Regulation of Cellular Respiration

Int. J. Med. Sci. 2011, 8 http://www.medsci.org 88 IInntteerrnnaattiioonnaall JJoouurrnnaall ooff MMeeddiiccaall SScciieenncceess 2011; 8(2):88-96 © Ivyspring International Publisher. All rights reserved. Research Paper Parvovirus B19 Nonstructural Protein-Induced Damage of Cellular DNA and Resultant Apoptosis Brian D. Poole1,2, Violetta Kivovich1,3,4,5, Leona Gilbert4,5 and Stanley J. Naides1, 5,6  1. Huck Institute for Life Sciences, Pennsylvania State University College of Medicine/Milton S. Hershey Medical Center, Hershey, PA, USA 2. Current: Department of Microbiology and Molecular Biology, Brigham Young University, Provo, UT, USA 3. MD/PhD Program, Pennsylvania State University College of Medicine/Milton S. Hershey Medical Center, Hershey, PA, USA 4. Chemical Biology Division, Department of Biological and Environmental Science, University of Jyväskylä, Jyväskylä, Finland 5. Division of Rheumatology, Department of Medicine, Pennsylvania State University College of Medicine/Milton S. Hershey Medical Center, Hershey, PA, USA 6. Current: Quest Diagnostics Nichols Institute, San Juan Capistrano, CA, USA  Corresponding author: Stanley J. Naides, MD, Immunology, Quest Diagnostics Nichols Institute, 33608 Ortega Highway, San Juan Capistrano, CA 92690. Tel. 949 728-4578; fax 949 728-7852; email: stanley.j.naides@questdiagnostics.com Received: 2010.12.02; Accepted: 2011.01.13; Published: 2011.01.15 Abstract Parvovirus B19 is a widespread virus with diverse clinical presentations. The viral non-structural protein, NS1, binds to and cleaves the viral genome, and induces apoptosis when transfected into nonpermissive cells, such as hepatocytes. We hypothesized that the cyto-toxicity of NS1 in such cells results from chromosomal DNA damage caused by the DNA-nicking and DNA-attaching activities of NS1. Upon testing this hypothesis, we found that NS1 covalently binds to cellular DNA and is modified by PARP, an enzyme involved in repairing single-s t r a n d e d D N A n i c k s . W e f u r t h e r m o r e d i s c o v e r e d t h a t t h e D N A n i c k r e p a i r pathway initiated by poly(ADPribose)polymerase and the DNA repair pathways initiated by ATM/ATR are necessary for efficient apoptosis resulting from NS1 expression. Key words: Parvovirus B19, DNA damage and repair, fulminant liver failure, apoptosis, autoan -tibody, systemic lupus erythematosus Introduction Parvovirus B19 (B19) is a common virus with multiple clinical presentations. Infection in children is typically seen as erythema infectiosum, or fifth dis-ease (1), while adults often experience arthropathy lasting up to several months (2). Autoantibodies are often found subsequent to B19 infection, and are as-sociated with arthropathy (3-5). In patients with chronic hemolytic anemias, such as sickle cell disease or hereditary spherocytosis, the destruction of the e r y t h r o i d p r e c u r s o r p o o l b y B 1 9 l e a d s t o a p l a s t i c c r i s i s (6). B19 infection is implicated in hepatitis non-A-E acute fulminant liver failure (7-16). Although these are the best-described clinical illnesses caused by B19, the virus has been implicated in a wide spectrum of other illnesses (17). B19 infects a variety of cell types, but predomi-nantly replicates in erythroid precursors (18). Infec-tion of other cell types results in a limited, non-replicative state with overexpression of the viral nonstructural protein, NS1, and little expression of genes for the Regulation of Cellular Respiration Regulation of Cellular Respiration Bởi: OpenStaxCollege Cellular respiration must be regulated in order to provide balanced amounts of energy in the form of ATP The cell also must generate a number of intermediate compounds that are used in the anabolism and catabolism of macromolecules Without controls, metabolic reactions would quickly come to a stand still as the forward and backward reactions reached a state of equilibrium Resources would be used inappropriately A cell does not need the maximum amount of ATP that it can make all the time: At times, the cell needs to shunt some of the intermediates to pathways for amino acid, protein, glycogen, lipid, and nucleic acid production In short, the cell needs to control its metabolism Regulatory Mechanisms A variety of mechanisms is used to control cellular respiration Some type of control exists at each stage of glucose metabolism Access of glucose to the cell can be regulated using the GLUT proteins that transport glucose ([link]) Different forms of the GLUT protein control passage of glucose into the cells of specific tissues 1/6 Regulation of Cellular Respiration GLUT4 is a glucose transporter that is stored in vesicles A cascade of events that occurs upon insulin binding to a receptor in the plasma membrane causes GLUT4-containing vesicles to fuse with the plasma membrane so that glucose may be transported into the cell Some reactions are controlled by having two different enzymes—one each for the two directions of a reversible reaction Reactions that are catalyzed by only one enzyme can go to equilibrium, stalling the reaction In contrast, if two different enzymes (each specific for a given direction) are necessary for a reversible reaction, the opportunity to control the rate of the reaction increases, and equilibrium is not reached A number of enzymes involved in each of the pathways—in particular, the enzyme catalyzing the first committed reaction of the pathway—are controlled by attachment of a molecule to an allosteric site on the protein The molecules most commonly used in this capacity are the nucleotides ATP, ADP, AMP, NAD+, and NADH These regulators, allosteric effectors, may increase or decrease enzyme activity, depending on the prevailing conditions The allosteric effector alters the steric structure of the enzyme, usually affecting the configuration of the active site This alteration of the protein’s (the enzyme’s) structure either increases or decreases its affinity for its substrate, with the effect of increasing or decreasing the rate of the reaction The attachment signals to the enzyme This binding can increase or decrease the enzyme’s activity, providing feedback This feedback type of control is effective as long as the chemical affecting it is attached to the enzyme Once the overall concentration of the chemical decreases, it will diffuse away from the protein, and the control is relaxed Control of Catabolic Pathways Enzymes, proteins, electron carriers, and pumps that play roles in glycolysis, the citric acid cycle, and the electron transport chain tend to catalyze non-reversible reactions In other words, if the initial reaction takes place, the pathway is committed to proceeding with the remaining reactions Whether a particular enzyme activity is released depends upon the energy needs of the cell (as reflected by the levels of ATP, ADP, and AMP) Glycolysis The control of glycolysis begins with the first enzyme in the pathway, hexokinase ([link]) This enzyme catalyzes the phosphorylation of glucose, which helps to prepare the compound for cleavage in a later step The presence of the negatively charged phosphate in the molecule also prevents the sugar from leaving the cell When hexokinase is inhibited, glucose diffuses out of the cell and does not become a substrate for the respiration pathways in that tissue The product of the hexokinase reaction is glucose-6-phosphate, which accumulates when a later enzyme, phosphofructokinase, is inhibited 2/6 Regulation of Cellular Respiration The glycolysis pathway is primarily regulated at the three key enzymatic steps (1, 2, and 7) as indicated Note that the first two steps that are regulated occur early in the pathway and involve hydrolysis of ATP Phosphofructokinase is the main enzyme controlled in glycolysis High levels of ATP, citrate, or a lower, more acidic pH decrease the enzyme’s activity An increase in citrate concentration can occur because of a blockage in the citric acid cycle Fermentation, with its production of organic acids like lactic acid, frequently accounts for the increased acidity in a cell; however, the products of fermentation not typically accumulate in cells The last step in glycolysis is catalyzed by pyruvate kinase The pyruvate produced can proceed to be catabolized or converted into the amino acid alanine If no more energy is needed and alanine is in adequate supply, the enzyme is inhibited The enzyme’s activity is increased when ...Calcium-induced contraction of sarcomeres changes the regulation of mitochondrial respiration in permeabilized cardiac cells Tiia Anmann 1 , Margus Eimre 2 , Andrey V. Kuznetsov 3,4 , Tatiana Andrienko 3 , Tuuli Kaambre 1 , Peeter Sikk 1 , Evelin Seppet 2 , Toomas Tiivel 1,2 , Marko Vendelin 3,5 , Enn Seppet 1 and Valdur A. Saks 1,3 1 Laboratory of Bioenergetics, National Institute of Chemical Physics and Biophysics, Tallinn, Estonia 2 Department of Pathophysiology, University of Tartu, Estonia 3 Laboratory of Fundamental and Applied Bioenergetics, INSERM E0221, Joseph Fourier University, Grenoble, France 4 Department of General and Transplant Surgery, Innsbruck Medical University, Austria 5 Institute of Cybernetics, Tallinn, Estonia Calcium ions play a central role in the excitation- contraction coupling in muscle cells [1,2] and partici- pate in regulating the activities of multiple enzymes and metabolic systems, including mitochondrial Krebs cycle dehydrogenases, in many types of cells [2–6]. The presence of sophisticated Ca-transport systems in mito- chondria allows these organelles to control the calcium cycle in the cytoplasmic space [7–15] and the lifetime of the cell, as overload of mitochondria with calcium results in opening of the mitochondrial permeability transition pore, which eventually leads to cell death [11–15]. It has also been proposed that, owing to the simultaneous activation of the contractile system and mitochondrial enzymes by calcium, the ATP produc- tion is matched to its demand in cells (‘parallel activa- tion’ mechanism) [16–20]. However, both experimental and theoretical studies with detailed mathematical modelling of the calcium effects on the mitochondria showed that calcium can induce, by stimulation of the steps of Krebs cycle, only twofold changes in the rate Keywords adenine nucleotides; calcium; cardio- myocytes; intracellular energetic units, mitochondria Correspondence V. A. Saks, Laboratory of Bioenergetics, Joseph Fourier University, 2280, Rue de la Piscine, BP53X – 38041, Grenoble Cedex 9, France Fax: +33 4 76514218 Tel: +33 4 76635627 E-mail: Valdur.Saks@ujf-grenoble.fr (Received 15 March 2005, revised 21 April 2005, accepted 22 April 2005) doi:10.1111/j.1742-4658.2005.04734.x The relationships between cardiac cell structure and the regulation of mitochondrial respiration were studied by applying fluorescent confocal microscopy and analysing the kinetics of mitochondrial ADP-stimulated respiration, during calcium-induced contraction in permeabilized cardiomyo- cytes and myocardial fibers, and in their ‘ghost’ preparations (after selective myosin extraction). Up to 3 lm free calcium, in the presence of ATP, induced strong contraction of permeabilized cardiomyocytes with intact sarcomeres, accompanied by alterations in mitochondrial arrangement and a significant decrease in the apparent K m for exogenous ADP and ATP in the kinetics of mitochondrial respiration. The V max of respiration showed a moderate (50%) increase, with an optimum at 0.4 lm free calcium and a decrease at higher calcium concentrations. At high free-calcium concentra- tions, the direct flux of ADP from ATPases to mitochondria was dimi- nished compared to that at low calcium levels. All of these effects were unrelated either to mitochondrial calcium overload or to mitochondrial permeability transition and were not observed in ‘ghost’ preparations [...]... annotation of genes and of pathways, respectively The websites listed under Pathway Maps in Table 2.2 are good sources of diagrams of many cellular pathways The websites Biomodels and CellML are repositories of published mathematical models of a diverse range of cellular processes References & further readings Alberts, B., Johnson, A., Lewis, J., Raff, M., Roberts, K., and Walter, P (2002) Molecular biology of. .. Multiscale modelling of cancer 11.1 Attributes of cancer 11.2 A multiscale model of avascular tumor growth 11.2.1 Cellular scale 11.2.2 Extracellular scale 11.2.3 Subcellular scale 11.3 A multiscale model of colorectal cancer 11.3.1 Gene level: a Boolean network 11.3.2 Cell level: a discrete cell-cycle model 11.3.3 Tissue level: colonies of cells and oxygen supply 11.4 Continuum models of solid tumor growth... concepts and models can be couched in molecular and mechanistic terms Just as mathematics was employed by physics to describe physical phenomena, increasingly detailed understanding of the molecular machinery of the cell is allowing the development of mechanistic and kinetic models of cellular phenomena A model is meant to be a replica of the system Where details are absent – be it due to lack of instruments... molecules to a living cell One of the striking features of life on earth is the universality (as far as we know) of the chemistry of the basic building blocks of cells; this is especially true in the case of the carrier of genetic information, the DNA This universality suggests that it is in the intrinsic physicochemical properties of these biomolecules where one can find the origins of spatiotemporal organization... complexity of cellular networks Figure 2.7 is a broad summary of these networks as they relate to the ‘DNA-to-RNA-to-protein’ flow of information; the general network shown in the 14 From molecules to a living cell METABOLIC PATHWAYS Metabolism of Complex Carbohydrates Metabolism of Cofactors & Vitamins Nucleotide Metabolism Metabolism of Complex Lipids Carbohydrate Metabolism Metabolism of other Amino... Modularization according to biological function is another way of stating the hypothesis that – in the midst of these large, highly connected intracellular networks – only certain subnetworks are essential in driving particular cellular processes It is the modelling of these cellular processes in terms of these subnetworks that is the subject of this book The cellular processes discussed here – although primarily... trajectories, Newton’s theory of universal gravitation provides a sufficient description of the system What are the current physical or chemical theories upon which models of biological processes are based? As illustrated in many of the models in this book, theories of chemical kinetics are assumed to apply (these are summarized in Chapter 3) In general, RESEARC H Open Access Cellular stress-induced up-regulation of FMRP promotes cell survival by modulating PI3K-Akt phosphorylation cascades Se Jin Jeon 1 , Jung Eun Seo 1 , Sung-Il Yang 4 , Ji Woong Choi 5 , David Wells 2 , Chan Young Shin 3,4 , Kwang Ho Ko 1* Abstract Background: Fragile X syndrome (FXS), the most commonly inherited mental retardation and single gene cause of autistic spectrum disorder, occurs when the Fmr1 gene is mutated. The product of Fmr1, fragile X linked mental retardation protein (FMRP) is widely expressed in HeLa cells, however the roles of FMRP within HeLa cells were not elucidated, yet. Interacting with a diverse range of mRNAs related to cellular survival regulatory signals, understanding the functions of FMRP in cellular context would provide better insights into the role of this interesting protein in FXS. Using HeLa cells treated with etoposide as a model, we tried to determine whether FMRP could play a role in cell survival. Methods: Apoptotic cell death was induced by etoposide treatment on Hela cells. After we transiently modulated FMRP expression (silencing or enhancing) by using molecular biotech nological methods such as small hairpin RNA virus-induced knock down and overexpression using transfection with FMRP expression vectors, cellular viability was me asured using propidium iodide staining, TUNEL staining, and FACS analysis along with the level of activation of PI3K-Akt pathway by Western blot. Expression level of FMR P and apoptotic regulator BcL-xL was analyzed by Western blot, RT-PCR and immunocytochemistry. Results: An increased FMRP expression was measured in etoposide-treated HeLa cells, which was induced by PI3K- Akt activation. Without FMRP expression, cellular defence mechanism via PI3K-Akt-Bcl-xL was weakened and resulted in an augmented cell death by etoposide. In addition, FMRP over-expression lead to the activation of PI3K- Akt signalling pathway as well as increased FMRP and BcL-xL expression, which culminates with the increased cell survival in etoposide-treated HeLa cells. Conclusions: Taken together, these results suggest that FMRP expression is an essential part of cellular survival mechanisms through the modulation of PI3K, Akt, and Bcl-xL signal pathways. Background Fragile X syndrome (FXS) is a well known neurodeve- lopmental disorder caused by loss of fragile X linked mental retar dation protein (FMRP) which is encoded by Fmr1 gene [1]. FXS pat ients typically show a wide spec- trum of cognitive and behavioral problems such as attention deficit, anxiety and mood disorder, increased risk of seizures, autist ic spectrum behaviors, and mental retardation [1]. F MRP is expressed in many tissues including testis, placenta, and brain [2,3] and in a variety of cell types including HeLa [4]. FMRP is a RNA binding protein, which r egulates translation of target mRNAs. A wide range of potential target mRNAs have been suggested, most of which have been correlated to the regulation of synaptic function as well as neuronal development (for a review, see [5,6]). Interestingly, many mRNAs encoding a diverse array of proteins having no known link to neuronal development and synaptogenesis were also suggested i ncluding phos- phoino sitide 3 kinase (PI3K) [7], amyloid precursor pro- tein (APP ) [8], and Bcl-2 interacting protein (Bnip) [9]. In addition, FMRP is found both in the nucleus and cytoplasm and shuttles between the two compartment * Correspondence: khk123@snu.ac.kr 1 Department of Pharmacology, College of Pharmacy and Research Institute of Pharmaceutical Sciences, Seoul National University, Seoul, Korea Full list of author information is available at the end of the article Jeon et al. Journal of Biomedical Science 2011, 18:17 http://www.jbiomedsci.com/content/18/1/17 © 2011 Jeon et al; licensee BioMed Central Ltd. T his is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons .org/licenses/by/2.0), which pe RESEARC H Open Access Intermolecular masking of the HIV-1 Rev NLS by the cellular protein HIC: Novel insights into the regulation of Rev nuclear import Lili Gu 1,2 , Takahiro Tsuji 1,3 , Mohamed Ali Jarboui 1 , Geok P Yeo 1 , Noreen Sheehy 1 , William W Hall 1 , Virginie W Gautier 1* Abstract Background: The HIV-1 regulatory protein Rev, which is essential for viral replication, mediates the nuclear export of unspliced viral transcripts. Rev nuclear function requires active nucleocytoplasmic shuttling, and Rev nuclear import is me diated by the recognition of its Nuclear Localisation Signal (NLS) by multiple import factors, which include transportin and importin b. However, it remains unclear which nuclear import pathway(s) predominate in vivo, and the cellular environment that modulates Rev nucleocytoplasmic shuttling remains to be characterised. Results: In our study, we have identified the cellular protein HIC (Human I-mfa domain-Containing protein) as a novel interactor of HIV-1 Rev. We demonstrate that HIC selectively interferes with Rev NLS interaction with importin b and impedes its nuclear import and function, but does not affect Rev nuclear import mediated by transportin. Hence, the molecular determinants mediating Rev-NLS recognition by importin b and transportin appear to be distinct. Furthermore, we have employed HIC and M9 M, a peptide specifically designed to inhibit the transportin- mediated nuclear import pathway, to characterise Rev nuclear import pathways within different cellular environments. Remarkably, we could show that in 293T, HeLa, COS7, Jurkat, U937, THP-1 and CEM cells, Rev nuc lear import is cell type specific and alternatively mediated by transportin or importin b, in a mutually exclusive fashion. Conclusions: Rev cytoplasmic sequestration by HIC may represent a novel mechanism for the control of Rev function. These studies highlight that the multivalent nature of the Rev NLS for different import receptors enables Rev to adapt its nuclear trafficking strategy. Background The HIV-1 regulatory protein Rev (18 kDa) is essential for HIV-1 replication [1,2]. Rev is predominantly loca- lised in the nucleus/nucleolus [3], and its primary func- tion is to mediate the nuclear export of partially spliced and unspliced viral transcripts. Rev has also been shown to modulate splicing and translation of viral transcripts, and their subsequent packaging, and to interfere with integration of the HIV-1 genome [4-7]. Rev nuclear export of unspliced viral transcripts requires active shut- tling of the protei n between the nucleus and c ytoplasm via nuclear pore complexes (NPCs) which is mediated by two major functional domains, the Nuclear Localisation Signal (NLS) and the Nuclear Export Signal (NES) [8,9]. The leucine-rich Rev NES binds directly to CRM1, which in concert with DDX3, a DEAD box RNA helicase, facili- tates Rev nu clear ex port of unspliced viral transcripts via the NPC [10-14]. Also , Rev-export function was shown to be inhibited by Nuclear Factor 90 (NF90)[15]. The basic argi nine-rich Rev NLS me diates both Rev nuclear import and binding to the Rev Response Element (RRE), a cis-acting RNA e lement present in all unspliced viral transcripts [16-18]. The Rev NLS is recognized by at least 5 different importin b family members, including impor- tin b, transp ortin, importin 5, importin 7 and importin 9, which facilitate its nuclear import [19-23]. Despite evi- dence showing the utilisation of multiple nuclear import receptors in vitro by Rev, it remains unclear if some are redundant and/or if, under specific conditions, one nuclear import pathway may predominate in vivo. Hutten * Correspondence: virginie.gautier@ucd.ie 1 UCD-Centre for Research in Infectious Diseases, School of Medicine and Medical Science, University College Dublin (UCD), Belfield, Dublin 4, Ireland Full list of author information is available at the end of the article Gu et al. Retrovirology 2011, 8:17 http://www.retrovirology.com/con ... Elevated levels of effector Effect on pathway activity glycolysis hexokinase glucose-6-phosphate decrease 4/6 Regulation of Cellular Respiration Summary of Feedback Controls in Cellular Respiration. .. control of the respiration processes is accomplished through the control of specific enzymes in the pathways This is a type of negative feedback, turning the 5/6 Regulation of Cellular Respiration. .. see an animation of the electron transport chain and ATP synthesis For a summary of feedback controls in cellular respiration, see [link] Summary of Feedback Controls in Cellular Respiration Pathway