Regulation of Mitochondrial Respiration by VDAC Is Enhanced by Membrane-Bound Inhibitors with Disordered Polyanionic C-Terminal Domains

The voltage-dependent anion channel (VDAC) is the primary regulating pathway of water-soluble metabolites and ions across the mitochondrial outer membrane. When reconstituted into lipid membranes, VDAC responds to sufficiently large transmembrane potentials by transitioning to gated states in which ATP/ADP flux is reduced and calcium flux is increased. Two otherwise unrelated cytosolic proteins, tubulin, and α-synuclein (αSyn), dock with VDAC by a novel mechanism in which the transmembrane potential draws their disordered, polyanionic C-terminal domains into and through the VDAC channel, thus physically blocking the pore. For both tubulin and αSyn, the blocked state is observed at much lower transmembrane potentials than VDAC gated states, such that in the presence of these cytosolic docking proteins, VDAC’s sensitivity to transmembrane potential is dramatically increased. Remarkably, the features of the VDAC gated states relevant for bioenergetics—reduced metabolite flux and increased calcium flux—are preserved in the blocked state induced by either docking protein. The ability of tubulin and αSyn to modulate mitochondrial potential and ATP production in vivo is now supported by many studies. The common physical origin of the interactions of both tubulin and αSyn with VDAC leads to a general model of a VDAC inhibitor, facilitates predictions of the effect of post-translational modifications of known inhibitors, and points the way toward the development of novel therapeutics targeting VDAC.     

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

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.     

ANT1 Activation and Inhibition Patterns Support the Fatty Acid Cycling Mechanism for Proton Transport

Adenine nucleotide translocase (ANT) is a well-known mitochondrial exchanger of ATP against ADP. In contrast, few studies have shown that ANT also mediates proton transport across the inner mitochondrial membrane. The results of these studies are controversial and lead to different hypotheses about molecular transport mechanisms. We hypothesized that the H⁺-transport mediated by ANT and uncoupling proteins (UCP) has a similar regulation pattern and can be explained by the fatty acid cycling concept. The reconstitution of purified recombinant ANT1 in the planar lipid bilayers allowed us to measure the membrane current after the direct application of transmembrane potential ΔΨ, which would correspond to the mitochondrial states III and IV. Experimental results reveal that ANT1 does not contribute to a basal proton leak. Instead, it mediates H⁺ transport only in the presence of long-chain fatty acids (FA), as already known for UCPs. It depends on FA chain length and saturation, implying that FA’s transport is confined to the lipid-protein interface. Purine nucleotides with the preference for ATP and ADP inhibited H⁺ transport. Specific inhibitors of ATP/ADP transport, carboxyatractyloside or bongkrekic acid, also decreased proton transport. The H⁺ turnover number was calculated based on ANT1 concentration determined by fluorescence correlation spectroscopy and is equal to 14.6 ± 2.5 s⁻¹. Molecular dynamic simulations revealed a large positively charged area at the protein/lipid interface that might facilitate FA anion’s transport across the membrane. ANT’s dual function—ADP/ATP and H⁺ transport in the presence of FA—may be important for the regulation of mitochondrial membrane potential and thus for potential-dependent processes in mitochondria. Moreover, the expansion of proton-transport modulating drug targets to ANT1 may improve the therapy of obesity, cancer, steatosis, cardiovascular and neurodegenerative diseases.     

Xanthohumol-Induced Rat Glioma C6 Cells Death by Triggering Mitochondrial Stress

AIM: To investigate the underlying mechanisms of xanthohumol (XN) on the proliferation inhibition and death of C6 glioma cells. METHODS: To determine the effects of XN on C6 cells, cell proliferation and mortality after XN treatment were assessed by SRB assay and trypan blue assay respectively. Apoptotic rates were evaluated by flowcytometry after Annexin V-FITC/PI double staining. The influence of XN on the activity of caspase-3 was determined by Western blot (WB); and nuclear transposition of apoptosis-inducing factor (AIF) was tested by immunocytochemistry and WB. By MitoSOX^TM staining, the mitochondrial ROS were detected. Mitochondrial function was also tested by MTT assay (content of succinic dehydrogenase), flow cytometry (mitochondrial membrane potential (MMP)—JC-1 staining; mitochondrial abundance—mito-Tracker green), immunofluorescence (MMP—JC-1 staining; mitochondrial morphology—mito-Tracker green), WB (mitochondrial fusion-fission protein—OPA1, mfn2, and DRP1; mitophagy-related proteins—Pink1, Parkin, LC3B, and P62), and high-performance liquid chromatography (HPLC) (energy charge). Finally, mitochondrial protein homeostasis of C6 cells after XN treatment with and without LONP1 inhibitor bortezomib was investigated by trypan blue assay (proliferative activity and mortality) and WB (mitochondrial protease LONP1). All cell morphology images were taken by a Leica Microsystems microscope. RESULTS: XN could lead to proliferation inhibition and death of C6 cells in a time- and dose-dependent manner and induce apoptosis of C6 cells through the AIF pathway. After long incubation of XN, mitochondria of C6 cells were seriously impaired, and mitochondria had a diffuse morphology and mitochondrial ROS were increased. The content of succinic dehydrogenase per cell was significantly decreased after XN insults of 24, 48, and 72 h. The energy charge was weakened after XN insult of 24 h. Furthermore, the MMP and mitochondrial abundance were significantly decreased; the protein expression levels of OPA1, mfn2, and DRP1 were down-regulated; and the protein expression levels of Pink1, Parkin, LC3B-II/LC3B-I, and p62 were up-regulated in long XN incubation times (24, 48, and 72 h). XN incubation with bortezomib for 48 h resulted in lower proliferative activity and higher mortality of C6 cells and caused the cell to have visible vacuoles. Moreover, the protein expression levels of LONP1 was up-regulated gradually as XN treatment time increased. CONCLUSION: These data supported that XN could induce AIF pathway apoptosis of the rat glioma C6 cells by affecting the mitochondria.     

Influence of succinylation on the properties of cast films from red bean protein isolate at various plasticizer levels

The effects of succinylation at three anhydride levels (0.2, 0.4, and 0.6 g g⁻¹) on the properties of cast films from red bean protein isolate (RPI) at three glycerol levels of 0.2, 0.4, and 0.6 g g⁻¹ were investigated. The tested properties included tensile strength (TS) and elongation at break (EB), surface hydrophobicity, moisture content (MC), total soluble matter (TSM), water vapor transmission rate, and permeability (WVTR and WVP), permeability coefficient of oil (PO). The results showed that the succinylation greatly improved the mechanical properties (especially the EB), but decreased the surface hydrophobicity of cast films. The MC, TSM, WVTR, WVP, and PO were considerably increased by the succinylation. The size exclusion chromatography analysis indicated that the succinylation resulted in protein aggregation or association, and transformation of insoluble precipitates (initially present) to soluble protein components. The dependence of the influence of succinylation upon some selected properties on the plasticizer level suggests interactions between introduced anionic succinic moieties and the hydroxyl groups of the plasticizer. These results suggest that the succinylation treatment could be applied to modify the mechanical properties of legume proteins, especially in the case that requires excellent flexibility of cast films. © 2010 Wiley Periodicals, Inc. J Appl Polym Sci, 2011     

Proteomics and Post-Translational Modifications of Starch Biosynthesis-Related Proteins in Developing Seeds of Rice

Rice (Oryza sativa L.) is a foremost staple food for approximately half the world’s population. The components of rice starch, amylose, and amylopectin are synthesized by a series of enzymes, which are responsible for rice starch properties and functionality, and then affect rice cooking and eating quality. Recently, proteomics technology has been applied to the establishment of the differentially expressed starch biosynthesis-related proteins and the identification of posttranslational modifications (PTMs) target starch biosynthesis proteins as well. It is necessary to summarize the recent studies in proteomics and PTMs in rice endosperm to deepen our understanding of starch biosynthesis protein expression and regulation, which will provide useful information to rice breeding programs and industrial starch applications. The review provides a comprehensive summary of proteins and PTMs involved in starch biosynthesis based on proteomic studies of rice developing seeds. Starch biosynthesis proteins in rice seeds were differentially expressed in the developing seeds at different developmental stages. All the proteins involving in starch biosynthesis were identified using proteomics methods. Most starch biosynthesis-related proteins are basically increased at 6–20 days after flowering (DAF) and decreased upon the high-temperature conditions. A total of 10, 14, 2, 17, and 7 starch biosynthesis related proteins were identified to be targeted by phosphorylation, lysine acetylation, succinylation, lysine 2-hydroxyisobutyrylation, and malonylation, respectively. The phosphoglucomutase is commonly targeted by five PTMs types. Research on the function of phosphorylation in multiple enzyme complex formation in endosperm starch biosynthesis is underway, while the functions of other PTMs in starch biosynthesis are necessary to be conducted in the near future.     

Therapeutic Application of Extracellular Vesicles-Capsulated Adeno-Associated Virus Vector via nSMase2/Smpd3, Satellite,and Immune Cells in Duchenne Muscular Dystrophy

Duchenne muscular dystrophy (DMD) is caused by loss-of-function mutations in the dystrophin gene on chromosome Xp21. Disruption of the dystrophin–glycoprotein complex (DGC) on the cell membrane causes cytosolic Ca²⁺ influx, resulting in protease activation, mitochondrial dysfunction, and progressive myofiber degeneration, leading to muscle wasting and fragility. In addition to the function of dystrophin in the structural integrity of myofibers, a novel function of asymmetric cell division in muscular stem cells (satellite cells) has been reported. Therefore, it has been suggested that myofiber instability may not be the only cause of dystrophic degeneration, but rather that the phenotype might be caused by multiple factors, including stem cell and myofiber functions. Furthermore, it has been focused functional regulation of satellite cells by intracellular communication of extracellular vesicles (EVs) in DMD pathology. Recently, a novel molecular mechanism of DMD pathogenesis—circulating RNA molecules—has been revealed through the study of target pathways modulated by the Neutral sphingomyelinase2/Neutral sphingomyelinase3 (nSMase2/Smpd3) protein. In addition, adeno-associated virus (AAV) has been clinically applied for DMD therapy owing to the safety and long-term expression of transduction genes. Furthermore, the EV-capsulated AAV vector (EV-AAV) has been shown to be a useful tool for the intervention of DMD, because of the high efficacy of the transgene and avoidance of neutralizing antibodies. Thus, we review application of AAV and EV-AAV vectors for DMD as novel therapeutic strategy.     

Chimeric Claudins: A New Tool to Study Tight Junction Structure and Function

The tight junction (TJ) is a structure composed of multiple proteins, both cytosolic and membranal, responsible for cell–cell adhesion in polarized endothelium and epithelium. The TJ is intimately connected to the cytoskeleton and plays a role in development and homeostasis. Among the TJ’s membrane proteins, claudins (CLDNs) are key to establishing blood–tissue barriers that protect organismal physiology. Recently, several crystal structures have been reported for detergent extracted recombinant CLDNs. These structural advances lack direct evidence to support quaternary structure of CLDNs. In this article, we have employed protein-engineering principles to create detergent-independent chimeric CLDNs, a combination of a 4-helix bundle soluble monomeric protein (PDB ID: 2jua) and the apical—50% of human CLDN1, the extracellular domain that is responsible for cell–cell adhesion. Maltose-binding protein-fused chimeric CLDNs (MBP-CCs) used in this study are soluble proteins that retain structural and functional aspects of native CLDNs. Here, we report the biophysical characterization of the structure and function of MBP-CCs. MBP-fused epithelial cadherin (MBP-eCAD) is used as a control and point of comparison of a well-characterized cell-adhesion molecule. Our synthetic strategy may benefit other families of 4-α-helix membrane proteins, including tetraspanins, connexins, pannexins, innexins, and more.     

Folding and Biogenesis of Mitochondrial Small Tim Proteins

Correct and timely folding is critical to the function of all proteins. The importance of this is illustrated in the biogenesis of the mitochondrial intermembrane space (IMS) “small Tim” proteins. Biogenesis of the small Tim proteins is regulated by dedicated systems or pathways, beginning with synthesis in the cytosol and ending with assembly of individually folded proteins into functional complexes in the mitochondrial IMS. The process is mostly centered on regulating the redox states of the conserved cysteine residues: oxidative folding is crucial for protein function in the IMS, but oxidized (disulfide bonded) proteins cannot be imported into mitochondria. How the redox-sensitive small Tim precursor proteins are maintained in a reduced, import-competent form in the cytosol is not well understood. Recent studies suggest that zinc and the cytosolic thioredoxin system play a role in the biogenesis of these proteins. In the IMS, the mitochondrial import and assembly (MIA) pathway catalyzes both import into the IMS and oxidative folding of the small Tim proteins. Finally, assembly of the small Tim complexes is a multistep process driven by electrostatic and hydrophobic interactions; however, the chaperone function of the complex might require destabilization of these interactions to accommodate the substrate. Here, we review how folding of the small Tim proteins is regulated during their biogenesis, from maintenance of the unfolded precursors in the cytosol, to their import, oxidative folding, complex assembly and function in the IMS.     

Identification of Novel Therapeutic Targets for Polyglutamine Diseases That Target Mitochondrial Fragmentation

Huntington’s disease (HD) is one of at least nine polyglutamine diseases caused by a trinucleotide CAG repeat expansion, all of which lead to age-onset neurodegeneration. Mitochondrial dynamics and function are disrupted in HD and other polyglutamine diseases. While multiple studies have found beneficial effects from decreasing mitochondrial fragmentation in HD models by disrupting the mitochondrial fission protein DRP1, disrupting DRP1 can also have detrimental consequences in wild-type animals and HD models. In this work, we examine the effect of decreasing mitochondrial fragmentation in a neuronal C. elegans model of polyglutamine toxicity called Neur-67Q. We find that Neur-67Q worms exhibit mitochondrial fragmentation in GABAergic neurons and decreased mitochondrial function. Disruption of drp-1 eliminates differences in mitochondrial morphology and rescues deficits in both movement and longevity in Neur-67Q worms. In testing twenty-four RNA interference (RNAi) clones that decrease mitochondrial fragmentation, we identified eleven clones—each targeting a different gene—that increase movement and extend lifespan in Neur-67Q worms. Overall, we show that decreasing mitochondrial fragmentation may be an effective approach to treating polyglutamine diseases and we identify multiple novel genetic targets that circumvent the potential negative side effects of disrupting the primary mitochondrial fission gene drp-1.     

Contribution of Massive Mitochondrial Fusion and Subsequent Fission in the Plant Life Cycle to the Integrity of the Mitochondrion and Its Genome

Plant mitochondria have large genomes to house a small number of key genes. Most mitochondria do not contain a whole genome. Despite these latter characteristics, the mitochondrial genome is faithfully maternally inherited. To maintain the mitochondrial genes—so important for energy production—the fusion and fission of mitochondria are critical. Fission in plants is better understood than fusion, with the dynamin-related proteins (DRP 3A and 3B) driving the constriction of the mitochondrion. How the endoplasmic reticulum and the cytoskeleton are linked to the fission process is not yet fully understood. The fusion mechanism is less well understood, as obvious orthologues are not present. However, there is a recently described gene, MIRO2, that appears to have a significant role, as does the ER and cytoskeleton. Massive mitochondrial fusion (MMF or hyperfusion) plays a significant role in plants. MMF occurs at critical times of the life cycle, prior to flowering, in the enlarging zygote and at germination, mixing the cells’ mitochondrial population—the so-called “discontinuous whole”. MMF in particular aids genome repair, the conservation of critical genes and possibly gives an energy boost to important stages of the life cycle. MMF is also important in plant regeneration, an important component of plant biotechnology.     

Functional and nutritional properties of acylated rapeseed proteins

Rapeseed flour was acylated to various degrees with acetic and succinic anhydrides to produce acetylated rapeseed protein concentrates (SRPC) and succinylated rapeseed protein concentrates (SRPC), respectively. With both acylating agents, approximately 5–82% (ARPC-5 to ARPC-82) and 5–56% (SRPC-5 to SRPC-56) of ε-amino groups of lysine were acylated. Changes in functional properties were monitored; nitrogen solubility, emulsifying properties and specific viscosity were improved by acylation. The effects on functional properties were more pronounced with succinylation than acetylation. The nutritive value of succinylated rapeseed protein concentrates was determined and compared with those of unmodified rapessed protein concentrate and rapessed flour. Succinylation reduced the availability of lysine and decreased (p<0.05) the net protein ratio (NPR) and apparent digestibility coefficient (ADC) of nitrogen in rapessed protein concentrates. Supplementation of SRPC with lysine improved the NPR compared to that of SRPC-56.     

Interference on Cytosolic DNA Activation Attenuates Sepsis Severity: Experiments on Cyclic GMP–AMP Synthase (cGAS) Deficient Mice

Although the enhanced responses against serum cell-free DNA (cfDNA) in cases of sepsis—a life-threatening organ dysfunction due to systemic infection—are understood, the influence of the cytosolic DNA receptor cGAS (cyclic guanosine monophosphate–adenosine monophosphate (GMP–AMP) synthase) on sepsis is still unclear. Here, experiments on cGAS deficient (cGAS^-/-) mice were conducted using cecal ligation and puncture (CLP) and lipopolysaccharide (LPS) injection sepsis models and macrophages. Severity of CLP in cGAS^-/- mice was less severe than in wildtype (WT) mice, as indicated by mortality, serum LPS, cfDNA, leukopenia, cytokines (TNF-α, IL-6 and IL-10), organ histology (lung, liver and kidney) and spleen apoptosis. With the LPS injection model, serum cytokines in cGAS^-/- mice were lower than in WT mice, despite the similar serum cfDNA level. Likewise, in LPS-activated WT macrophages, the expression of several mitochondria-associated genes (as revealed by RNA sequencing analysis) and a profound reduction in mitochondrial parameters, including maximal respiration (determined by extracellular flux analysis), DNA (mtDNA) and mitochondrial abundance (revealed by fluorescent staining), were demonstrated. These data implied the impact of cfDNA resulting from LPS-induced cell injury. In parallel, an additive effect of bacterial DNA on LPS, seen in comparison with LPS alone, was demonstrated in WT macrophages, but not in cGAS^-/- cells, as indicated by supernatant cytokines (TNF-α and IL-6), M1 proinflammatory polarization (iNOS and IL-1β), cGAS, IFN-γ and supernatant cyclic GMP–AMP (cGAMP). In conclusion, cGAS activation by cfDNA from hosts (especially mtDNA) and bacteria was found to induce an additive proinflammatory effect on LPS-activated macrophages which was perhaps responsible for the more pronounced sepsis hyperinflammation observed in WT mice compared with the cGAS^-/- group.     

The ɛ‐Amino Group of Protein Lysine Residues Is Highly Susceptible to Nonenzymatic Acylation by Several Physiological Acyl‐CoA Thioesters

Mitochondrial enzymes implicated in the pathophysiology of diabetes, cancer, and metabolic syndrome are highly regulated by acetylation. However, mitochondrial acetyltransferases have not been identified. Here, we show that acetylation and also other acylations are spontaneous processes that depend on pH value, acyl‐CoA concentration and the chemical nature of the acyl residue. In the case of a peptide derived from carbamoyl phosphate synthetase 1, the rates of succinylation and glutarylation were up to 150 times than for acetylation. These results were confirmed by using the protein substrate cyclophilin A (CypA). Deacylation experiments revealed that SIRT3 exhibits deacetylase activity but is not able to remove any of the succinyl groups from CypA, whereas SIRT5 is an effective protein desuccinylase. Thus, the acylation landscape on lysine residues might largely depend on the enzymatic activity of specific sirtuins, and the availability and reactivity of acyl‐CoA compounds.     

A Molecular Perspective on Sirtuin Activity

The protein acetylation of either the α-amino groups of amino-terminal residues or of internal lysine or cysteine residues is one of the major posttranslational protein modifications that occur in the cell with repercussions at the protein as well as at the metabolome level. The lysine acetylation status is determined by the opposing activities of lysine acetyltransferases (KATs) and lysine deacetylases (KDACs), which add and remove acetyl groups from proteins, respectively. A special group of KDACs, named sirtuins, that require NAD⁺ as a substrate have received particular attention in recent years. They play critical roles in metabolism, and their abnormal activity has been implicated in several diseases. Conversely, the modulation of their activity has been associated with protection from age-related cardiovascular and metabolic diseases and with increased longevity. The benefits of either activating or inhibiting these enzymes have turned sirtuins into attractive therapeutic targets, and considerable effort has been directed toward developing specific sirtuin modulators. This review summarizes the protein acylation/deacylation processes with a special focus on the current developments in the sirtuin research field.     

The Antibiotic Doxycycline Impairs Cardiac Mitochondrial and Contractile Function

Tetracycline antibiotics act by inhibiting bacterial protein translation. Given the bacterial ancestry of mitochondria, we tested the hypothesis that doxycycline—which belongs to the tetracycline class—reduces mitochondrial function, and results in cardiac contractile dysfunction in cultured H9C2 cardiomyoblasts, adult rat cardiomyocytes, in Drosophila and in mice. Ampicillin and carbenicillin were used as control antibiotics since these do not interfere with mitochondrial translation. In line with its specific inhibitory effect on mitochondrial translation, doxycycline caused a mitonuclear protein imbalance in doxycycline-treated H9C2 cells, reduced maximal mitochondrial respiration, particularly with complex I substrates, and mitochondria appeared fragmented. Flux measurements using stable isotope tracers showed a shift away from OXPHOS towards glycolysis after doxycycline exposure. Cardiac contractility measurements in adult cardiomyocytes and Drosophila melanogaster hearts showed an increased diastolic calcium concentration, and a higher arrhythmicity index. Systolic and diastolic dysfunction were observed after exposure to doxycycline. Mice treated with doxycycline showed mitochondrial complex I dysfunction, reduced OXPHOS capacity and impaired diastolic function. Doxycycline exacerbated diastolic dysfunction and reduced ejection fraction in a diabetes mouse model vulnerable for metabolic derangements. We therefore conclude that doxycycline impairs mitochondrial function and causes cardiac dysfunction.     

tRNA-Dependent Import of a Transit Sequence-Less Aminoacyl-tRNA Synthetase (LeuRS2) into the Mitochondria of Arabidopsis

Aminoacyl-tRNA synthetases (AaRS) charge tRNAs with amino acids for protein translation. In plants, cytoplasmic, mitochondrial, and chloroplast AaRS exist that are all coded for by nuclear genes and must be imported from the cytosol. In addition, only a few of the mitochondrial tRNAs needed for translation are encoded in mitochondrial DNA. Despite considerable progress made over the last few years, still little is known how the bulk of cytosolic AaRS and respective tRNAs are transported into mitochondria. Here, we report the identification of a protein complex that ties AaRS and tRNA import into the mitochondria of Arabidopsis thaliana. Using leucyl-tRNA synthetase 2 (LeuRS2) as a model for a mitochondrial signal peptide (MSP)-less precursor, a ≈30 kDa protein was identified that interacts with LeuRS2 during import. The protein identified is identical with a previously characterized mitochondrial protein designated HP30-2 (encoded by At3g49560) that contains a sterile alpha motif (SAM) similar to that found in RNA binding proteins. HP30-2 is part of a larger protein complex that contains with TIM22, TIM8, TIM9 and TIM10 four previously identified components of the translocase for MSP-less precursors. Lack of HP30-2 perturbed mitochondrial biogenesis and function and caused seedling lethality during greening, suggesting an essential role of HP30-2 in planta.     

Mitochondrial Unfolded Protein Responses in White Adipose Tissue: Lipoatrophy,Whole-Body Metabolism and Lifespan

The mitochondrial unfolded protein response (UPR^mt) is a stress response mediated by the expression of genes such as chaperones, proteases, and mitokines to maintain mitochondrial proteostasis. Certain genetically modified mice, which defect mitochondrial proteins specifically in adipocytes, developed atrophy of the white adipose tissue, resisted diet-induced obesity, and had altered whole-body metabolism. UPR^mt, which has beneficial functions for living organisms, is termed “mitohormesis”, but its specific characteristics and detailed regulatory mechanism have not been elucidated to date. In this review, we discuss the function of UPR^mt in adipose atrophy (lipoatrophy), whole-body metabolism, and lifespan based on the concept of mitohormesis.     

Molecular and Functional Characterization of MobK Protein—A Novel-Type Relaxase Involved in Mobilization for Conjugational Transfer of Klebsiella pneumoniae Plasmid pIGRK

Conjugation, besides transformation and transduction, is one of the main mechanisms of horizontal transmission of genetic information among bacteria. Conjugational transfer, due to its essential role in shaping bacterial genomes and spreading of antibiotics resistance genes, has been widely studied for more than 70 years. However, new and intriguing facts concerning the molecular basis of this process are still being revealed. Most recently, a novel family of conjugative relaxases (Mob proteins) was distinguished. The characteristic feature of these proteins is that they are not related to any of Mobs described so far. Instead of this, they share significant similarity to tyrosine recombinases. In this study MobK—a tyrosine recombinase-like Mob protein, encoded by pIGRK cryptic plasmid from the Klebsiella pneumoniae clinical strain, was characterized. This study revealed that MobK is a site-specific nuclease and its relaxase activity is dependent on both a conserved catalytic tyrosine residue (Y¹⁷⁹) that is characteristic of tyrosine recombinases and the presence of Mg²⁺ divalent cations. The pIGRK minimal origin of transfer sequence (oriT) was also characterized. This is one of the first reports presenting tyrosine recombinase-like conjugative relaxase protein. It also demonstrates that MobK is a convenient model for studying this new protein family.