International Journal of Molecular Sciences

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19 pages, 3701 KiB

Open AccessArticle

Phosphorylation of SARS-CoV-2 Orf9b Regulates Its Targeting to Two Binding Sites in TOM70 and Recruitment of Hsp90

by Lukas Brandherm, Antonio Mario Kobaš, Mara Klöhn, Yannick Brüggemann, Stephanie Pfaender, Joachim Rassow and Sebastian Kreimendahl

Int. J. Mol. Sci. 2021, 22(17), 9233; https://doi.org/10.3390/ijms22179233 - 26 Aug 2021

Cited by 14 | Viewed by 3638

Abstract

SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) is the causative agent of the COVID19 pandemic. The SARS-CoV-2 genome encodes for a small accessory protein termed Orf9b, which targets the mitochondrial outer membrane protein TOM70 in infected cells. TOM70 is involved in a signaling [...] Read more.

SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) is the causative agent of the COVID19 pandemic. The SARS-CoV-2 genome encodes for a small accessory protein termed Orf9b, which targets the mitochondrial outer membrane protein TOM70 in infected cells. TOM70 is involved in a signaling cascade that ultimately leads to the induction of type I interferons (IFN-I). This cascade depends on the recruitment of Hsp90-bound proteins to the N-terminal domain of TOM70. Binding of Orf9b to TOM70 decreases the expression of IFN-I; however, the underlying mechanism remains elusive. We show that the binding of Orf9b to TOM70 inhibits the recruitment of Hsp90 and chaperone-associated proteins. We characterized the binding site of Orf9b within the C-terminal domain of TOM70 and found that a serine in position 53 of Orf9b and a glutamate in position 477 of TOM70 are crucial for the association of both proteins. A phosphomimetic variant Orf9b_S53E showed drastically reduced binding to TOM70 and did not inhibit Hsp90 recruitment, suggesting that Orf9b–TOM70 complex formation is regulated by phosphorylation. Eventually, we identified the N-terminal TPR domain of TOM70 as a second binding site for Orf9b, which indicates a so far unobserved contribution of chaperones in the mitochondrial targeting of the viral protein. Full article

(This article belongs to the Special Issue Mitochondrial Protein Network: From Biogenesis to Bioenergetics in Health and Disease 2.0)

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15 pages, 13306 KiB

Open AccessArticle

Thyroid Hormone Induces Ca²⁺-Mediated Mitochondrial Activation in Brown Adipocytes

by Minh-Hanh Thi Nguyen, Dat Da Ly, Nhung Thi Nguyen, Xu-Feng Qi, Hyon-Seung Yi, Minho Shong, Seung-Kuy Cha, Sangkyu Park and Kyu-Sang Park

Int. J. Mol. Sci. 2021, 22(16), 8640; https://doi.org/10.3390/ijms22168640 - 11 Aug 2021

Cited by 4 | Viewed by 2360

Abstract

Thyroid hormones, including 3,5,3′-triiodothyronine (T₃), cause a wide spectrum of genomic effects on cellular metabolism and bioenergetic regulation in various tissues. The non-genomic actions of T₃ have been reported but are not yet completely understood. Acute T₃ treatment significantly [...] Read more.

Thyroid hormones, including 3,5,3′-triiodothyronine (T₃), cause a wide spectrum of genomic effects on cellular metabolism and bioenergetic regulation in various tissues. The non-genomic actions of T₃ have been reported but are not yet completely understood. Acute T₃ treatment significantly enhanced basal, maximal, ATP-linked, and proton-leak oxygen consumption rates (OCRs) of primary differentiated mouse brown adipocytes accompanied with increased protein abundances of uncoupling protein 1 (UCP1) and mitochondrial Ca²⁺ uniporter (MCU). T₃ treatment depolarized the resting mitochondrial membrane potential (Ψ_m) but augmented oligomycin-induced hyperpolarization in brown adipocytes. Protein kinase B (AKT) and mammalian target of rapamycin (mTOR) were activated by T₃, leading to the inhibition of autophagic degradation. Rapamycin, as an mTOR inhibitor, blocked T₃-induced autophagic suppression and UCP1 upregulation. T₃ increases intracellular Ca²⁺ concentration ([Ca²⁺]_i) in brown adipocytes. Most of the T₃ effects, including mTOR activation, UCP1 upregulation, and OCR increase, were abrogated by intracellular Ca²⁺ chelation with BAPTA-AM. Calmodulin inhibition with W7 or knockdown of MCU dampened T₃-induced mitochondrial activation. Furthermore, edelfosine, a phospholipase C (PLC) inhibitor, prevented T₃ from acting on [Ca²⁺]_i, UCP1 abundance, Ψ_m, and OCR. We suggest that short-term exposure of T₃ induces UCP1 upregulation and mitochondrial activation due to PLC-mediated [Ca²⁺]_i elevation in brown adipocytes. Full article

(This article belongs to the Special Issue Mitochondrial Protein Network: From Biogenesis to Bioenergetics in Health and Disease 2.0)

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14 pages, 2390 KiB

Open AccessArticle

Mitochondrial Metabolism behind Region-Specific Resistance to Ischemia-Reperfusion Injury in Gerbil Hippocampus. Role of PKCβII and Phosphate-Activated Glutaminase

by Małgorzata Beręsewicz-Haller, Olga Krupska, Paweł Bochomulski, Danuta Dudzik, Anita Chęcińska, Wojciech Hilgier, Coral Barbas, Krzysztof Zablocki and Barbara Zablocka

Int. J. Mol. Sci. 2021, 22(16), 8504; https://doi.org/10.3390/ijms22168504 - 07 Aug 2021

Cited by 5 | Viewed by 2555

Abstract

Ischemic episodes are a leading cause of death worldwide with limited therapeutic interventions. The current study explored mitochondrial phosphate-activated glutaminase (GLS1) activity modulation by PKCβII through GC-MS untargeted metabolomics approach. Mitochondria were used to elucidate the endogenous resistance of hippocampal CA2-4 and dentate [...] Read more.

Ischemic episodes are a leading cause of death worldwide with limited therapeutic interventions. The current study explored mitochondrial phosphate-activated glutaminase (GLS1) activity modulation by PKCβII through GC-MS untargeted metabolomics approach. Mitochondria were used to elucidate the endogenous resistance of hippocampal CA2-4 and dentate gyrus (DG) to transient ischemia and reperfusion in a model of ischemic episode in gerbils. In the present investigation, male gerbils were subjected to bilateral carotids occlusion for 5 min followed by reperfusion (IR). Gerbils were randomly divided into three groups as vehicle-treated sham control, vehicle-treated IR and PKCβII specific inhibitor peptide βIIV5-3-treated IR. Vehicle or βIIV5-3 (3 mg/kg, i.v.) were administered at the moment of reperfusion. The gerbils hippocampal tissue were isolated at various time of reperfusion and cell lysates or mitochondria were isolated from CA1 and CA2-4,DG hippocampal regions. Recombinant proteins PKCβII and GLS1 were used in in vitro phosphorylation reaction and organotypic hippocampal cultures (OHC) transiently exposed to NMDA (25 μM) to evaluate the inhibition of GLS1 on neuronal viability. PKCβII co-precipitates with GAC (GLS1 isoform) in CA2-4,DG mitochondria and phosphorylates GLS1 in vitro. Cell death was dose dependently increased when GLS1 was inhibited by BPTA while inhibition of mitochondrial pyruvate carrier (MPC) attenuated cell death in NMDA-challenged OHC. Fumarate and malate were increased after IR 1h in CA2-4,DG and this was reversed by βIIV5-3 what correlated with GLS1 activity increases and earlier showed elevation of neuronal death (Krupska et al., 2017). The present study illustrates that CA2-4,DG resistance to ischemic episode at least partially rely on glutamine and glutamate utilization in mitochondria as a source of carbon to tricarboxylic acid cycle. This phenomenon depends on modulation of GLS1 activity by PKCβII and remodeling of MPC: all these do not occur in ischemia-vulnerable CA1. Full article

(This article belongs to the Special Issue Mitochondrial Protein Network: From Biogenesis to Bioenergetics in Health and Disease 2.0)

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32 pages, 11265 KiB

Open AccessArticle

Progressive Mitochondrial SOD1^G93A Accumulation Causes Severe Structural, Metabolic and Functional Aberrations through OPA1 Down-Regulation in a Mouse Model of Amyotrophic Lateral Sclerosis

by Iago Méndez-López, Francisco J. Sancho-Bielsa, Tobias Engel, Antonio G. García and Juan Fernando Padín

Int. J. Mol. Sci. 2021, 22(15), 8194; https://doi.org/10.3390/ijms22158194 - 30 Jul 2021

Cited by 11 | Viewed by 2882

Abstract

In recent years, the “non-autonomous motor neuron death” hypothesis has become more consolidated behind amyotrophic lateral sclerosis (ALS). It postulates that cells other than motor neurons participate in the pathology. In fact, the involvement of the autonomic nervous system is fundamental since patients [...] Read more.

In recent years, the “non-autonomous motor neuron death” hypothesis has become more consolidated behind amyotrophic lateral sclerosis (ALS). It postulates that cells other than motor neurons participate in the pathology. In fact, the involvement of the autonomic nervous system is fundamental since patients die of sudden death when they become unable to compensate for cardiorespiratory arrest. Mitochondria are thought to play a fundamental role in the physiopathology of ALS, as they are compromised in multiple ALS models in different cell types, and it also occurs in other neurodegenerative diseases. Our study aimed to uncover mitochondrial alterations in the sympathoadrenal system of a mouse model of ALS, from a structural, bioenergetic and functional perspective during disease instauration. We studied the adrenal chromaffin cell from mutant SOD1^G93A mouse at pre-symptomatic and symptomatic stages. The mitochondrial accumulation of the mutated SOD1^G93A protein and the down-regulation of optic atrophy protein-1 (OPA1) provoke mitochondrial ultrastructure alterations prior to the onset of clinical symptoms. These changes affect mitochondrial fusion dynamics, triggering mitochondrial maturation impairment and cristae swelling, with increased size of cristae junctions. The functional consequences are a loss of mitochondrial membrane potential and changes in the bioenergetics profile, with reduced maximal respiration and spare respiratory capacity of mitochondria, as well as enhanced production of reactive oxygen species. This study identifies mitochondrial dynamics regulator OPA1 as an interesting therapeutic target in ALS. Additionally, our findings in the adrenal medulla gland from presymptomatic stages highlight the relevance of sympathetic impairment in this disease. Specifically, we show new SOD1^G93A toxicity pathways affecting cellular energy metabolism in non-motor neurons, which offer a possible link between cell specific metabolic phenotype and the progression of ALS. Full article

(This article belongs to the Special Issue Mitochondrial Protein Network: From Biogenesis to Bioenergetics in Health and Disease 2.0)

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19 pages, 7176 KiB

Open AccessArticle

Transcriptomic and Metabolic Network Analysis of Metabolic Reprogramming and IGF-1 Modulation in SCA3 Transgenic Mice

by Yu-Te Lin, Yong-Shiou Lin, Wen-Ling Cheng, Jui-Chih Chang, Yi-Chun Chao, Chin-San Liu and An-Chi Wei

Int. J. Mol. Sci. 2021, 22(15), 7974; https://doi.org/10.3390/ijms22157974 - 26 Jul 2021

Cited by 6 | Viewed by 2735

Abstract

Spinocerebellar ataxia type 3 (SCA3) is a genetic neurodegenerative disease for which a cure is still needed. Growth hormone (GH) therapy has shown positive effects on the exercise behavior of mice with cerebellar atrophy, retains more Purkinje cells, and exhibits less DNA damage [...] Read more.

Spinocerebellar ataxia type 3 (SCA3) is a genetic neurodegenerative disease for which a cure is still needed. Growth hormone (GH) therapy has shown positive effects on the exercise behavior of mice with cerebellar atrophy, retains more Purkinje cells, and exhibits less DNA damage after GH intervention. Insulin-like growth factor 1 (IGF-1) is the downstream mediator of GH that participates in signaling and metabolic regulation for cell growth and modulation pathways, including SCA3-affected pathways. However, the underlying therapeutic mechanisms of GH or IGF-1 in SCA3 are not fully understood. In the present study, tissue-specific genome-scale metabolic network models for SCA3 transgenic mice were proposed based on RNA-seq. An integrative transcriptomic and metabolic network analysis of a SCA3 transgenic mouse model revealed that metabolic signaling pathways were activated to compensate for the metabolic remodeling caused by SCA3 genetic modifications. The effect of IGF-1 intervention on the pathology and balance of SCA3 disease was also explored. IGF-1 has been shown to invoke signaling pathways and improve mitochondrial function and glycolysis pathways to restore cellular functions. As one of the downregulated factors in SCA3 transgenic mice, IGF-1 could be a potential biomarker and therapeutic target. Full article

(This article belongs to the Special Issue Mitochondrial Protein Network: From Biogenesis to Bioenergetics in Health and Disease 2.0)

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19 pages, 4183 KiB

Open AccessArticle

Loss of Individual Mitochondrial Ribonuclease P Complex Proteins Differentially Affects Mitochondrial tRNA Processing In Vivo

by Maithili Saoji, Aditya Sen and Rachel T. Cox

Int. J. Mol. Sci. 2021, 22(11), 6066; https://doi.org/10.3390/ijms22116066 - 04 Jun 2021

Cited by 7 | Viewed by 2796

Abstract

Over a thousand nucleus-encoded mitochondrial proteins are imported from the cytoplasm; however, mitochondrial (mt) DNA encodes for a small number of critical proteins and the entire suite of mt:tRNAs responsible for translating these proteins. Mitochondrial RNase P (mtRNase P) is a three-protein complex [...] Read more.

Over a thousand nucleus-encoded mitochondrial proteins are imported from the cytoplasm; however, mitochondrial (mt) DNA encodes for a small number of critical proteins and the entire suite of mt:tRNAs responsible for translating these proteins. Mitochondrial RNase P (mtRNase P) is a three-protein complex responsible for cleaving and processing the 5′-end of mt:tRNAs. Mutations in any of the three proteins can cause mitochondrial disease, as well as mutations in mitochondrial DNA. Great strides have been made in understanding the enzymology of mtRNase P; however, how the loss of each protein causes mitochondrial dysfunction and abnormal mt:tRNA processing in vivo has not been examined in detail. Here, we used Drosophila genetics to selectively remove each member of the complex in order to assess their specific contributions to mt:tRNA cleavage. Using this powerful model, we find differential effects on cleavage depending on which complex member is lost and which mt:tRNA is being processed. These data revealed in vivo subtleties of mtRNase P function that could improve understanding of human diseases. Full article

(This article belongs to the Special Issue Mitochondrial Protein Network: From Biogenesis to Bioenergetics in Health and Disease 2.0)

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Review

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17 pages, 1966 KiB

Open AccessReview

Mitochondrial Carriers and Substrates Transport Network: A Lesson from Saccharomyces cerevisiae

by Alessandra Ferramosca and Vincenzo Zara

Int. J. Mol. Sci. 2021, 22(16), 8496; https://doi.org/10.3390/ijms22168496 - 07 Aug 2021

Cited by 10 | Viewed by 4849

Abstract

The yeast Saccharomyces cerevisiae is one of the most widely used model organisms for investigating various aspects of basic cellular functions that are conserved in human cells. This organism, as well as human cells, can modulate its metabolism in response to specific growth [...] Read more.

The yeast Saccharomyces cerevisiae is one of the most widely used model organisms for investigating various aspects of basic cellular functions that are conserved in human cells. This organism, as well as human cells, can modulate its metabolism in response to specific growth conditions, different environmental changes, and nutrient depletion. This adaptation results in a metabolic reprogramming of specific metabolic pathways. Mitochondrial carriers play a fundamental role in cellular metabolism, connecting mitochondrial with cytosolic reactions. By transporting substrates across the inner membrane of mitochondria, they contribute to many processes that are central to cellular function. The genome of Saccharomyces cerevisiae encodes 35 members of the mitochondrial carrier family, most of which have been functionally characterized. The aim of this review is to describe the role of the so far identified yeast mitochondrial carriers in cell metabolism, attempting to show the functional connections between substrates transport and specific metabolic pathways, such as oxidative phosphorylation, lipid metabolism, gluconeogenesis, and amino acids synthesis. Analysis of the literature reveals that these proteins transport substrates involved in the same metabolic pathway with a high degree of flexibility and coordination. The understanding of the role of mitochondrial carriers in yeast biology and metabolism could be useful for clarifying unexplored aspects related to the mitochondrial carrier network. Such knowledge will hopefully help in obtaining more insight into the molecular basis of human diseases. Full article

(This article belongs to the Special Issue Mitochondrial Protein Network: From Biogenesis to Bioenergetics in Health and Disease 2.0)

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29 pages, 2371 KiB

Open AccessReview

Mitochondrial HSP70 Chaperone System—The Influence of Post-Translational Modifications and Involvement in Human Diseases

by Henrieta Havalová, Gabriela Ondrovičová, Barbora Keresztesová, Jacob A. Bauer, Vladimír Pevala, Eva Kutejová and Nina Kunová

Int. J. Mol. Sci. 2021, 22(15), 8077; https://doi.org/10.3390/ijms22158077 - 28 Jul 2021

Cited by 28 | Viewed by 6463

Abstract

Since their discovery, heat shock proteins (HSPs) have been identified in all domains of life, which demonstrates their importance and conserved functional role in maintaining protein homeostasis. Mitochondria possess several members of the major HSP sub-families that perform essential tasks for keeping the [...] Read more.

Since their discovery, heat shock proteins (HSPs) have been identified in all domains of life, which demonstrates their importance and conserved functional role in maintaining protein homeostasis. Mitochondria possess several members of the major HSP sub-families that perform essential tasks for keeping the organelle in a fully functional and healthy state. In humans, the mitochondrial HSP70 chaperone system comprises a central molecular chaperone, mtHSP70 or mortalin (HSPA9), which is actively involved in stabilizing and importing nuclear gene products and in refolding mitochondrial precursor proteins, and three co-chaperones (HSP70-escort protein 1—HEP1, tumorous imaginal disc protein 1—TID-1, and Gro-P like protein E—GRPE), which regulate and accelerate its protein folding functions. In this review, we summarize the roles of mitochondrial molecular chaperones with particular focus on the human mtHsp70 and its co-chaperones, whose deregulated expression, mutations, and post-translational modifications are often considered to be the main cause of neurological disorders, genetic diseases, and malignant growth. Full article

(This article belongs to the Special Issue Mitochondrial Protein Network: From Biogenesis to Bioenergetics in Health and Disease 2.0)

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