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1.
Background
The dependence of malignant properties of colorectal cancer (CRC) cells on IGF1R signaling has been demonstrated and several IGF1R antagonists are currently in clinical trials. Recently, we identified a novel pathway in which cAMP independent PKA activation by TGFβ signaling resulted in the destabilization of survivin/XIAP complex leading to increased cell death. In this study, we evaluated the effect of IGF1R inhibition or activation on PKA activation and its downstream cell survival signaling mechanisms.Methods
Small molecule IGF1R kinase inhibitor OSI-906 was used to test the effect of IGF1R inhibition on PKA activation, AKAP association and its downstream cell survival signaling. In a complementary approach, ligand mediated activation of IGF1R was performed and AKAP/PKA signaling was analyzed for their downstream survival effects.Results
We demonstrate that the inhibition of IGF1R in the IGF1R-dependent CRC subset generates cell death through a novel mechanism involving TGFβ stimulated cAMP independent PKA activity that leads to disruption of cell survival by survivin/XIAP mediated inhibition of caspase activity. Importantly, ligand mediated activation of the IGF1R in CRC cells results in the generation of cAMP dependent PKA activity that functions in cell survival by inhibiting caspase activity. Therefore, this subset of CRC demonstrates 2 opposing pathways organized by 2 different AKAPs in the cytoplasm that both utilize activation of PKA in a manner that leads to different outcomes with respect to life and death. The cAMP independent PKA activation pathway is dependent upon mitochondrial AKAP149 for its apoptotic functions. In contrast, Praja2 (Pja2), an AKAP-like E3 ligase protein was identified as a key element in controlling cAMP dependent PKA activity and pro-survival signaling. Genetic manipulation of AKAP149 and Praja2 using siRNA KD had opposing effects on PKA activity and survivin/XIAP regulation.Conclusions
We had identified 2 cytoplasmic pathways dependent upon the same enzymatic activity with opposite effects on cell fate in terms of life and death. Understanding the specific mechanistic functions of IGF1R with respect to determining the PKA survival functions would have potential for impact upon the development of new therapeutic strategies by exploiting the IGF1R/cAMP-PKA survival signaling in cancer.2.
Arlet Loza-Huerta Rosario Vera-Estrella Alberto Darszon Carmen Beltrán 《Biochimica et Biophysica Acta (BBA)/General Subjects》2013
Background
Sea urchin sperm motility is regulated by Speract, a sperm-activating peptide (SAP) secreted from the outer egg coat. Upon binding to its receptor in the sperm flagellum, Speract induces a series of ionic and metabolic changes in Strongylocentrotus purpuratus spermatozoa that regulate their motility. Among these events, protein phosphorylation is one of the most relevant and evidence indicates that some proteins of the Speract signaling cascade localize in low density detergent-insoluble membranes (LD-DIM).Methods
LD-DIM-derived proteins from immotile, motile or Speract-stimulated S. purpuratus sperm were resolved in 2-D gels and the PKA and PKC substrates detected with specific antibodies were identified by LC–MS/MS.Results
Differential PKA and PKC substrate phosphorylation levels among the LD-DIM isolated from sperm in different motility conditions were found and identified by mass spectrometry as: ATP synthase, creatine kinase, NADH dehydrogenase (ubiquinone) flavoprotein 2, succinyl-CoA ligase and the voltage-dependent anion channel 2 (VDAC2), which are mitochondrial proteins, as well as, the cAMP-dependent protein kinase type II regulatory (PKA RII) subunit, Tubulin β chain and Actin Cy I changed their phosphorylation state.Conclusions
Some mitochondrial proteins regulated by PKA or PKC may influence sea urchin sperm motility.General significance
The fact that a high percentage (66%) of the PKA or PKC substrates identified in LD-DIM are mitochondrial proteins suggests that the phosphorylation of these proteins modulates sea urchin sperm motility via Speract stimulation by providing sufficient energy to sperm physiology. Those mitochondrial proteins are indeed PKA- or PKC-substrates in the sea urchin spermatozoa. 相似文献3.
Victor M. Victor Susana Rovira-Llopis Vanessa Saiz-Alarcon Maria C. Sangüesa Luis Rojo-Bofill Celia Ba?uls Rosa Falcón Raquel Castelló Luis Rojo Milagros Rocha Antonio Hernández-Mijares 《PloS one》2014,9(9)
Context
Anorexia nervosa is a common illness among adolescents and is characterised by oxidative stress.Objective
The effects of anorexia on mitochondrial function and redox state in leukocytes from anorexic subjects were evaluated.Design and setting
A multi-centre, cross-sectional case-control study was performed.Patients
Our study population consisted of 20 anorexic patients and 20 age-matched controls, all of which were Caucasian women.Main outcome measures
Anthropometric and metabolic parameters were evaluated in the study population. To assess whether anorexia nervosa affects mitochondrial function and redox state in leukocytes of anorexic patients, we measured mitochondrial oxygen consumption, membrane potential, reactive oxygen species production, glutathione levels, mitochondrial mass, and complex I and III activity in polymorphonuclear cells.Results
Mitochondrial function was impaired in the leukocytes of the anorexic patients. This was evident in a decrease in mitochondrial O2 consumption (P<0.05), mitochondrial membrane potential (P<0.01) and GSH levels (P<0.05), and an increase in ROS production (P<0.05) with respect to control subjects. Furthermore, a reduction of mitochondrial mass was detected in leukocytes of the anorexic patients (P<0.05), while the activity of mitochondrial complex I (P<0.001), but not that of complex III, was found to be inhibited in the same population.Conclusions
Oxidative stress is produced in the leukocytes of anorexic patients and is closely related to mitochondrial dysfunction. Our results lead us to propose that the oxidative stress that occurs in anorexia takes place at mitochondrial complex I. Future research concerning mitochondrial dysfunction and oxidative stress should aim to determine the physiological mechanism involved in this effect and the physiological impact of anorexia. 相似文献4.
Gerrida M Uys Amsha Ramburan Benjamin Loos Craig J Kinnear Lundi J Korkie Jomien Mouton Johann Riedemann Johanna C Moolman-Smook 《BMC cell biology》2011,12(1):18
Background
Cardiac contractility is regulated by dynamic phosphorylation of sarcomeric proteins by kinases such as cAMP-activated protein kinase A (PKA). Efficient phosphorylation requires that PKA be anchored close to its targets by A-kinase anchoring proteins (AKAPs). Cardiac Myosin Binding Protein-C (cMyBPC) and cardiac troponin I (cTNI) are hypertrophic cardiomyopathy (HCM)-causing sarcomeric proteins which regulate contractility in response to PKA phosphorylation. 相似文献5.
Background
Loss of function mutations in the DJ-1 gene have been linked to recessively inherited forms of Parkinsonism. Mitochondrial dysfunction and increased oxidative stress are thought to be key events in the pathogenesis of Parkinson’s disease. Although it has been reported that DJ-1 serves as scavenger for reactive oxidative species (ROS) by oxidation on its cysteine residues, how loss of DJ-1 affects mitochondrial function is less clear.Methodology/Principal Findings
Using primary mouse embryonic fibroblasts (MEFs) or brains from DJ-1−/− mice, we found that loss of DJ-1 does not affect mitochondrial respiration. Specifically, endogenous respiratory activity as well as basal and maximal respiration are normal in intact DJ-1−/− MEFs, and substrate-specific state 3 and state 4 mitochondrial respiration are also unaffected in permeabilized DJ-1−/− MEFs and in isolated mitochondria from the cerebral cortex of DJ-1−/− mice at 3 months or 2 years of age. Expression levels and activities of all individual complexes composing the electron transport system are unchanged, but ATP production is reduced in DJ-1−/− MEFs. Mitochondrial transmembrane potential is decreased in the absence of DJ-1. Furthermore, mitochondrial permeability transition pore opening is increased, whereas mitochondrial calcium levels are unchanged in DJ-1−/− cells. Consistent with earlier reports, production of reactive oxygen species (ROS) is increased, though levels of antioxidative enzymes are unaltered. Interestingly, the decreased mitochondrial transmembrane potential and the increased mitochondrial permeability transition pore opening in DJ-1−/− MEFs can be restored by antioxidant treatment, whereas oxidative stress inducers have the opposite effects on mitochondrial transmembrane potential and mitochondrial permeability transition pore opening.Conclusions/Significance
Our study shows that loss of DJ-1 does not affect mitochondrial respiration or mitochondrial calcium levels but increases ROS production, leading to elevated mitochondrial permeability transition pore opening and reduced mitochondrial transmembrane potential. 相似文献6.
7.
David A Hughes Martin Jastroch Mark Stoneking Martin Klingenspor 《BMC evolutionary biology》2009,9(1):4
Background
Uncoupling protein 1 (UCP1) is a mitochondrial anion carrier, expressed in brown adipose tissue (BAT) of Eutherians. UCP1 is responsible for uncoupling mitochondrial proton transport from the production of ATP, thereby dissipating heat; it is essential for non-shivering thermogenesis (NST) in mammalian BAT. UCP1 orthologs have been identified in non-Eutherian mammals, fish and amphibians. Yet, UCP1 has a unique function in Eutherians in that it is necessary in the production of heat (NST). As such, this study aims to determine the evolutionary mode of UCP1 in Eutherians, where there is clear evidence of UCP1-dependent NST in BAT. 相似文献8.
Nadège Calmels Hervé Seznec Pascal Villa Laurence Reutenauer Marcel Hibert Jacques Haiech Pierre Rustin Michel Koenig Hélène Puccio 《BMC neurology》2009,9(1):46-10
Background
Pharmacological high-throughput screening (HTS) represents a powerful strategy for drug discovery in genetic diseases, particularly when the full spectrum of pathological dysfunctions remains unclear, such as in Friedreich ataxia (FRDA). FRDA, the most common recessive ataxia, results from a generalized deficiency of mitochondrial and cytosolic iron-sulfur cluster (ISC) proteins activity, due to a partial loss of frataxin function, a mitochondrial protein proposed to function as an iron-chaperone for ISC biosynthesis. In the absence of measurable catalytic function for frataxin, a cell-based assay is required for HTS assay. 相似文献9.
Background
Protein kinase A (cAMP-dependent kinase, PKA) is a serine/threonine kinase, for which ca. 150 substrate proteins are known. Based on a refinement of the recognition motif using the available experimental data, we wished to apply the simplified substrate protein binding model for accurate prediction of PKA phosphorylation sites, an approach that was previously successful for the prediction of lipid posttranslational modifications and of the PTS1 peroxisomal translocation signal. 相似文献10.
Eugene J Fine Anna Miller Edward V Quadros Jeffrey M Sequeira Richard D Feinman 《Cancer cell international》2009,9(1):14-11
Background
Recent evidence suggests that several human cancers are capable of uncoupling of mitochondrial ATP generation in the presence of intact tricarboxylic acid (TCA) enzymes. The goal of the current study was to test the hypothesis that ketone bodies can inhibit cell growth in aggressive cancers and that expression of uncoupling protein 2 is a contributing factor. The proposed mechanism involves inhibition of glycolytic ATP production via a Randle-like cycle while increased uncoupling renders cancers unable to produce compensatory ATP from respiration. 相似文献11.
Background
DNA replication requires contributions from various proteins, such as DNA helicases; in mitochondria Twinkle is important for maintaining and replicating mitochondrial DNA. Twinkle helicases are predicted to also possess primase activity, as has been shown in plants; however this activity appears to have been lost in metazoans. Given this, the study of Twinkle in other organisms is required to better understand the evolution of this family and the roles it performs within mitochondria.Results
Here we describe the characterization of a Twinkle homologue, Twm1, in the amoeba Dictyostelium discoideum, a model organism for mitochondrial genetics and disease. We show that Twm1 is important for mitochondrial function as it maintains mitochondrial DNA copy number in vivo. Twm1 is a helicase which unwinds DNA resembling open forks, although it can act upon substrates with a single 3′ overhang, albeit less efficiently. Furthermore, unlike human Twinkle, Twm1 has primase activity in vitro. Finally, using a novel in bacterio approach, we demonstrated that Twm1 promotes DNA replication.Conclusions
We conclude that Twm1 is a replicative mitochondrial DNA helicase which is capable of priming DNA for replication. Our results also suggest that non-metazoan Twinkle could function in the initiation of mitochondrial DNA replication. While further work is required, this study has illuminated several alternative processes of mitochondrial DNA maintenance which might also be performed by the Twinkle family of helicases.12.
Ulaganathan Mabalirajan Tanveer Ahmad Rakhshinda Rehman Geeta Devi Leishangthem Amit Kumar Dinda Anurag Agrawal Balaram Ghosh Surendra Kumar Sharma 《PloS one》2013,8(4)
Background
Baicalein, a bioflavone present in the dry roots of Scutellaria baicalensis Georgi, is known to reduce eotaxin production in human fibroblasts. However, there are no reports of its anti-asthma activity or its effect on airway injury.Methodology/Principal Findings
In a standard experimental asthma model, male Balb/c mice that were sensitized with ovalbumin (OVA), treated with baicalein (10 mg/kg, ip) or a vehicle control, either during (preventive use) or after OVA challenge (therapeutic use). In an alternate model, baicalein was administered to male Balb/c mice which were given either IL-4 or IL-13 intranasally. Features of asthma were determined by estimating airway hyperresponsiveness (AHR), histopathological changes and biochemical assays of key inflammatory molecules. Airway injury was determined with apoptotic assays, transmission electron microscopy and assessing key mitochondrial functions. Baicalein treatment reduced AHR and inflammation in both experimental models. TGF-β1, sub-epithelial fibrosis and goblet cell metaplasia, were also reduced. Furthermore, baicalein treatment significantly reduced 12/15-LOX activity, features of mitochondrial dysfunctions, and apoptosis of bronchial epithelia.Conclusion/Significance
Our findings demonstrate that baicalein can attenuate important features of asthma, possibly through the reduction of airway injury and restoration of mitochondrial function. 相似文献13.
Background
The unique metabolism of tumors was described many years ago by Otto Warburg, who identified tumor cells with increased glycolysis and decreased mitochondrial activity. However, “aerobic glycolysis” generates fewer ATP per glucose molecule than mitochondrial oxidative phosphorylation, so in terms of energy production, it is unclear how increasing a less efficient process provides tumors with a growth advantage.Methods/Findings
We carried out a screen for loss of genetic elements in pancreatic tumor cells that accelerated their growth as tumors, and identified mitochondrial ribosomal protein L28 (MRPL28). Knockdown of MRPL28 in these cells decreased mitochondrial activity, and increased glycolysis, but paradoxically, decreased cellular growth in vitro. Following Warburg''s observations, this mutation causes decreased mitochondrial function, compensatory increase in glycolysis and accelerated growth in vivo. Likewise, knockdown of either mitochondrial ribosomal protein L12 (MRPL12) or cytochrome oxidase had a similar effect. Conversely, expression of the mitochondrial uncoupling protein 1 (UCP1) increased oxygen consumption and decreased tumor growth. Finally, treatment of tumor bearing animals with dichloroacetate (DCA) increased pyruvate consumption in the mitochondria, increased total oxygen consumption, increased tumor hypoxia and slowed tumor growth.Conclusions
We interpret these findings to show that non-oncogenic genetic changes that alter mitochondrial metabolism can regulate tumor growth through modulation of the consumption of oxygen, which appears to be a rate limiting substrate for tumor proliferation. 相似文献14.
Amoêdo ND Rodrigues MF Pezzuto P Galina A da Costa RM de Almeida FC El-Bacha T Rumjanek FD 《PloS one》2011,6(7):e22264
Background
Tumor cells are characterized by accelerated growth usually accompanied by up-regulated pathways that ultimately increase the rate of ATP production. These cells can suffer metabolic reprogramming, resulting in distinct bioenergetic phenotypes, generally enhancing glycolysis channeled to lactate production. In the present work we showed metabolic reprogramming by means of inhibitors of histone deacetylase (HDACis), sodium butyrate and trichostatin. This treatment was able to shift energy metabolism by activating mitochondrial systems such as the respiratory chain and oxidative phosphorylation that were largely repressed in the untreated controls.Methodology/Principal Findings
Various cellular and biochemical parameters were evaluated in lung cancer H460 cells treated with the histone deacetylase inhibitors (HDACis), sodium butyrate (NaB) and trichostatin A (TSA). NaB and TSA reduced glycolytic flux, assayed by lactate release by H460 cells in a concentration dependent manner. NaB inhibited the expression of glucose transporter type 1 (GLUT 1), but substantially increased mitochondria bound hexokinase (HK) activity. NaB induced increase in HK activity was associated to isoform HK I and was accompanied by 1.5 fold increase in HK I mRNA expression and cognate protein biosynthesis. Lactate dehydrogenase (LDH) and pyruvate kinase (PYK) activities were unchanged by HDACis suggesting that the increase in the HK activity was not coupled to glycolytic flux. High resolution respirometry of H460 cells revealed NaB-dependent increased rates of oxygen consumption coupled to ATP synthesis. Metabolomic analysis showed that NaB altered the glycolytic metabolite profile of intact H460 cells. Concomitantly we detected an activation of the pentose phosphate pathway (PPP). The high O2 consumption in NaB-treated cells was shown to be unrelated to mitochondrial biogenesis since citrate synthase (CS) activity and the amount of mitochondrial DNA remained unchanged.Conclusion
NaB and TSA induced an increase in mitochondrial function and oxidative metabolism in H460 lung tumor cells concomitant with a less proliferative cellular phenotype. 相似文献15.
《Biochimica et Biophysica Acta (BBA)/General Subjects》2014,1840(12):3404-3412
BackgroundMitochondria are the major source of ATP to power sperm motility. Phosphorylation of mitochondrial proteins has been proposed as a major regulatory mechanism for mitochondrial bioenergetics.MethodsSperm motility was measured by a computer-assisted analyzer, protein detection by western blotting, membrane potential by tetramethylrhodamine, cellular ATP by luciferase assay and localization of PKA by immuno-electron microscopy.ResultsBicarbonate is essential for the creation of mitochondrial electro-chemical gradient, ATP synthesis and sperm motility. Bicarbonate stimulates PKA-dependent phosphorylation of two 60 kDa proteins identified as Tektin and glucose-6-phosphate isomerase. This phosphorylation was inhibited by respiration inhibition and phosphorylation could be restored by glucose in the presence of bicarbonate. However, this effect of glucose cannot be seen when the mitochondrial ATP/ADP exchanger was inhibited indicating that glycolytic-produced ATP is transported into the mitochondria and allows PKA-dependent protein phosphorylation inside the mitochondria.ConclusionsBicarbonate activates mitochondrial soluble adenylyl cyclase (sAC) which catalyzes cAMP production leading to the activation of mitochondrial PKA. Glucose can overcome the lack of ATP in the absence of bicarbonate but it cannot affect the mitochondrial sAC/PKA system, therefore the PKA-dependent phosphorylation of the 60 kDa proteins does not occur in the absence of bicarbonate.General significanceProduction of CO2 in Krebs cycle, which is converted to bicarbonate is essential for sAC/PKA activation leading to mitochondrial membrane potential creation and ATP synthesis. 相似文献
16.
17.
Background
While there is agreement that exercise is a powerful stimulus to increase both mitochondrial function and content, we do not know the optimal training stimulus to maximise improvements in mitochondrial biogenesis.Scope of review
This review will focus predominantly on the effects of exercise on mitochondrial function and content, as there is a greater volume of published research on these adaptations and stronger conclusions can be made.Major conclusions
The results of cross-sectional studies, as well as training studies involving rats and humans, suggest that training intensity may be an important determinant of improvements in mitochondrial function (as determined by mitochondrial respiration), but not mitochondrial content (as assessed by citrate synthase activity). In contrast, it appears that training volume, rather than training intensity, may be an important determinant of exercise-induced improvements in mitochondrial content. Exercise-induced mitochondrial adaptations are quickly reversed following a reduction or cessation of physical activity, highlighting that skeletal muscle is a remarkably plastic tissue. Due to the small number of studies, more research is required to verify the trends highlighted in this review, and further studies are required to investigate the effects of different types of training on the mitochondrial sub-populations and also mitochondrial adaptations in different fibre types. Further research is also required to better understand how genetic variants influence the large individual variability for exercise-induced changes in mitochondrial biogenesis.General significance
The importance of mitochondria for both athletic performance and health underlines the importance of better understanding the factors that regulate exercise-induced changes in mitochondrial biogenesis. This article is part of a Special Issue entitled Frontiers of Mitochondrial Research. 相似文献18.
Astrid C Schauss Huiyan Huang Seok-Yong Choi Liqun Xu Sébastien Soubeyrand Patricia Bilodeau Rodolfo Zunino Peter Rippstein Michael A Frohman Heidi M McBride 《BMC biology》2010,8(1):100
Background
Mitochondria are highly dynamic organelles whose morphology and position within the cell is tightly coupled to metabolic function. There is a limited list of essential proteins that regulate mitochondrial morphology and the mechanisms that govern mitochondrial dynamics are poorly understood. However, recent evidence indicates that the core machinery that governs mitochondrial dynamics is linked within complex intracellular signalling cascades, including apoptotic pathways, cell cycle transitions and nuclear factor kappa B activation. Given the emerging importance of mitochondrial plasticity in cell signalling pathways and metabolism, it is essential that we develop tools to quantitatively analyse the processes of fission and fusion. In terms of mitochondrial fusion, the field currently relies upon on semi-quantitative assays which, even under optimal conditions, are labour-intensive, low-throughput and require complex imaging techniques. 相似文献19.
David HT Yen Lih-Chi Chen Yuh-Chiang Shen Ying-Chen Chiu I-Chun Ho Ya-Jou Lou I-Chin Chen Jiin-Cherng Yen 《Journal of biomedical science》2011,18(1):32
Background
Adrenomedullin (ADM) exerts its biological functions through the receptor-mediated enzymatic mechanisms that involve protein kinase A (PKA), or neuronal nitric oxide synthase (nNOS). We previously demonstrated that the receptor-mediated cAMP/PKA pathway involves in ADM-enhanced baroreceptor reflex (BRR) response. It remains unclear whether ADM may enhance BRR response via activation of nNOS-dependent mechanism in the nucleus tractus solitarii (NTS). 相似文献20.