The transmembrane prolines of the mitochondrial ADP/ATP carrier are involved in nucleotide binding and transport and its biogenesis

The mitochondrial ADP/ATP carrier (Ancp) is a paradigm of the mitochondrial carrier family, which allows cross-talk between mitochondria, where cell energy is mainly produced, and cytosol, where cell energy is mainly consumed. The members of this family share numerous structural and functional characteristics. Resolution of the atomic structure of the bovine Ancp, in a complex with one of its specific inhibitors, revealed interesting features and suggested the involvement of some particular residues in the movements of the protein to perform translocation of nucleotides from one side of the membrane to the other. They correspond to three prolines located in the odd-numbered transmembrane helices (TMH), Pro-27, Pro-132, and Pro-229. The corresponding residues of the yeast Ancp (Pro-43, Ser-147, and Pro-247) were mutated into alanine or leucine, one at a time and analysis of the various mutants evidenced a crucial role of Pro-43 and Pro-247 during nucleotide transport. Beside, replacement of Ser-147 with proline does not inactivate Ancp and this is discussed in view of the conservation of the three prolines at equivalent positions in the Ancp sequences. These prolines belong to the signature sequences of the mitochondrial carriers and we propose they play a dual role in the mitochondrial ADP/ATP carrier function and biogenesis. Unexpectedly their mutations cause more general effects on mitochondrial biogenesis and morphology, as evidenced by measurements of respiratory rates, cytochrome contents, and also clearly highlighted by fluorescence microscopy.     

Regulation of mitochondrial bioenergetics by the non-canonical roles of mitochondrial dynamics proteins in the heart

Recent advancement in mitochondrial research has significantly extended our knowledge on the role and regulation of mitochondria in health and disease. One important breakthrough is the delineation of how mitochondrial morphological changes, termed mitochondrial dynamics, are coupled to the bioenergetics and signaling functions of mitochondria. In general, it is believed that fusion leads to an increased mitochondrial respiration efficiency and resistance to stress-induced dysfunction while fission does the contrary. This concept seems not applicable to adult cardiomyocytes. The mitochondria in adult cardiomyocytes exhibit fragmented morphology (tilted towards fission) and show less networking and movement as compared to other cell types. However, being the most energy-demanding cells, cardiomyocytes in the adult heart possess vast number of mitochondria, high level of energy flow, and abundant mitochondrial dynamics proteins. This apparent discrepancy could be explained by recently identified new functions of the mitochondrial dynamics proteins. These “non-canonical” roles of mitochondrial dynamics proteins range from controlling inter-organelle communication to regulating cell viability and survival under metabolic stresses. Here, we summarize the newly identified non-canonical roles of mitochondrial dynamics proteins. We focus on how these fission and fusion independent roles of dynamics proteins regulate mitochondrial bioenergetics. We also discuss potential molecular mechanisms, unique intracellular location, and the cardiovascular disease relevance of these non-canonical roles of the dynamics proteins. We propose that future studies are warranted to differentiate the canonical and non-canonical roles of dynamics proteins and to identify new approaches for the treatment of heart diseases. This article is part of a Special issue entitled Cardiac adaptations to obesity, diabetes and insulin resistance, edited by Professors Jan F.C. Glatz, Jason R.B. Dyck and Christine Des Rosiers.     

PGAM5: A crucial role in mitochondrial dynamics and programmed cell death

In response to mitochondrial damage, mitochondria activate mitochondrial dynamics to maintain normal functions, and an imbalance in mitochondrial dynamics triggers multiple programmed cell death processes. Recent studies have shown that phosphoglycerate mutase 5 (PGAM5) is associated with mitochondrial damage. PGAM5 activates mitochondrial biogenesis and mitophagy to promote a cellular compensatory response when mitochondria are mildly damaged, whereas severe damage to mitochondria leads to PGAM5 inducing excessive mitochondria fission, disruption to mitochondrial movement, and amplification of apoptosis, necroptosis and mitophagic death signals, which eventually evoke cell death. PGAM5 functions mainly through protein-protein interactions and specific Ser/Thr/His protein phosphatase activity. PGAM5 is also regulated by mitochondrial proteases. Detection of PGAM5 and its interacting protein partners should enable a more accurate evaluation of mitochondrial damage and a more precise method for the diagnosis and treatment of diseases.     

The mitochondrial elongation factors MIEF1 and MIEF2 exert partially distinct functions in mitochondrial dynamics

Mitochondria are dynamic organelles whose morphology is regulated by a complex balance of fission and fusion processes, and we still know relatively little about how mitochondrial dynamics is regulated. MIEF1 (also called MiD51) has recently been characterized as a key regulator of mitochondrial dynamics and in this report we explore the functions of its paralog MIEF2 (also called MiD49), to learn to what extent MIEF2 is functionally distinct from MIEF1. We show that MIEF1 and MIEF2 have many functions in common. Both are anchored in the mitochondrial outer membrane, recruit Drp1 from the cytoplasm to the mitochondrial surface and cause mitochondrial fusion, and MIEF2, like MIEF1, can interact with Drp1 and hFis1. MIEF1 and MIEF2, however, also differ in certain aspects. MIEF1 and MIEF2 are differentially expressed in human tissues during development. When overexpressed, MIEF2 exerts a stronger fusion-promoting effect than MIEF1, and in line with this, hFis1 and Mff can only partially revert the MIEF2-induced fusion phenotype, whereas MIEF1-induced fusion is reverted to a larger extent by hFis1 and Mff. MIEF2 forms high molecular weight oligomers, while MIEF1 is largely present as a dimer. Furthermore, MIEF1 and MIEF2 use distinct domains for oligomerization: in MIEF1, the region from amino acid residues 109–154 is required, whereas oligomerization of MIEF2 depends on amino acid residues 1 to 49, i.e. the N-terminal end. We also show that oligomerization of MIEF1 is not required for its mitochondrial localization and interaction with Drp1. In conclusion, our data suggest that the mitochondrial regulators MIEF1 and MIEF2 exert partially distinct functions in mitochondrial dynamics.     

Structural dynamics of the mitochondrial ADP/ATP carrier revealed by molecular dynamics simulation studies

The mitochondrial adenosine diphosphate/adenosine triphosphate (ADP/ATP) carrier has been recently crystallized in complex with its specific inhibitor carboxyatractyloside (CATR). In the crystal structure, the six-transmembrane helix bundle that defines the nucleotide translocation pathway is closed on the matrix side due to sharp kinks in the odd-numbered helices. The closed conformation is further sealed by the loops protruding into the matrix that interact through an intricate network of charge-pairs. To gain insight into its structural dynamics we performed molecular dynamics (MD) simulation studies of the ADP/ATP carrier with and without its cocrystallized inhibitor. The two trajectories sampled a conformational space around two different configurations characterized by distinct salt-bridge networks with a significant shift from inter- to intrarepeat bonding on the matrix side in the absence of CATR. Analysis of the geometrical parameters defining the transmembrane helices showed that even-numbered helices can undergo a face rotation, whereas odd-numbered helices can undergo a change in the wobble angle with a conserved proline acting as molecular hinge. Our results provide new information on the dynamical properties of the ADP/ATP carrier and for the first time yield a detailed picture of a stable carrier conformation in absence of the inhibitor.     

Mutant huntingtin, abnormal mitochondrial dynamics, defective axonal transport of mitochondria, and selective synaptic degeneration in Huntington's disease

Huntington's disease (HD) is a progressive, fatal neurodegenerative disease caused by expanded polyglutamine repeats in the HD gene. HD is characterized by chorea, seizures, involuntary movements, dystonia, cognitive decline, intellectual impairment and emotional disturbances. Research into mutant huntingtin (Htt) and mitochondria has found that mutant Htt interacts with the mitochondrial protein dynamin-related protein 1 (Drp1), enhances GTPase Drp1 enzymatic activity, and causes excessive mitochondrial fragmentation and abnormal distribution, leading to defective axonal transport of mitochondria and selective synaptic degeneration. This article summarizes latest developments in HD research and focuses on the role of abnormal mitochondrial dynamics and defective axonal transport in HD neurons. This article also discusses the therapeutic strategies that decrease mitochondrial fragmentation and neuronal damage in HD.     

The genetic control of plant mitochondrial morphology and dynamics

Little is known about the genetic control of mitochondrial morphology and dynamics in higher plants. We used a genetic screen involving fluorescence microscopic analysis of ethyl methane sulphonate (EMS)-mutated Arabidopsis thaliana seedlings expressing GFP targeted to mitochondria to isolate eight mutants displaying distinct perturbations of the normal mitochondrial morphology or distribution. We describe five mutants with distinct and unique mitochondrial phenotypes, which map to five different loci, not previously implicated in mitochondrial behaviour in plants. We have used a combination of forward and reverse genetics to identify one of the genes, friendly mitochondria (FMT), a homologue of the CluA gene of Dictyostelium discoideum, which is involved in the correct distribution of mitochondria in the cell. The five mutants constitute a powerful resource to aid our understanding of mitochondrial dynamics in higher plants.     

Regulation of autophagy,mitochondrial dynamics,and cellular bioenergetics by 4-hydroxynonenal in primary neurons

The production of reactive species contributes to the age-dependent accumulation of dysfunctional mitochondria and protein aggregates, all of which are associated with neurodegeneration. A putative mediator of these effects is the lipid peroxidation product 4-hydroxynonenal (4-HNE), which has been shown to inhibit mitochondrial function, and accumulate in the postmortem brains of patients with neurodegenerative diseases. This deterioration in mitochondrial quality could be due to direct effects on mitochondrial proteins, or through perturbation of the macroautophagy/autophagy pathway, which plays an essential role in removing damaged mitochondria. Here, we use a click chemistry-based approach to demonstrate that alkyne-4-HNE can adduct to specific mitochondrial and autophagy-related proteins. Furthermore, we found that at lower concentrations (5–10 μM), 4-HNE activates autophagy, whereas at higher concentrations (15 μM), autophagic flux is inhibited, correlating with the modification of key autophagy proteins at higher concentrations of alkyne-4-HNE. Increasing concentrations of 4-HNE also cause mitochondrial dysfunction by targeting complex V (the ATP synthase) in the electron transport chain, and induce significant changes in mitochondrial fission and fusion protein levels, which results in alterations to mitochondrial network length. Finally, inhibition of autophagy initiation using 3-methyladenine (3MA) also results in a significant decrease in mitochondrial function and network length. These data show that both the mitochondria and autophagy are critical targets of 4-HNE, and that the proteins targeted by 4-HNE may change based on its concentration, persistently driving cellular dysfunction.     

烟粉虱在不同寄主作物上的种群动态及化学防治

20 0 1年在河北省廊坊市研究了烟粉虱Bemisiatabaci(Gennadius)在主要寄主作物棉花、大豆、花生和玉米上的种群动态和化学防治方法。研究结果表明 ,烟粉虱的种群数量在寄主的生育期内持续增长 ,在 8月 2 2日左右达到高峰 ,之后逐渐减少。烟粉虱虽然寄主范围广泛 ,但在每种寄主上的发生量并不一致 ,调查结果表明烟粉虱在 4种不同寄主上的种群数量比较为 :棉花 >大豆 >花生 ,玉米上没有发现烟粉虱的卵及若虫 ,只有极少量的成虫在植株上逗留。在药效试验中 1 0 %吡虫啉WP、2 5 %扑虱灵WP和 1 0 %高效氯氰菊酯LC在厂家推荐浓度下对烟粉虱成虫的防治效果分别为 2 0 0 7%～ 3 3 1 9% ,2 0 62 %～ 44 88%和 4 0 5 %～ 6 78% ;对若虫的防治效果分别为 5 1 91 %～ 64 46% ,5 4 3 0 %～ 63 99%和 1 6 73 %～ 2 6 97%。     

Cyclin-dependent protein kinase 2 activity is required for mitochondrial translocation of Bax and disruption of mitochondrial transmembrane potential during etoposide-induced apoptosis

Previous studies have suggested that upregulation of Cyclin A-dependent protein kinase 2 (Cdk2) activity is an essential event in apoptotic progression and the mitochondrial permeability transition in human cancer cells. Here, we show that upregulated Cyclin A/Cdk2 activity precedes the proteolytic cleavage of PARP and is correlated with the mitochondrial translocation of Bax and the loss of mitochondrial transmembrane potential (Δψm) during etoposide-induced apoptosis in human cervical adenocarcinoma (HeLa) cells. Etoposide-induced apoptotic cell death is efficiently prevented in cells that overexpress a dominant negative mutant of Cdk2 (Cdk2-dn) or p21^WAF1/CIP1, a specific Cdk inhibitor. Conversely, apoptotic cell death is promoted in Cyclin A-expressing cells. Disruption of the mitochondrial transmembrane potential in etoposide-induced cells is prevented in cells that overexpress Cdk2-dn or p21^WAF1/CIP1, while this transition is prominently promoted in Cyclin A-expressing cells. We screened for mitochondrial Cdk2 targets in the etoposide-induced cells and found that the mitochondrial level of Bax is elevated by more than three fold in etoposide-treated cells and this elevation is effectively prevented in cells expressing Cdk2-dn under the same conditions. Thus, we suggest that Cdk2 activity is involved in the mitochondrial translocation of Bax, which plays an important role in the mitochondrial membrane permeability transition during apoptotic progression.     

Interaction against different environmental dynamics during a leg extension task is controlled by temporal rather than amplitude scaling of muscular activity

Force exertion against different mechanical environments can affect motor control strategies in order to account for the altered environmental dynamics and to maintain the ability to produce force. Here, we investigated the change of muscular activity of selected muscles of the lower extremities while the participants interacted with an external mechanical device of variable stability. Twenty-five healthy participants exerted force against the device by performing a unilateral ballistic leg extension task under 1 or 3 degrees of freedom (DoF). Directional force data and electromyographic responses from four leg muscles (TA, VM, GM, PL) were recorded. Muscle responses to the altered experimental conditions were analyzed by calculating time to peak electrical activity (TTP), peak electrical activity (PEA), slope of EMG-signal and muscle activity. It was found that neuromuscular system adjustments to the task are expressed mainly by temporal (TTP) rather than amplitude (PEA) scaling of muscular activity. This change was specific for the investigated muscles. Moreover, a selective increase of muscle activity occurred while increasing external DoF. This scheme was accompanied by a significant reduction of applicable force against the device in the unstable 3 DoF condition. The findings suggest that orchestration of movement control is linked to environmental dynamics also affecting the ability to produce force under dynamic conditions. The adjustments of the neuromuscular system are rather temporal in nature being consistent with the impulse timing hypothesis of motor control.     

The role of short-term synaptic dynamics in motor control

During the past few years, much attention has been given to the role of short-term synaptic plasticity, in particular depression and facilitation, in sculpting network activity. A recent study shows that synaptic depression in rhythmic motor networks could switch the control of network frequency from intrinsic neuronal properties to the synaptic dynamics. Short-term synaptic plasticity is also involved in the stabilization and reconfiguration of motor circuits and in the initiation, maintenance and modulation of motor programs.     

B-plexins control microtubule dynamics and dendrite morphology of hippocampal neurons

Semaphorins and their receptors plexins are implicated in various processes in the nervous system, but how B-plexins regulate the growth of dendrites remains poorly characterized. We had previously observed that Plexin-B1 and B3 interact with microtubule end-binding proteins (EBs) that are central adapters at growing microtubule tips, and this interaction is involved in neurite growth. Therefore, we hypothesized that plexins regulate microtubule dynamics and through that also dendritogenesis. The role of all three B-plexins was systematically examined in these processes. B-plexins and their ligand Semaphorin-4D influence the dynamics of microtubule tips both EB-dependently and independendently. EB3 as well as Plexin-B1, B2 and B3 turned out to have a significant role in the development of dendritic arbor of rat hippocampal neurons. Our results clearly indicate that semaphorin-plexin-EB pathway is one molecular mechanism how extracellular guidance cues are translated into intracellular mechanics. Taken together, Semaphorin-4D and B-plexins modulate the dynamic behavior of microtubule tips, and are therefore important in neurite growth.     

Reversible metabolic depression in lamprey hepatocytes during prespawning migration: dynamics of mitochondrial membrane potential

The lamprey (Lampetra fluviatilis L.) is an extant representative of the ancient vertebrate group of Agnathans. During the prespawning migration (the river period of life from autumn until spring) lamprey hepatocytes exhibit widely different energy states: a high-energy state in autumn and spring, corresponding to a normal physiological standard, and a low-energy state in winter, which is provoked by prolonged starvation and profound metabolic arrest. In spring the restoration of energy status (return to an active state) is associated with hormonally induced lipolysis of the lipid droplets stored in the cells. Lamprey hepatocytes demonstrate an aerobic metabolism based on oxidation of free fatty acids. The dynamics of mitochondrial membrane potential (MMP) were measured throughout the prespawning migration. Pharmacological inhibition of the electron transport chain decreased the MMP and caused extensive depletion of cellular ATP without loss of cell viability. The potential molecular mechanisms responsible for winter metabolic depression in lamprey hepatocytes are discussed.     

Genetic diversity of golden takin (Budorcas taxicolor bedfordi) population from Qinling Mountains in China revealed by sequence analysis of mitochondrial DNA control region

The golden takin is an endangered species, listed as a First Grade Protected animal and found only in the Qinling Mountains in China. A great deal of research on the golden takin's living habitat, population size, and home range has been conducted. Here, we employed sequence analysis of the mitochondrial DNA control region to study the genetic diversity of the golden takin from three separate nature reserve parks in the Qinling Mountains. We also compared the results of our study with previously published data on the genetic diversity of mixed takin species located in the Qinling Mountains and the Minshan area. Based on 62 sampled golden takin individuals, we found an overall mean genetic haplotype diversity of 0.687. There is no significant geographic genetic diversity across different golden takin populations within the Qinling Mountains. However, we did show significant diversity between golden takin from the Qinling Mountains and from Minshan. These original data provide a foundation for the genetic diversity of golden takin, and will yield comprehensive information for better supporting the management in the national reserve parks.     

Functional Characterization of TbMCP5, a Conserved and Essential ADP/ATP Carrier Present in the Mitochondrion of the Human Pathogen Trypanosoma brucei

Trypanosoma brucei is a kinetoplastid parasite of medical and veterinary importance. Its digenetic life cycle alternates between the bloodstream form in the mammalian host and the procyclic form (PCF) in the bloodsucking insect vector, the tsetse fly. PCF trypanosomes rely in the glucose-depleted environment of the insect vector primarily on the mitochondrial oxidative phosphorylation of proline for their cellular ATP provision. We previously identified two T. brucei mitochondrial carrier family proteins, TbMCP5 and TbMCP15, with significant sequence similarity to functionally characterized ADP/ATP carriers from other eukaryotes. Comprehensive sequence analysis confirmed that TbMCP5 contains canonical ADP/ATP carrier sequence features, whereas they are not conserved in TbMCP15. Heterologous expression in the ANC-deficient yeast strain JL1Δ2Δ3u⁻ revealed that only TbMCP5 was able to restore its growth on the non-fermentable carbon source lactate. Transport studies in yeast mitochondria showed that TbMCP5 has biochemical properties and ADP/ATP exchange kinetics similar to those of Anc2p, the prototypical ADP/ATP carrier of S. cerevisiae. Immunofluorescence microscopy and Western blot analysis confirmed that TbMCP5 is exclusively mitochondrial and is differentially expressed with 4.5-fold more TbMCP5 in the procyclic form of the parasite. Silencing of TbMCP5 expression in PCF T. brucei revealed that this ADP/ATP carrier is essential for parasite growth, particularly when depending on proline for energy generation. Moreover, ADP/ATP exchange in isolated T. brucei mitochondria was eliminated upon TbMCP5 depletion. These results confirmed that TbMCP5 functions as the main ADP/ATP carrier in the trypanosome mitochondrion. The important role of TbMCP5 in the T. brucei energy metabolism is further discussed.     

三化螟种群动态分析及防治

分析了 1 980～ 2 0 0 0年三化螟种群数量变动情况 ,发现年度间的变动不一致 ,有明显间歇性。认为冬春气候 (气温、雨量 )、耕作制度、天敌因素等与三化螟为害密切相关 ,并提出科学实用的防治方法     

FGFR1 is essential for N-acetyl-seryl-aspartyl-lysyl-proline regulation of mitochondrial dynamics by upregulating microRNA let-7b-5p

Fibroblast growth factor receptor (FGFR) 1 plays a key role in endothelial homeostasis by inducing microRNA (miR) let-7. Our previous paper showed that anti-fibrotic effects of N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP) were associated with restoring diabetes-suppressed expression of FGFR1 and miR let-7, the key contributor of mitochondrial biogenesis, which is regulated by mitochondrial membrane GTPase proteins (MFN2 and OPA1). Here, we found that the FGFR1 signaling pathway was critical for AcSDKP in maintaining endothelial mitochondrial biogenesis through induction of miR let-7b-5p. In endothelial cells, AcSDKP restored the triple cytokines (TGF-β2, interleukin-1β, tumor necrosis factor-α)-suppressed miR let-7b-5p and protein levels of the mitochondrial membrane GTPase. This effect of AcSDKP was lost with either fibroblast growth factor receptor substrate 2 (FRS2) siRNA or neutralizing FGFR1-treated cells. Similarly, AcSDKP had no effect on the miR let-7b-5p inhibitor-suppressed GTPase levels in endothelial cells. In addition, a miR let-7b-5p mimic restored the levels of FRS2 siRNA-reduced GTPases in endothelial cells. These findings were also confirmed using MitoTracker Green and an immunofluorescence assay. Our results demonstrated that the AcSDKP-FGFR1 signaling pathway is critical for maintaining mitochondrial dynamics by control of miR let-7b-5p in endothelial cells.     

The mitochondrial ADP/ATP carrier: functional and structural studies in the route of elucidating pathophysiological aspects

The mitochondrial ADP/ATP carrier plays a central role in aerobic cell energetics by providing to the cytosol the ATP generated by oxidative phosphorylation. Though discovered around 40 years ago owing to the existence of unique inhibitors and in spite of numerous experimental approaches, this carrier, which stands as a model of the mitochondrial solute carriers keeps some long-standing mystery. There are still open challenging questions among them the precise ADP/ATP transport mechanism, the functional oligomeric state of the carrier and relationships between human ADP/ATP carrier dysfunctioning and pathologies. Deciphering the 3D structure of this carrier afforded a considerable progress of the knowledge but requires now additional data focused on molecular dynamics from this static picture. State of the art in this topic is reviewed and debated in this paper in view of better comprehending origin of the discrepancies in these questions and, finally, the multiple physiological roles of this carrier in eukaryotic cell economy.