Abnormal Mitochondrial Dynamics—A Novel Therapeutic Target for Alzheimer's Disease?

Mitochondria are dynamic organelles that undergo continuous fission and fusion, which could affect all aspects of mitochondrial function. Mitochondrial dysfunction has been well documented in Alzheimer’s disease (AD). In the past few years, emerging evidence indicates that an imbalance of mitochondrial dynamics is involved in the pathogenesis of AD. In this review, we discuss in detail the abnormal mitochondrial dynamics in AD and how such abnormal dynamics may impact mitochondrial and neuronal function and contribute to the course of disease. Based on this discussion, we propose that mitochondrial dynamics could be a potential therapeutic target for AD.     

Human F1F0 ATP Synthase,Mitochondrial Ultrastructure and OXPHOS Impairment: A (Super-)Complex Matter?

Mitochondrial morphogenesis is a key process of cell physiology. It is essential for the proper function of this double membrane-delimited organelle, as it ensures the packing of the inner membrane in a very ordered pattern called cristae. In yeast, the mitochondrial ATP synthase is able to form dimers that can assemble into oligomers. Two subunits (e and g) are involved in this supramolecular organization. Deletion of the genes encoding these subunits has no effect on the ATP synthase monomer assembly or activity and only affects its dimerization and oligomerization. Concomitantly, the absence of subunits e and g and thus, of ATP synthase supercomplexes, promotes the modification of mitochondrial ultrastructure suggesting that ATP synthase oligomerization is involved in cristae morphogenesis. We report here that in mammalian cells in culture, the shRNA-mediated down-regulation of subunits e and g affects the stability of ATP synthase and results in a 50% decrease of the available functional enzyme. Comparable to what was shown in yeast, when subunits e and g expression are repressed, ATP synthase dimers and oligomers are less abundant when assayed by native electrophoresis. Unexpectedly, mammalian ATP synthase dimerization/oligomerization impairment has functional consequences on the respiratory chain leading to a decrease in OXPHOS activity. Finally these structural and functional alterations of the ATP synthase have a strong impact on the organelle itself leading to the fission of the mitochondrial network and the disorganization of mitochondrial ultrastructure. Unlike what was shown in yeast, the impairment of the ATP synthase oligomerization process drastically affects mitochondrial ATP production. Thus we propose that mutations or deletions of genes encoding subunits e and g may have physiopathological implications.     

Mitochondrial Protein Import: A Matter of Death?

Mitochondria play a central role not only in energy generation but also for apoptosis. A key step in mitochondrial apoptosis is the release of mitochondrial proteins, most importantly cytochrome c. This release is orchestrated by the pro- and anti-apoptotic members of the Bcl-2 protein family. The functions of these Bcl-2 family members are clear in terms of order and of principle: the pro-apoptotic BH3-only protein group contains the triggers, which cause the activation of the effectors Bax and Bak, while the anti-apoptotic Bcl-2-like proteins prevent this activation. However, the molecular details are still insufficiently clear and the proposed models have certain gaps and are partly contradictory. We have recently presented evidence that targeting to mitochondria of at least one BH3-only protein is essential for its pro-apoptotic functions. Here we discuss how this mechanism might fit into and expand existing models and speculate about the potential implications of this finding.     

Mitochondrial Dysfunction—A Pharmacological Target in Alzheimer's Disease

Increasing evidences suggest that mitochondrial dysfunction plays an important role in the pathogenesis of neurodegenerative diseases including Alzheimer's disease (AD). Alterations of mitochondrial efficiency and function are mainly related to alterations in mitochondrial content, amount of respiratory enzymes, or changes in enzyme activities leading to oxidative stress, mitochondrial permeability transition pore opening, and enhanced apoptosis. More recently, structural changes of the network are related to bioenergetic function, and its consequences are a matter of intensive research. Several mitochondria-targeting compounds with potential efficacy in AD including dimebon, methylene blue, piracetam, simvastatin, Ginkgo biloba, curcumin, and omega-3 polyunsaturated fatty acids have been identified. The majority of preclinical data indicate beneficial effects, whereas most controlled clinical trials did not meet the expectations. Since mitochondrial dysfunction represents an early event in disease progression, one reason for the disappointing clinical results could be that pharmacological interventions might came too late. Thus, more studies are needed that focus on therapeutic strategies starting before severe disease progress.     

Nuclear Recruitment of Neuronal Nitric-oxide Synthase by α-Syntrophin Is Crucial for the Induction of Mitochondrial Biogenesis

Neuronal nitric-oxide synthase (nNOS) has various splicing variants and different subcellular localizations. nNOS can be found also in the nucleus; however, its exact role in this compartment is still not completely defined. In this report, we demonstrate that the PDZ domain allows the recruitment of nNOS to nuclei, thus favoring local NO production, nuclear protein S-nitrosylation, and induction of mitochondrial biogenesis. In particular, overexpression of PDZ-containing nNOS (nNOSα) increases S-nitrosylated CREB with consequent augmented binding on cAMP response element consensus sequence on peroxisome proliferator-activated receptor γ co-activator (PGC)-1α promoter. The resulting PGC-1α induction is accompanied by the expression of mitochondrial genes (e.g., TFAM, MtCO1) and increased mitochondrial mass. Importantly, full active nNOS lacking PDZ domain (nNOSβ) does not localize in nuclei and fails in inducing the expression of PGC-1α. Moreover, we substantiate that the mitochondrial biogenesis normally accompanying myogenesis is associated with nuclear translocation of nNOS. We demonstrate that α-Syntrophin, which resides in nuclei of myocytes, functions as the upstream mediator of nuclear nNOS translocation and nNOS-dependent mitochondrial biogenesis. Overall, our results indicate that altered nNOS splicing and nuclear localization could be contributing factors in human muscular diseases associated with mitochondrial impairment.     

Fast-Evolving Mitochondrial DNA in Ceriantharia: A Reflection of Hexacorallia Paraphyly?

The low evolutionary rate of mitochondrial genes in Anthozoa has challenged their utility for phylogenetic and systematic purposes, especially for DNA barcoding. However, the evolutionary rate of Ceriantharia, one of the most enigmatic “orders” within Anthozoa, has never been specifically examined. In this study, the divergence of mitochondrial DNA of Ceriantharia was compared to members of other Anthozoa and Medusozoa groups. In addition, nuclear markers were used to check the relative phylogenetic position of Ceriantharia in relation to other Cnidaria members. The results demonstrated a pattern of divergence of mitochondrial DNA completely different from those estimated for other anthozoans, and phylogenetic analyses indicate that Ceriantharia is not included within hexacorallians in most performed analyses. Thus, we propose that the Ceriantharia should be addressed as a separate clade.     

Immunohistochemical Localization of Glycogen Synthase and GSK3β: Control of Glycogen Content in Retina

Glycogen has an important role in energy handling in several brain regions. In the brain, glycogen is localized in astrocytes and its role in several normal and pathological processes has been described, whereas in the retina, glycogen metabolism has been scarcely investigated. The enzyme glycogen phosphorylase has been located in retinal Müller cells; however the cellular location of glycogen synthase (GS) and its regulatory partner, glycogen synthase kinase 3β (GSK3β), has not been investigated. Our aim was to localize these enzymes in the rat retina by immunofluorescence techniques. We found both GS and GSK3β in Müller cells in the synaptic layers, and within the inner segments of photoreceptor cells. The presence of these enzymes in Müller cells suggests that glycogen could be regulated within the retina as in other tissues. Indeed, we showed that glycogen content in the whole retina in vitro was increased by high glucose concentrations, glutamate, and insulin. In contrast, retina glycogen levels were not modified by norepinephrine nor by depolarization with high KCl concentrations. Insulin also induced an increase in glycogen content in cultured Müller cells. The effect of insulin in both, whole retina and cultured Müller cells was blocked by inhibitors of phosphatidyl-inositol 3-kinase, strongly suggesting that glycogen content in retina is modulated by the insulin signaling pathway. The expression of GS and GSK3β in the synaptic layers and photoreceptor cells suggests an important role of GSK3β regulating glycogen synthase in neurons, which opens multiple feasible roles of insulin within the retina.     

Synaptotagmin 4: A New Antiobesity Target?

In this issue of Neuron, Zhang et?al. show that Synaptotagmin 4 (Syt4) is specifically induced in adult hypothalamic oxytocin neurons by high-fat diet. Evidence is provided to support a critical role for Syt4 in negative regulation of oxytocin release, which in turn is responsible for diet-induced obesity, raising the possibility of using Syt4 as a new antiobesity target.     

The Nuclear Localization of Glycogen Synthase Kinase 3β Is Required Its Putative PY-Nuclear Localization Sequences

Glycogen synthase kinase-3β (GSK-3β), which is a member of the serine/threonine kinase family, has been shown to be crucial for cellular survival, differentiation, and metabolism. Here, we present evidence that GSK-3β is associated with the karyopherin β2 (Kap β2) (102-kDa), which functions as a substrate for transportation into the nucleus. A potential PY-NLS motif (¹⁰⁹IVRLRYFFY¹¹⁷) was observed, which is similar with the consensus PY NLS motif (R/K/H)X_2–5PY in the GSK-3β catalytic domain. Using a pull down approach, we observed that GSK-3β physically interacts with Kap β2 both in vivo and in vitro. Secondly, GSK-3β and Kap β2 were shown to be co-localized by confocal microscopy. The localization of GSK-3β to the nuclear region was disrupted by putative Kap β2 binding site mutation. Furthermore, in transient transfection assays, the Kap β2 binding site mutant induced a substantial reduction in the in vivo serine/threonine phosphorylation of GSK-3β, where- as the GSK-3β wild type did not. Thus, our observations indicated that Kap β2 imports GSK-3β through its putative PY NLS motif from the cytoplasm to the nucleus and increases its kinase activity.     

What Is the Role of ε in the Escherichia coli ATP Synthase?

The ATP synthase from Escherichia coli is a prototype of the ATP synthases that are found in many bacteria, in the mitochondria of eukaryotes, and in the chloroplasts of plants. It contains eight different types of subunits that have traditionally been divided into F₁, a water-soluble catalytic sector, and F_o, a membrane-bound ion transporting sector. In the current rotary model for ATP synthesis, the subunits can be divided into rotor and stator subunits. Several lines of evidence indicate that is one of the three rotor subunits, which rotate through 360 degrees. The three-dimensional structure of is known and its interactions with other subunits have been explored by several approaches. In light of recent work by our group and that of others, the role of in the ATP synthase from E. coli is discussed.     

Conformational Transitions of Subunit ? in ATP Synthase from Thermophilic Bacillus PS3

Subunit ɛ of bacterial and chloroplast F_OF₁-ATP synthase is responsible for inhibition of ATPase activity. In Bacillus PS3 enzyme, subunit ɛ can adopt two conformations. In the “extended”, inhibitory conformation, its two C-terminal α-helices are stretched along subunit γ. In the “contracted”, noninhibitory conformation, these helices form a hairpin. The transition of subunit ɛ from an extended to a contracted state was studied in ATP synthase incorporated in Bacillus PS3 membranes at 59°C. Fluorescence energy resonance transfer between fluorophores introduced in the C-terminus of subunit ɛ and in the N-terminus of subunit γ was used to follow the conformational transition in real time. It was found that ATP induced the conformational transition from the extended to the contracted state (half-maximum transition extent at 140 μM ATP). ADP could neither prevent nor reverse the ATP-induced conformational change, but it did slow it down. Acid residues in the DELSEED region of subunit β were found to stabilize the extended conformation of ɛ. Binding of ATP directly to ɛ was not essential for the ATP-induced conformational change. The ATP concentration necessary for the half-maximal transition (140 μM) suggests that subunit ɛ probably adopts the extended state and strongly inhibits ATP hydrolysis only when the intracellular ATP level drops significantly below the normal value.     

7 The Molecular Biology of Tryptophan Synthase: A Model for Protein–Protein Interaction

Abstract

The use of plastic produced from non-renewable resources constitutes a major environmental problem of the modern society. Polylactide polymers (PLA) have recently gained enormous attention as one possible substitution of petroleum derived polymers. A prerequisite for high quality PLA production is the provision of optically pure lactic acid, which cannot be obtained by chemical synthesis in an economical way. Microbial fermentation is therefore the commercial option to obtain lactic acid as monomer for PLA production. However, one major economic hurdle for commercial lactic acid production as basis for PLA is the costly separation procedure, which is needed to recover and purify the product from the fermentation broth. Yeasts, such as Saccharomyces cerevisiae (bakers yeast) offer themselves as production organisms because they can tolerate low pH and grow on mineral media what eases the purification of the acid. However, naturally yeasts do not produce lactic acid. By metabolic engineering, ethanol was exchanged with lactic acid as end product of fermentation. A vast amount of effort has been invested into the development of yeasts for lactic acid production since the first paper on this topic by Dequin and process insight. If pH stress is used as basis for DNA microarray analyses, in order to improve the host, what exactly is addressed? Growth? Or productivity? They might be connected, but can be negatively correlated. A better growing strain might not be a better producer. So if the question was growth, the answer might not be what was initially intended (productivity).

A major task for the future is to learn to ask the right questions – a lot of studies intended to lead to better productivity, did lead to interesting results, but NOT to better production strains.

Taking together what we learned from lactic acid production with yeasts, we see a bright future for bulk and fine chemical production with these versatile hosts.     

Aquifex aeolicus 3-Deoxy-d-manno-2-Octulosonic Acid 8-Phosphate Synthase: A New Class of KDO 8-P Synthase?

The relationship between 3-deoxy-D-manno-2-octulosonic acid 8-phosphate (KDO 8-P) synthase and 3-deoxy-D-arabino-2-heptulosonic acid 7-phosphate (DAH 7-P) synthase has not been adequately addressed in the literature. Based on recent reports of a metal requiring KDO 8-P synthase and the newly solved X-ray crystal structures of both Escherichia coli KDO 8-P synthase and DAH 7-P synthase, we begin to address the evolutionary kinship between these catalytically similar enzymes. Using a maximum likelihood-based grouping of 29 KDO 8-P synthase sequences, we demonstrate the existence of a new class of KDO 8-P synthase, the members of which we propose to require a metal cofactor for catalysis. Similarly, we hypothesize a class of DAH 7-P synthase that does not have the metal requirement of the heretofore model E. coli enzyme. Based on this information and a careful investigation of the reported X-ray crystal structures, we also propose that KDO 8-P synthase and DAH 7-P synthase are the product of a divergent evolutionary process from a common ancestor.     

Type I Interferon: Potential Therapeutic Target for Psoriasis?

Background

Psoriasis is an immune-mediated disease characterized by aberrant epidermal differentiation, surface scale formation, and marked cutaneous inflammation. To better understand the pathogenesis of this disease and identify potential mediators, we used whole genome array analysis to profile paired lesional and nonlesional psoriatic skin and skin from healthy donors.

Methodology/Principal Findings

We observed robust overexpression of type I interferon (IFN)–inducible genes and genomic signatures that indicate T cell and dendritic cell infiltration in lesional skin. Up-regulation of mRNAs for IFN-α subtypes was observed in lesional skin compared with nonlesional skin. Enrichment of mature dendritic cells and 2 type I IFN–inducible proteins, STAT1 and ISG15, were observed in the majority of lesional skin biopsies. Concordant overexpression of IFN-γ and TNF-α–inducible gene signatures occurred at the same disease sites.

Conclusions/Significance

Up-regulation of TNF-α and elevation of the TNF-α–inducible gene signature in lesional skin underscore the importance of this cytokine in psoriasis; these data describe a molecular basis for the therapeutic activity of anti–TNF-α agents. Furthermore, these findings implicate type I IFNs in the pathogenesis of psoriasis. Consistent and significant up-regulation of type I IFNs and their associated gene signatures in psoriatic skin suggest that type I IFNs may be potential therapeutic targets in psoriasis treatment.     

Immunohistochemical Localization of Mitochondrial Fatty Acid ��-Oxidation Enzymes in Rat Testis

The testis consists of two types of tissues, the interstitial tissue and the seminiferous tubule, which have different functions and are assumed to have different nutritional metabolism. The localization of enzymes of the mitochondrial fatty acid β-oxidation system in the testis was investigated to obtain a better understanding of nutrient metabolism in the testis. Adult rat testis tissues were subjected to immunoblot analysis for quantitation of the amounts of enzyme proteins, to DNA microarray analysis for gene expression, and to immunofluorescence and immunoelectron microscopy for localization. Quantitative analysis by immunoblot and DNA microarray revealed that enzymes occur abundantly in Leydig cells in the interstitial tissue but much less so in the seminiferous tubules. Immunohistochemistry revealed that Leydig cells in the interstitial tissue and Sertoli cells in the seminiferous tubules contain a full set of mitochondrial fatty acid β-oxidation enzymes in relatively plentiful amounts among the cells in the testis, but that this is not so in spermatogenic cells. This characteristic localization of the mitochondrial fatty acid β-oxidation system in the testis needs further elucidation in terms of a possible role for it in the nutritional metabolism of spermatogenesis. (J Histochem Cytochem 58:195–206, 2010)     

Structure and Function of ε-Subunit of ATP Synthase in Higher Plant Chloroplast

Theε-subunit is the smallest subunit of chloroplast ATP synthase, and is known to inhibit ATPase activity in isolated CF1-ATPase. As a result ε is sometimes called an inhibitory subunit. In addition, and perhaps more importantly, theε-subunit is essential for the coupling of proton translocation to ATP synthesis (as proton gate). The relation between the structure and function ofε-subunit of ATP synthase in higher plant chloroplast has been studied by molecular biological methods such as site-directed mu-tagenesis and truncations for C- or N-terminus ofε-subunit. The results showed that: (1) Thr42 ofε-subunit is important for its structure and function; (2) compared with theε-subunit in E.. coli, theε-subunit of chloroplast ATP synthase is more sensitive to C- or N-terminus truncations.     

Mitochondrial translation in trypanosomatids: a novel target for chemotherapy?

Trypanosomatids cause widespread disease in humans and animals. Treatment of many of these diseases is hampered by the lack of efficient and safe drugs. New strategies for drug development are therefore urgently needed. It has long been known that the single mitochondrion of trypanosomatids exhibits many unique features. Recently, the mitochondrial translation machinery of trypanosomatids has been the focus of several studies, which revealed interesting variations to the mammalian system. It is the aim of this article to review these unique features and to discuss them in the larger biological context. It is our opinion that some of these features represent promising novel targets for chemotherapeutic intervention that should be studied in more detail.