ID: Q9VMK1), were aligned using Jalview software 2.11.3.2 with T-Coffee server, visualizing similarity among sequences with percentage of identity option which colors the residues according to the ( > 80%, >60%,  > 40%,  < 40%). (B) The identity matrix between UCP4 sequences was generated using UniProt.

Figure 5

NCS1 regulates UCP4 protein levels and mitochondrial Ca²⁺ homeostasis. NCS1 might regulate UCP4 gene expression in two ways: a) via a cytosolic Ca²⁺ concentration dependent manner or b) independent from cytosolic Ca²⁺. (a) Cytosolic Ca²⁺ activates NSC1 initiating interaction with IP3R located in the perinuclear region. IP3R increases nuclear Ca²⁺ targeting transcription factors like CREB. (b) NSC1 interacts with IP3R inducing Ca²⁺ release from ER (endoplasmic reticulum) which activates CaMKII. CaMKII promotes PGC-1α translocation to the nuclei, where it regulates mitochondrial biogenesis and cellular survival mechanisms. (c) NCS1/UCP4 can regulate mitochondrial Ca²⁺ stabilizing its uptake. Efficient Ca²⁺ transfer from ER to mitochondria at contact sites requires orchestrated activation of proteins involved in ER Ca²⁺ release (NCS1/IP3R) and mitochondrial Ca²⁺ uptake (VDAC1/2 and MCU). While activation of NCS1/IP3R/VDAC1 and MCU provides Ca²⁺ transfer from ER to mitochondria, UCP4 regulates the ability of mitochondria to buffer Ca²⁺ via modulation of mitochondrial membrane potential and its pH.

Figure 6

Chemical structure of known uncouplers. Chemical structures were drawn using ChemSketch (Freeware) 2023.1.2.