ACTIN Anchors the Highly Oligomeric DRP1 at Mitochondria-Sarcoplasmic Reticulum Contact Sites in Adult Murine Heart: Its Functional Implication

2021 ◽  
Author(s):  
Celia Fernandez Sanz ◽  
Sergio De la Fuente Perez ◽  
Zuzana Nichtova ◽  
Yuexing Yuan ◽  
Sebastian Lanvermann ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Sarcoplasmic Reticulum ◽  
High Molecular Weight ◽  
Critical Role ◽  
Cardiac Contraction ◽  
Cardiac Physiology ◽  
Functional Implication ◽  
Contact Sites ◽  
Ec Coupling ◽  
Adult Cardiomyocytes

Rationale: Mitochondrial fission and fusion are relatively infrequent in adult cardiomyocytes compared to other cell types. This is surprising considering that proteins involved in mitochondrial dynamics are highly expressed in the heart. It has been previously reported that dynamin related protein 1 (DRP1) has a critical role in mitochondrial fitness and cardiac protection. Cardiac DRP1 ablation in the adult heart evokes a progressive dilated cardiac myopathy and lethal heart failure. Nevertheless, the conditional cardiacspecific DRP1 knock out animals present a significantly longer survival rate compared with global DRP1 KO models. We have described before the great importance for cardiac physiology of the strategic positioning of mitochondrial proteins in the cardiac tissue. Therefore, we hypothesize that DRP1 plays a regulatory role in cardiac physiology and mitochondrial fitness by preferentially accumulating at mitochondria and junctional sarcoplasmic reticulum (jSR) contact sites, where the high Ca2+ microdomain is formed during excitation-contraction (EC) coupling. Objective: This study aims to determine whether mitochondria-associated DRP1 is preferentially accumulated in the mitochondria and jSR contact sites and if indeed this is the case, what is the mechanism responsible for such a biased distribution and what is the functional implication. Methods and Results: Using high-resolution imaging approaches, we found that mitochondria-associated DRP1 in cardiomyocytes was localized in the discrete regions where T-tubule, jSR, and mitochondria are adjacent to each other. Western blot results showed that mitochondria-bound DRP1 was restricted to the mitochondria-associated membranes (MAM), with undetectable levels in purified mitochondria. Furthermore, in comparison to the cytosolic DRP1, the membrane-bound DRP1 in SR and MAM fractions formed high molecular weight oligomers. In both electrically paced adult cardiomyocytes and Langendorff-perfused beating hearts, the oscillatory Ca2+ pulses preserved MAM-associated DRP1 accumulation. Interestingly, similar to DRP1, all mitochondria-bound βACTIN only exists in MAM and not in the purified mitochondria. Additionally, co-immunoprecipitation pulls down both DRP1 and βACTIN together. Inhibition of βACTIN polymerization with Cytochalasin D disrupts the tight association between DRP1 and βACTIN. In cardiac specific DRP1 knockout mouse after 6 weeks of tamoxifen induction the cardiomyocytes show disarray of sarcomere, a decrease of cardiac contraction, loss of mitochondrial membrane potential significantly decreased spare respiratory capacity, and frequent occurrence of earl after contraction, suggesting the heart is susceptible for failure and arrhythmias. Despite of this phenotype, DRP1icKo animal have a longer life spam than other DRP1 KO models. We also observed that DRP1icKO. Strikingly, DRP1 levels are is only modestly decreased in the MAM when compared with the rest of the cellular fractions. These preserved levels were accompanied with preservation of the mitochondrial pool in the MAM fraction obtained from the DRP1icKO hearts. Conclusions: The results show that in adult cardiomyocytes, mitochondria bound DRP1 clusters in high molecular weight protein complexes at MAM. This clustering is fortified by EC coupling mediated Ca2+ transients and requires its interaction with βACTIN. Together with the better preserved dRP1 levels in the DRP1icKO model in the MAM, we conclude that DRP1 is anchored in mitochondria-SR interface through βACTIN and position itself to play a fundamental role in regulating mitochondrial quality control in the working heart.

Derivation of the material models for ultra-high molecular-weight polyethylene fiber-reinforced armor-grade composites with different architectures

2016 ◽  
Vol 33 (3) ◽  
Author(s):  
Mica Grujicic ◽  
Jennifer Snipes ◽  
S. Ramaswami ◽  
Vasudeva Avuthu ◽  
Chian-Fong Yen ◽  
...  
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Design Methodology ◽  
Critical Role ◽  
Fiber Reinforcement ◽  
Mechanical Tests ◽  
Transverse Impact ◽  
Material Models ◽  
Polyethylene Fiber ◽  
Ultra High Molecular Weight

Purpose To overcome the problem of inferior through-the-thickness mechanical properties displayed by armor-grade composites based on 2-D reinforcement architectures, armor-grade composites based on 3D fiber-reinforcement architectures have recently been investigated experimentally. Design/methodology/approach The subject of the present work is armor-grade composite materials reinforced using ultra-high-molecular-weight polyethylene fibers and having four (two 2D and two 3D) prototypical architectures, as well as the derivation of the corresponding material models. The effect of the reinforcement architecture is accounted for by constructing the appropriate unit cells (within which the constituent materials and their morphologies are represented explicitly) and subjecting them to a series of virtual mechanical tests. The results obtained are used within a post-processing analysis to derive and parameterize the corresponding homogenized-material models. One of these models (specifically, the one for 0°/90° cross-collimated fiber architecture) was directly validated by comparing its predictions with the experimental counterparts. The other models are validated by examining their physical soundness and details of their predictions. Lastly, the models are integrated as user-material subroutines, and linked with a commercial finite-element package, in order to carry out a transient non-linear dynamics analysis of ballistic transverse impact of armor-grade composite-material panels with different reinforcement architectures. Findings It is found that the reinforcement architecture plays a critical role in the overall ballistic limit of the armor panel, as well as in its structural and damage/failure response. Originality/value To the authors’ knowledge, the present work is the first reported attempt to assess, computationally, the utility and effectiveness of 3D fiber-reinforcement architectures for ballistic impact applications.

High-Molecular-Weight, High-Unsaturation Copolymers of Isobutylene and Conjugated Dienes. I. Synthesis

1976 ◽  
Vol 49 (4) ◽  
pp. 960-966 ◽  
Author(s):  
W. A. Thaler ◽  
D. J. Buckley
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Cationic Polymerization ◽  
Critical Role ◽  
Conjugated Dienes ◽  
Solution Properties ◽  
Insight Into

Abstract This investigation has provided insight into the critical role of solution properties on the course of cationic polymerization and has led to a technique for reconciling the opposing demands for synthesis of isoolefin-diene copolymers both free from gel and high in molecular weight. By this technique, the synthesis of a broad range of intermediate-unsaturation elastomers of quality has been realized.

scholarly journals Single particle characterization of biomass burning organic aerosol (BBOA): evidence for non-uniform mixing of high molecular weight organics and potassium

2015 ◽  
Vol 15 (22) ◽  
pp. 32157-32183 ◽  
Author(s):  
A. K. Y. Lee ◽  
M. D. Willis ◽  
R. M. Healy ◽  
J. M. Wang ◽  
C.-H. Jeong ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Black Carbon ◽  
High Molecular Weight ◽  
Biomass Burning ◽  
Single Particle ◽  
Critical Role ◽  
Organic Aerosol ◽  
Mixing State ◽  
Particle Measurements ◽  
Near Ultraviolet

Abstract. Biomass burning is a major source of black carbon (BC) and primary organic aerosol globally. In particular, biomass burning organic aerosol (BBOA) is strongly associated with atmospheric brown carbon (BrC) that absorbs near ultraviolet and visible light, resulting in significant impacts on regional visibility degradation and radiative forcing. The mixing state of BBOA can play a critical role in the prediction of aerosol optical properties. In this work, single particle measurements from a soot-particle aerosol mass spectrometer coupled with a light scattering module (LS-SP-AMS) were performed to examine the mixing state of BBOA, refractory black carbon (rBC) and potassium (K+, a tracer for biomass burning aerosol) in an air mass influenced by aged biomass burning. Cluster analysis of single particle measurements identified five BBOA-related particle types. rBC accounted for 3–14 w.t. % of these particle types on average. Only one particle type exhibited a strong ion signal for K+, with mass spectra characterized by low molecular weight organic species. The remaining four particle types were classified based on the apparent molecular weight of the BBOA constituents. Two particle types were associated with low potassium content and significant amounts of high molecular weight (HMW) organic compounds. Our observations indicate non-uniform mixing of particles within a biomass burning plume in terms of molecular weight and illustrate that HMW BBOA can be a key contributor to low-volatility BrC observed in BBOA particles.

scholarly journals Serum stimulation of CCR7 chemotaxis due to coagulation factor XIIa-dependent production of high-molecular-weight kininogen domain 5

2016 ◽  
Vol 113 (45) ◽  
pp. E7059-E7068 ◽  
Author(s):  
Manish P. Ponda ◽  
Jan L. Breslow
Keyword(s):  
Molecular Weight ◽  
Lymph Nodes ◽  
High Molecular Weight ◽  
Matrix Protein ◽  
Critical Role ◽  
Coagulation Factor ◽  
Extracellular Matrix Protein ◽  
High Molecular Weight Kininogen ◽  
Factor Xiia ◽  
Cofactor Activity

Chemokines and their receptors play a critical role in immune function by directing cell-specific movement. C-C chemokine receptor 7 (CCR7) facilitates entry of T cells into lymph nodes. CCR7-dependent chemotaxis requires either of the cognate ligands C-C chemokine ligand 19 (CCL19) or CCL21. Although CCR7-dependent chemotaxis can be augmented through receptor up-regulation or by increased chemokine concentrations, we found that chemotaxis is also markedly enhanced by serum in vitro. Upon purification, the serum cofactor activity was ascribed to domain 5 of high-molecular-weight kininogen. This peptide was necessary and sufficient for accelerated chemotaxis. The cofactor activity in serum was dependent on coagulation factor XIIa, a serine protease known to induce cleavage of high-molecular-weight kininogen (HK) at sites of inflammation. Within domain 5, we synthesized a 24-amino acid peptide that could recapitulate the activity of intact serum through a mechanism distinct from up-regulating CCR7 expression or promoting chemokine binding to CCR7. This peptide interacts with the extracellular matrix protein thrombospondin 4 (TSP4), and antibodies to TSP4 neutralize its activity. In vivo, an HK domain 5 peptide stimulated homing of both T and B cells to lymph nodes. A circulating cofactor that is activated at inflammatory foci to enhance lymphocyte chemotaxis represents a powerful mechanism coupling inflammation to adaptive immunity.

High molecular weight forms of IGF-II ('big-IGF-II') released by Wilms' tumor cells

1997 ◽  
pp. 396-401 ◽  
Author(s):  
S Schmitt ◽  
Q Ren-Qiu ◽  
T Torresani ◽  
M Doebeli ◽  
J Zapf ◽  
...  
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Tumor Cells ◽  
Culture Medium ◽  
Wilms Tumor ◽  
Critical Role ◽  
Polyclonal Antiserum ◽  
Mrna Levels ◽  
Rabbit Polyclonal Antiserum

Insulin-like growth factor-II (IGF-II) is thought to play a critical role in the development of embryonic tumors such as Wilms' tumor. However, despite highly elevated IGF-II mRNA levels in tumors, IGF-II is not elevated in the serum of patients with Wilms' tumors. Recently high molecular weight forms of IGF-II ('big'- or pro-IGF-II) have been found to be produced by some tumors. In order to prove whether or not high molecular weight forms of IGF-II are produced by Wilms' tumor cells and secreted into the culture medium, we established Wilms' tumor cell lines. After column chromatography of the culture medium, IGF-II and pro-IGF-II concentrations were measured. For pro-IGF-II measurement we established a pro-IGF-II RIA using a rabbit polyclonal antiserum directed against amino acids 7-21 (E7-21) of the E-domain of pro-IGF-II. Gel electrophoresis and Western blotting with anti-IGF-II antibodies revealed a band at 7.5 kDa corresponding to fully processed IGF-II and bands between 10 and 20 kDa. Using pro-IGF-II antiserum, bands between 10 and 25 kDa were detected. We conclude that in vitro cultured Wilms' tumor cells produce and release various forms of 'big IGF-II' with molecular masses between 10 and 25 kDa. It remains uncertain whether these high molecular weight forms of IGF-II represent normal precursors of IGF-II or incorrectly processed IGF-II.

scholarly journals Single-particle characterization of biomass burning organic aerosol (BBOA): evidence for non-uniform mixing of high molecular weight organics and potassium

2016 ◽  
Vol 16 (9) ◽  
pp. 5561-5572 ◽  
Author(s):  
Alex K. Y. Lee ◽  
Megan D. Willis ◽  
Robert M. Healy ◽  
Jon M. Wang ◽  
Cheol-Heon Jeong ◽  
...  
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Biomass Burning ◽  
Single Particle ◽  
Radiative Forcing ◽  
Critical Role ◽  
Organic Aerosol ◽  
Mixing State ◽  
Particle Measurements ◽  
Near Ultraviolet

Abstract. Biomass burning organic aerosol (BBOA) can be emitted from natural forest fires and human activities such as agricultural burning and domestic energy generation. BBOA is strongly associated with atmospheric brown carbon (BrC) that absorbs near-ultraviolet and visible light, resulting in significant impacts on regional visibility degradation and radiative forcing. The mixing state of BBOA can play a critical role in the prediction of aerosol optical properties. In this work, single-particle measurements from a Soot-Particle Aerosol Mass Spectrometer coupled with a light scattering module (LS-SP-AMS) were performed to examine the mixing state of BBOA, refractory black carbon (rBC), and potassium (K, a tracer for biomass burning aerosol) in an air mass influenced by wildfire emissions transported from northern Québec to Toronto, representing aged biomass burning plumes. Cluster analysis of single-particle measurements identified five BBOA-related particle types. rBC accounted for 3–14 wt % of these particle types on average. Only one particle type exhibited a strong ion signal for K+, with mass spectra characterized by low molecular weight organic species. The remaining four particle types were classified based on the apparent molecular weight of the BBOA constituents. Two particle types were associated with low potassium content and significant amounts of high molecular weight (HMW) organic compounds. Our observations indicate non-uniform mixing of particles within a biomass burning plume in terms of molecular weight and illustrate that HMW BBOA can be a key contributor to low-volatility BrC observed in BBOA particles. The average mass absorption efficiency of low-volatility BBOA is about 0.8–1.1 m2 g−1 based on a theoretical closure calculation. Our estimates indicate that low-volatility BBOA contributes ∼ 33–44 % of thermo-processed particle absorption at 405 nm; and almost all of the BBOA absorption was associated with low-volatility organics.

scholarly journals Modification of Synaptic Plasma Membrane Ca 2+ -ATPase in Ischemic Injury in an Animal Model of Global Ischemia

Stroke ◽  
2001 ◽  
Vol 32 (suppl_1) ◽  
pp. 354-354
Author(s):  
Asma Zaidi ◽  
Angela M Cross ◽  
Jennifer L Bean ◽  
Mary L Michaelis
Keyword(s):  
Molecular Weight ◽  
Animal Model ◽  
Plasma Membrane ◽  
High Molecular Weight ◽  
Critical Role ◽  
Ischemic Injury ◽  
Global Ischemia ◽  
Synaptic Membranes ◽  
Synaptic Plasma Membrane ◽  
Reducing Conditions

P83 Regulation of [Ca 2+ ] i is altered in neurons during ischemic injury in stroke, but the precise mechanism(s) underlying the Ca 2+ dysregulation are not known. The plasma membrane Ca 2+ -ATPase (PMCA) is one of the two main Ca 2+ extrusion systems that play a critical role in maintaining neuronal Ca 2+ homeostasis. We have substantial evidence showing that this enzyme is very sensitive to oxidative stress. When exposed to very low concentrations of physiologically relevant oxidants, the PMCA has been shown to form high molecular weight aggregates and this is accompanied by the decline in its enzymatic activity. Because stroke-induced hypoxia is associated with the overproduction of reactive oxygen species (ROS) and a loss of Ca 2+ homeostasis, we carried out studies to determine if the PMCA is modified in an in vivo animal model of stroke. Global ischemia was induced in Sprague Dawley rats by occlusion of the common carotid arteries for defined time periods. The V max of PMCA activity in brain homogenates as well as in purified synaptic plasma membranes was significantly reduced following occlusion of the vessels, and the reduction was proportional to the time of ischemia. The loss in PMCA activity could not be reversed by addition of exogenous ATP, suggesting an alteration in the protein itself. Immunoblots of the synaptic membranes run under non-reducing conditions showed crosslinking of PMCA molecules to form high molecular weight adducts, and this too increased with increasing periods of ischemia. The PMCA aggregates were partially reversed under reducing conditions, indicating the involvement of sulphydryl group oxidation. These observations support the hypothesis that increased formation of ROS under ischemic conditions can oxidatively modify specific proteins critical for the regulation of free [Ca 2+ ] i levels. Such oxidative damage is likely to contribute to the loss of neuronal Ca 2+ regulation and neuronal viability in stroke. (Supported by AHA 9960343Z, AG 12993, and the Higuchi Biosciences Center, University of Kansas)

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