A novel Alex3/Gαq protein complex regulating mitochondrial dynamics, dendritic complexity, and neuronal survival

In neurons, mitochondrial dynamics and trafficking are essential to provide the energy required for neurotransmission and neuronal activity. Recent studies point to GPCR and G proteins as important regulators of mitochondrial dynamics and energy metabolism. Here we show that activation of Gαq negatively regulates mitochondrial dynamics and trafficking in neurons. Gαq interacts with the mitochondrial trafficking protein Alex3. By generating a CNS-specific armcx3 knock-out mouse line, we demonstrate that Alex3 is required for Gαq effects on mitochondrial dynamics and trafficking, and dendritic growth. Armcx3-deficient mice present decreased OXPHOS complex and ER stress response protein levels, which correlate with increased neuronal death, motor neuron and neuromuscular synaptic loss, and severe motor alterations. Finally, we show that Alex3 disassembles from the Miro1/Gαq complex upon calcium rise. These data uncover a novel Alex3/Gαq complex that regulates neuronal mitochondrial dynamics and neuronal death and allows the control of mitochondrial functions by GPCRs.

The mitospecific domain of Mrp7 (bL27) supports mitochondrial translation during fermentation and is required for effective adaptation to respiration

We demonstrate here that mitoribosomal protein synthesis, responsible for the synthesis of oxidative phosphorylation (OXPHOS) subunits encoded by mitochondrial genome, occurs at high levels during glycolysis fermentation and in a manner uncoupled from OXPHOS complex assembly regulation. Furthermore, we provide evidence that the mitospecific domain of Mrp7 (bL27), a mitoribosomal component, is required to maintain mitochondrial protein synthesis during fermentation, but is not required under respiration growth conditions. Maintaining mitotranslation under high glucose fermentation conditions also involves Mam33 (p32/gC1qR homolog), a binding partner of Mrp7’s mitospecific domain, and together they confer a competitive advantage for a cell's ability to adapt to respiration-based metabolism when glucose becomes limiting. Furthermore, our findings support that the mitoribosome, and specifically the central protuberance (CP) region, may be differentially regulated and/or assembled, under the different metabolic conditions of fermentation and respiration. Based on our findings, we propose the purpose of mitotranslation is not limited to the assembly of OXPHOS complexes, but also plays a role in mitochondrial signaling critical for switching cellular metabolism from a glycolysis- to a respiratory-based state.

Ca2+ overload- and ROS-associated mitochondrial dysfunction contributes to δ-tocotrienol-mediated paraptosis in melanoma cells

Melanoma is an aggressive tumor with still poor therapy outcomes. δ-tocotrienol (δ-TT) is a vitamin E derivative displaying potent anti-cancer properties. Previously, we demonstrated that δ-TT triggers apoptosis in human melanoma cells. Here, we investigated whether it might also activate paraptosis, a non-canonical programmed cell death. In accordance with the main paraptotic features, δ-TT was shown to promote cytoplasmic vacuolization, associated with endoplasmic reticulum/mitochondrial dilation and protein synthesis, as well as MAPK activation in A375 and BLM cell lines. Moreover, treated cells exhibited a significant reduced expression of OXPHOS complex I and a marked decrease in oxygen consumption and mitochondrial membrane potential, culminating in decreased ATP synthesis and AMPK phosphorylation. This mitochondrial dysfunction resulted in ROS overproduction, found to be responsible for paraptosis induction. Additionally, δ-TT caused Ca2+ homeostasis disruption, with endoplasmic reticulum-derived ions accumulating in mitochondria and activating the paraptotic signaling. Interestingly, by using both IP3R and VDAC inhibitors, a close cause-effect relationship between mitochondrial Ca2+ overload and ROS generation was evidenced. Collectively, these results provide novel insights into δ-TT anti-melanoma activity, highlighting its ability to induce mitochondrial dysfunction-mediated paraptosis. Graphic Abstract δ-tocotrienol induces paraptotic cell death in human melanoma cells, causing endoplasmic reticulum dilation and mitochondrial swelling. These alterations induce an impairment of mitochondrial function, ROS production and calcium overload.

Tetramethylpyrazine Improves Cognitive Impairment and Modifies the Hippocampal Proteome in Two Mouse Models of Alzheimer's Disease

Alzheimer's disease (AD), one of the most common neurodegenerative diseases, has no effective treatment. We studied the potential effects of tetramethylpyrazine (TMP), an alkaloid in the rhizome of Ligusticum chuanxiong Hort. used in Traditional Chinese Medicine (chuānxiong) to treat ischemic stroke, on AD progression in two AD mouse models. Eight-month-old 3xTg-AD mice received TMP treatment (10 mg/kg/d) for 1 month, and 4-month-old APP/PS1-AD mice received TMP treatment (10 mg/kg/d) for 2 months. Behavioral tests, including step-down passive avoidance (SDA), new object recognition (NOR), Morris water maze (MWM), and Contextual fear conditioning test showed that TMP significantly improved the learning and memory of the two AD-transgenic mice. In addition, TMP reduced beta-amyloid (Aß) levels and tau phosphorylation (p-tau). Venny map pointed out that 116 proteins were commonly changed in 3xTg mice vs. wild type (WT) mice and TMP-treated mice vs. -untreated mice. The same 130 proteins were commonly changed in APP/PS1 mice vs. WT mice and TMP-treated mice vs. -untreated mice. The functions of the common proteins modified by TMP in the two models were mainly involved in mitochondrial, synaptic, cytoskeleton, ATP binding, and GTP binding. Mitochondrial omics analysis revealed 21 and 20 differentially expressed mitochondrial proteins modified by TMP in 3xTg-AD mice and APP/PS1 mice, respectively. These differential proteins were located in the mitochondrial inner membrane, mitochondrial outer membrane, mitochondrial gap, and mitochondrial matrix, and the function of some proteins is closely related to oxidative phosphorylation (OXPHOS). Western-blot analysis confirmed that TMP changed the expression of OXPHOS complex proteins (sdhb, ndufa10, uqcrfs1, cox5b, atp5a) in the hippocampus of the two AD mice. Taken together, we demonstrated that TMP treatment changed the hippocampal proteome, reduced AD pathology, and reduced cognitive impairment in the two AD models. The changes might be associated with modification of the mitochondrial protein profile by TMP. The results of the study suggest that TMP can improve the symptoms of AD.

A Mitochondrial Polymorphism Alters Immune Cell Metabolism and Protects Mice from Skin Inflammation

Several genetic variants in the mitochondrial genome (mtDNA), including ancient polymorphisms, are associated with chronic inflammatory conditions, but investigating the functional consequences of such mtDNA polymorphisms in humans is challenging due to the influence of many other polymorphisms in both mtDNA and the nuclear genome (nDNA). Here, using the conplastic mouse strain B6-mtFVB, we show that in mice, a maternally inherited natural mutation (m.7778G > T) in the mitochondrially encoded gene ATP synthase 8 (mt-Atp8) of complex V impacts on the cellular metabolic profile and effector functions of CD4+ T cells and induces mild changes in oxidative phosphorylation (OXPHOS) complex activities. These changes culminated in significantly lower disease susceptibility in two models of inflammatory skin disease. Our findings provide experimental evidence that a natural variation in mtDNA influences chronic inflammatory conditions through alterations in cellular metabolism and the systemic metabolic profile without causing major dysfunction in the OXPHOS system.

Effect of Ubiquinol on Glaucomatous Neurodegeneration and Oxidative Stress: Studies for Retinal Ganglion Cell Survival and/or Visual Function

Oxidative stress is one of major causal factors in glaucomatous neurodegeneration. Ubiquinol promotes retinal ganglion cell (RGC) survival against glaucomatous insults such as oxidative stress. Here we investigated the effect of ubiquinol on RGC survival and/or visual function in mouse models of glaucoma and oxidative stress. DBA/2J and age-matched DBA/2J-Gpnmb+ (D2-Gpnmb+), which do not develop intraocular pressure elevation, or C57BL/6J mice were fed with ubiquinol (1%) or control diet daily for 5 or 2 months. We assessed RGC survival by Brn3a immunohistochemistry and measured expression levels of active and total BAX, peroxisome proliferator-activated receptor-gamma coactivator 1α, transcription factor A (TFAM) and oxidative phosphorylation (OXPHOS) complex protein. Following induction of oxidative stress by paraquat injection, we also assessed visual function. In glaucomatous retina, ubiquinol supplementation significantly promoted RGC survival, blocked BAX activation and increased TFAM and OXPHOS complex II protein expression. Also, ubiquinol supplementation ameliorated oxidative stress-induced visual dysfunction. These findings indicate that ubiquinol promotes RGC survival by increasing TFAM expression and OXPHOS complex II activity in glaucomatous neurodegeneration, and that ubiquinol enhances RGC survival and preserves visual function against oxidative stress. We propose that ubiquinol has a therapeutic potential for treating oxidative stress-associated glaucomatous neurodegeneration.

Mitochondrial OXPHOS Biogenesis: Co-Regulation of Protein Synthesis, Import, and Assembly Pathways

The assembly of mitochondrial oxidative phosphorylation (OXPHOS) complexes is an intricate process, which—given their dual-genetic control—requires tight co-regulation of two evolutionarily distinct gene expression machineries. Moreover, fine-tuning protein synthesis to the nascent assembly of OXPHOS complexes requires regulatory mechanisms such as translational plasticity and translational activators that can coordinate mitochondrial translation with the import of nuclear-encoded mitochondrial proteins. The intricacy of OXPHOS complex biogenesis is further evidenced by the requirement of many tightly orchestrated steps and ancillary factors. Early-stage ancillary chaperones have essential roles in coordinating OXPHOS assembly, whilst late-stage assembly factors—also known as the LYRM (leucine–tyrosine–arginine motif) proteins—together with the mitochondrial acyl carrier protein (ACP)—regulate the incorporation and activation of late-incorporating OXPHOS subunits and/or co-factors. In this review, we describe recent discoveries providing insights into the mechanisms required for optimal OXPHOS biogenesis, including the coordination of mitochondrial gene expression with the availability of nuclear-encoded factors entering via mitochondrial protein import systems.

RNA sequencing identifies a cryptic exon caused by a deep intronic variant in NDUFB10 resulting in isolated Complex I deficiency.

The diagnosis of mitochondrial disorders remains a challenging and often unmet need. We sought to investigate a sibling pair with suspected mitochondrial disease and a clinical presentation notable for global developmental delay, poor growth, sensorineural hearing loss, and brain MRI abnormalities, both with early death. Following uninformative exome and genome sequencing of the family quartet, RNA sequencing was pursued as an orthogonal testing strategy. RNA sequencing of fibroblasts from the older sibling identified the presence of a cryptic exon in intron 1 of NDUFB10, that included an in-frame stop codon. NDUFB10 encodes a subunit of mitochondrial OXPHOS complex I. Differential expression analysis relative to control samples suggested significantly decreased expression. The cryptic exon was found to contain a rare intronic variant, NM_004548.3:c.131-442G>C, that was homozygous in both affected siblings and absent from population allele frequency databases. Immunoblot and quantitative proteomic analysis of fibroblasts from the older sibling revealed decreased abundance of complex I subunits associated with NDUFB10, providing evidence of isolated complex I deficiency. Biallelic variants in NDUFB10 have previously been reported in a single individual with infantile-onset mitochondrial disease. We present data implicating a deep intronic variant in NDUFB10 as the cause of mitochondrial disease in two further individuals. This variant results in loss of expression and overall destabilization of mitochondrial OXPHOS complex I and highlights the importance of RNA sequencing as a complementary diagnostic tool in patients undergoing genome-wide diagnostic evaluation.

ECSIT prevents Alzheimer’s disease pathology by regulating neuronal mitochondrial ROS and mitophagy

Altered mitochondrial fitness is a potential triggering factor in Alzheimer’s disease (AD). Mitochondrial quality control pathways are dysfunctional and mitochondrially-derived reactive oxygen species (mROS) levels are increased in AD patient brains. However, the pathways responsible for dysregulated mROS accumulation have remained relatively unclear. In this study, we demonstrate that levels of ECSIT, a mitochondrial oxidative phosphorylation (OxPhos) complex I (CI)-associated protein, are reduced in AD-affected brains. Neuronal ECSIT downregulation increased mROS generation and impaired mitophagy of defective mitochondria. Consequently, decreasing neuronal ECSIT caused AD-like changes, including memory loss and neuropathology. In contrast, augmented neuronal expression of ECSIT protected against the development of an AD-like phenotype. Decreased levels of ECSIT in AD patient brains therefore likely contribute to oxidative stress, neuroinflammation and AD pathogenesis.