bluuebunny/crossref_metadata_2025_split huggingface dataset:
Isozymes of mammalian hexokinase: structure, subcellular localization and metabolic function
Authors: Wilson John E. 
Published: May 2003

SUMMARY
The first step in metabolism of glucose (Glc) is usually phosphorylation,catalyzed by hexokinase. However, the Glc-6-P produced can then enter one or more of several alternative pathways. Selective expression of isozymic forms of hexokinase, differing in catalytic and regulatory properties as well as subcellular localization, is likely to be an important factor in determining the pattern of Glc metabolism in mammalian tissues/cells. Despite their overall structural similarity, the Type I, Type II and Type III isozymes differ in important respects. All three isozymes are inhibited by the product,Glc-6-P, but with the Type I isozyme, this inhibition is antagonized by PI, whereas with the Type II and Type III isozymes, Piactually causes additional inhibition. Reciprocal changes in intracellular levels of Glc-6-P and Pi are closely associated with cellular energy status, and it is proposed that the response of the Type I isozyme to these effectors adapts it for catabolic function, introducing Glc into glycolytic metabolism for energy production. In contrast, the Type II, and probably the Type III, isozymes are suggested to serve primarily anabolic functions, e.g. to provide Glc-6-P for glycogen synthesis or metabolism via the pentose phosphate pathway for lipid synthesis. Type I hexokinase binds to mitochondria through interaction with porin, the protein that forms channels through which metabolites traverse the outer mitochondrial membrane. Several experimental approaches have led to the conclusion that the Type I isozyme, bound to actively phosphorylating mitochondria, selectively uses intramitochondrial ATP as substrate. Such interactions are thought to facilitate coordination of the introduction of Glc into glycolysis, via the hexokinase reaction, with the terminal oxidative stages of Glc metabolism occurring in the mitochondria, thus ensuring an overall rate of Glc metabolism commensurate with cellular energy demands and avoiding excessive production of lactate. The Type II isozyme also binds to mitochondria. Whether such coupling occurs with mitochondrially bound Type II hexokinase in normal tissues, and how it might be related to the proposed anabolic role of this isozyme, remain to be determined. The Type III isozyme lacks the hydrophobic N-terminal sequence known to be critical for binding of the Type I and Type II isozymes to mitochondria. Immunolocalization studies have indicated that, in many cell types, the Type III has a perinuclear localization, the possible metabolic consequences of which remain unclear.
Metabolic consequences of functional complexes of mitochondria,myofibrils and sarcoplasmic reticulum in muscle cells
Authors: Andrienko T. , Kuznetsov A. V. , Kaambre T. , Usson Y. , Orosco A. , Appaix F. , Tiivel T. , Sikk P. , Vendelin M. , Margreiter R. , Saks V. A. 
Published: May 2003

SUMMARY
Regulation of mitochondrial respiration both by endogenous and exogenous ADP in the cells in situ was studied in isolated and permeabilized cardiomyocytes, permeabilized cardiac fibers and `ghost' fibers (all with a diameter of 10–20 μm) at different (0–3 μmoll-1)free Ca2+ concentrations in the medium. In all these preparations,the apparent Km of mitochondrial respiration for exogenous ADP at free Ca2+ concentrations of 0–0.1μmoll-1 was very high, in the range of 250–350μmoll-1, in contrast to isolated mitochondria in vitro(apparent Km for ADP is approximately 20μmoll-1). An increase in the free Ca2+ concentration(up to 3 μmoll-1, which is within physiological range), resulted in a very significant decrease of the apparent Km value to 20–30 μmoll-1, a decrease of Vmax of respiration in permeabilized intact fibers and a strong contraction of sarcomeres. In ghost cardiac fibers, from which myosin was extracted but mitochondria were intact, neither the high apparent Km for ADP (300–350 μmoll-1) nor Vmax of respiration changed in the range of free Ca2+ concentration studied, and no sarcomere contraction was observed. The exogenous-ADP-trapping system (pyruvate kinase + phosphoenolpyruvate) inhibited endogenous-ADP-supported respiration in permeabilized cells by no more than 40%, and this inhibition was reversed by creatine due to activation of mitochondrial creatine kinase. These results are taken to show strong structural associations (functional complexes) among mitochondria, sarcomeres and sarcoplasmic reticulum. Inside these complexes, mitochondrial functional state is controlled by channeling of ADP, mostly via energy- and phosphoryl-transfer networks, and apparently depends on the state of sarcomere structures.
Myoglobin function reassessed
Authors: Wittenberg Jonathan B. , Wittenberg Beatrice A. 
Published: May 2003

SUMMARYThe heart and those striated muscles that contract for long periods, having available almost limitless oxygen, operate in sustained steady states of low sarcoplasmic oxygen pressure that resist change in response to changing muscle work or oxygen supply. Most of the oxygen pressure drop from the erythrocyte to the mitochondrion occurs across the capillary wall. Within the sarcoplasm,myoglobin, a mobile carrier of oxygen, is developed in response to mitochondrial demand and augments the flow of oxygen to the mitochondria. Myoglobin-facilitated oxygen diffusion, perhaps by virtue of reduction of dimensionality of diffusion from three dimensions towards two dimensions in the narrow spaces available between mitochondria, is rapid relative to other parameters of cell respiration. Consequently, intracellular gradients of oxygen pressure are shallow, and sarcoplasmic oxygen pressure is nearly the same everywhere. Sarcoplasmic oxygen pressure, buffered near 0.33 kPa (2.5 torr; equivalent to approximately 4 μmol l-1 oxygen) by equilibrium with myoglobin, falls close to the operational Km of cytochrome oxidase for oxygen, and any small increment in sarcoplasmic oxygen pressure will be countered by increased oxygen utilization. The concentration of nitric oxide within the myocyte results from a balance of endogenous synthesis and removal by oxymyoglobin-catalyzed dioxygenation to the innocuous nitrate. Oxymyoglobin,by controlling sarcoplasmic nitric oxide concentration, helps assure the steady state in which inflow of oxygen into the myocyte equals the rate of oxygen consumption.
Mitochondria are morphologically heterogeneous within cells
Authors: Collins Tony J. , Bootman Martin D. 
Published: May 2003

SUMMARY
Mitochondria play key roles in the life and death of cells. We investigated whether mitochondria represent morphologically continuous entities within single intact cells. Physical continuity of mitochondria was determined by three-dimensional reconstruction of fluorescence from mitochondrially targeted DsRed1 or tetra-methyl rhodamine ethyl ester (TMRE). The mitochondria of pancreatic acinar, porcine aortic endothelial (PAE) cells, COS-7 cells and SH-SY5Y cells and neocortical astrocytes all displayed heterogeneous distributions and were of varying sizes. In general, there was a denser aggregation of mitochondria in perinuclear positions than in the cell periphery, where individual isolated mitochondria could clearly be seen. DsRed1 was found to be highly mobile within the matrix of individual mitochondria, with an estimated linear diffusion rate of 1μm s-1. High-intensity irradiation of subcellular regions bleached the fluorescence of mitochondrially targeted DsRed1, but did not cause the mitochondria to depolarise or fragment. A lack of rapid fluorescence-recovery-after-photobleaching (FRAP) of DsRed1 indicated lumenal discontinuity between mitochondria. We observed a slow (half-time approx. 20 min) recovery of DsRed1 fluorescence within the irradiated area that was attributed to mitochondrial movement or fusion of unbleached and bleached organelles. Mitochondria were not electrically coupled, since typically only individual mitochondria were observed to depolarise following irradiation of TMRE-loaded cells. Our data indicate that the mitochondria within individual cells are morphologically heterogeneous and unconnected, thus allowing them to have distinct functional properties.
Mitochondrial movement and positioning in axons: the role of growth factor signaling
Authors: Chada Sonita R. , Hollenbeck Peter J. 
Published: May 2003

SUMMARYThe extreme length of axonal processes requires that aerobic ATP production and Ca2+ homeostasis are non-uniformly organized in the cytoplasm. As a result, the transport and positioning of mitochondria along axons is essential for neuronal homeostasis. Mitochondria undergo rapid but intermittent transport in both the anterograde and retrograde directions in axons. We have shown that in chick embryonic sensory neurons, the transport of mitochondria responds to physiological changes in the cell and, particularly,to growth cone activity. When an axon is actively elongating, mitochondria move preferentially anterograde and then become stationary, accumulating in the region of the active growth cone. When axonal elongation ceases,mitochondria in the distal axon resume movement but undergo net retrograde transport and become uniformly distributed along the axon. This redistribution of mitochondria is achieved in two ways: there is a transition between motile and stationary mitochondria and a large up- and downregulation of their anterograde, but not retrograde, motor activity. Mitochondrial transport does not respond to the experimentally induced elongation of axons in the absence of an active growth cone, implying that signals from the active growth cone regulate transport. To determine the nature of these signals, we have focally stimulated the shafts of sensory axons in culture with nerve growth factor(NGF) covalently conjugated to polystyrene beads. We find that mitochondria accumulate at regions of focal NGF stimulation. This response is specific to mitochondria and does not result from general disruption of the cytoskeleton in the region of stimulation. Disruption of the phosphoinositide 3-kinase (PI 3-kinase) pathway, one of the signaling pathways downstream from NGF–receptor binding, completely eliminates NGF effects on mitochondrial behavior in axons. We propose that mitochondrial transport and/or docking are regulated in part via NGF/TrkA/PI 3-kinase signaling.
Four-dimensional organization of protein kinase signaling cascades: the roles of diffusion, endocytosis and molecular motors
Authors: Kholodenko Boris N 
Published: May 2003

SUMMARYExtracellular signals received by membrane receptors are processed, encoded and transferred to the nucleus via phosphorylation and spatial relocation of protein members of multiple component pathways, such as mitogen activated protein kinase (MAPK) cascades. The receptor-induced membrane recruitment of the cytoplasmic protein SOS results in the activation of the Ras/MAPK cascade. It has been suggested that the membrane recruitment of signaling proteins causes an increase in the diffusion-limited rates. We have recently shown that this increase is too small to be responsible for enhanced signal transduction. Instead we demonstrate that the function of membrane localization is to increase the number (or average lifetime) of complexes between signaling partners. A hallmark of signaling pathways is the spatial separation of activation and deactivation mechanisms; e.g. a protein can be phosphorylated at the cell surface by a membrane-bound kinase and dephosphorylated in the cytosol by a cytosolic phosphatase. Given the measured values of protein diffusion coefficients and of phosphatase and kinase activities, the spatial separation is shown to result in precipitous phospho-protein gradients. When information transfer is hampered by slow protein diffusion and rapid dephosphorylation, phospho-protein trafficking within endocytic vesicles may be an efficient way to deliver messages to physiologically relevant locations. The proposed mechanism explains recent observations that various inhibitors of endocytosis can inhibit MAPK activation. Additional mechanisms facilitating the relay of signals from cell-surface receptors to the nucleus can involve the assembly of protein kinases on a scaffolding protein and active transport of signaling complexes by molecular motors. We also discuss long-range signaling within a cell, such as survival signaling in neurons. We hypothesize that ligand-independent waves of receptor activation or/and traveling waves of phosphorylated kinases emerge to spread the signals over long distances.
The caveolar nitric oxide synthase/arginine regeneration system for NO production in endothelial cells
Authors: Solomonson Larry P. , Flam Brenda R. , Pendleton Laura C. , Goodwin Bonnie L. , Eichler Duane C. 
Published: May 2003

SUMMARY
The enzyme endothelial nitric oxide synthase (eNOS) catalyzes the conversion of arginine, oxygen and NADPH to NO and citrulline. Previous results suggest an efficient, compartmentalized system for recycling of citrulline to arginine utilized for NO production. In support of this hypothesis, the recycling enzymes, argininosuccinate synthase (AS) and argininosuccinate lyase (AL), have been shown to colocalize with eNOS in caveolae, a subcompartment of the plasma membrane. Under unstimulated conditions, the degree of recycling is minimal. Upon stimulation of NO production by bradykinin, however, recycling is co-stimulated to the extent that more than 80% of the citrulline produced is recycled to arginine. These results suggest an efficient caveolar recycling complex that supports the receptor-mediated stimulation of endothelial NO production. To investigate the molecular basis for the unique location and function of endothelial AS and AL,endothelial AS mRNA was compared with liver AS mRNA. No differences were found in the coding region of the mRNA species, but significant differences were found in the 5′-untranslated region (5′-UTR). The results of these studies suggest that sequence in the endothelial AS-encoding gene, represented by position -92 nt to -43 nt from the translation start site in the extended AS mRNA 5′-UTRs, plays an important role in differential and tissue-specific expression. Overall, a strong evidential case has been developed supporting the proposal that arginine availability, governed by a caveolar-localized arginine regeneration system, plays a key role in receptor-mediated endothelial NO production.
Shaken and stirred: muscle structure and metabolism
Authors: Suarez Raul K. 
Published: May 2003

SUMMARYMuscles are ideal models with which to examine the relationship between structure and metabolism because they are some of the most highly structured cells, and are capable of the largest and most rapid metabolic transitions as well as the highest metabolic rates known. Studies of metabolism have traditionally been conducted within what can considered as the kinetic paradigm provided by `solution biochemistry'; i.e. the rates of enzymatic reactions are studied in terms of their regulation by mass-action and allosteric effectors and, most recently, metabolic control analysis of pathways. This approach has served biology well and continues to be useful. Here, we consider the diffusion of small and large molecules in muscles and energy metabolism in the context of intracellular space. We find that in attempting to explain certain phenomena, a purely kinetic paradigm appears insufficient. Instead, phenomena such as the `shuttling' of high-energy phosphate donors and acceptors and the binding of metabolic enzymes to intracellular structures or to each other are better understood when metabolic rates and their regulation are considered in the context of intracellular compartments, distances, gradients and diffusion. As in all of biology,however, complexity dominates, and to such a degree that one pathway may consist of several reactions that each behave according to different rules.`Soluble' creatine kinase operates at or near equilibrium, while mitochondrial and myofibrillar creatine kinases directly channel substrate to (or from) the adenine nucleotide translocase and actomyosin-ATPase, their operation being thus displaced from equilibrium. Hexose 6-phosphate metabolism appears to obey the rules of solution biochemistry, e.g. phosphoglucoisomerase behaves as Haldane would have predicted in 1930. In contrast, given low steady-state substrate and product concentrations and high flux rates, a number of glycolytic reactions further downstream must be catalyzed by enzymes localized in close proximity to each other. Metabolites may be channeled within these complexes. When observed, mechanistic differences between species in the same steps or processes should not be surprising, considering how animals vary so much in structures, mechanical properties, mitochondrial contents and metabolic rates. This analysis suggests that declarations of the triumph of one mechanism or paradigm over all others, as well as calls for the abandonment of solution biochemistry, are unwarranted. Rather, metabolic biochemistry would seem better served by reconciling the old and the new.
Analysis of glycolytic enzyme co-localization in Drosophilaflight muscle
Authors: Sullivan David T. , MacIntyre R. , Fuda N. , Fiori J. , Barrilla J. , Ramizel L. 
Published: May 2003

SUMMARY
In Drosophila flight muscles, glycolytic enzymes are co-localized along sarcomeres at M-lines and Z-discs and co-localization is required for normal flight. We have extended our analysis of this phenomenon to include a set of six glycolytic enzymes that catalyze consecutive reactions along the glycolytic pathway: aldolase, glycerol-3-phosphate dehydrogenase (GPDH),glyceraldehyde-3-phosphate dehydrogenase (GAPDH), triose phosphate isomerase,phosphoglycerate kinase and phosphoglycerol mutase (PGLYM). Each of these enzymes has an identical pattern of localization. In mutants null for GPDH,localization of none of the other enzymes occurs and therefore is interdependent. In optimally fixed preparations of myofibrils, accumulation of the enzymes at M-lines is much greater than at Z-discs. However, localization at M-lines is more labile, as shown by loss of localization when fixation is delayed. We have begun to analyze the protein—protein interaction involved in glycolytic enzyme co-localization using the yeast two-hybrid system. We have identified two pair-wise interactions. One is between GPDH and GAPDH and another is between GPDH and PGLYM.
Body size as a latent variable in a structural equation model: thermal acclimation and energetics of the leaf-eared mouse
Authors: Nespolo Roberto F. , Arim Matías , Bozinovic Francisco 
Published: May 2003

SUMMARYBody size is one of the most important determinants of energy metabolism in mammals. However, the usual physiological variables measured to characterize energy metabolism and heat dissipation in endotherms are strongly affected by thermal acclimation, and are also correlated among themselves. In addition to choosing the appropriate measurement of body size, these problems create additional complications when analyzing the relationships among physiological variables such as basal metabolism, non-shivering thermogenesis,thermoregulatory maximum metabolic rate and minimum thermal conductance, body size dependence, and the effect of thermal acclimation on them.We measured these variables in Phyllotis darwini, a murid rodent from central Chile, under conditions of warm and cold acclimation. In addition to standard statistical analyses to determine the effect of thermal acclimation on each variable and the body-mass-controlled correlation among them, we performed a Structural Equation Modeling analysis to evaluate the effects of three different measurements of body size (body mass, mb; body length, Lb and foot length, Lf) on energy metabolism and thermal conductance. We found that thermal acclimation changed the correlation among physiological variables. Only cold-acclimated animals supported our a priori path models, and mb appeared to be the best descriptor of body size (compared with Lb and Lf) when dealing with energy metabolism and thermal conductance. However, while mb appeared to be the strongest determinant of energy metabolism, there was an important and significant contribution of Lb (but not Lf) to thermal conductance. This study demonstrates how additional information can be drawn from physiological ecology and general organismal studies by applying Structural Equation Modeling when multiple variables are measured in the same individuals.
Occlusable corneas in toadfishes: light transmission, movement and ultrastruture of pigment during light- and dark-adaptation
Authors: Siebeck Ulrike E. , Collin Shaun P. , Ghoddusi Majid , Marshall N. Justin 
Published: May 2003

SUMMARYThe toadfishes Tetractenos hamiltoni and Torquigener pleurogramma (Tetraodontidae) possess occlusable yellow corneas. We examine the light transmission and location of the yellow/orange pigment throughout the cornea, the temporal properties of pigment migration and the ultrastructure of the pigmented processes during light- and dark-adaptation. Each species was dark-adapted during the day and light-adapted during the night and then exposed to either sun illumination or darkness for different lengths of time (0–70 min). Movement of corneal pigment could be induced in both species regardless of time of day or night. The pigment was able to migrate in a dorsal or ventral direction and changed from minimal to maximal pigmentation within 60 min. Three types of transmission curves were found with varying degrees of transmission in the 400–500 nm waveband, indicating that the pigment distribution is not uniform across the cornea; some areas of the cornea transmit near UV light, while others absorb blue light. The gradual change of the transmission characteristics in different areas of the cornea indicates the presence of different concentrations of a single type of pigment. Ultrastructural examination of the corneas showed that the layer containing the pigment is situated within the scleral cornea either surrounding (T. pleurogramma) or abutting (T. hamiltoni) an iridescent layer. Long sheet-like processes or chromatophores extending centrally from dorsal and ventral reservoirs are filled with pigment during the light-adapted state but empty in the dark-adapted state.