DOCSLIB.ORG

Proteasomal-Dependent Aggregate Reversal and Absence of Cell Death in a Conditional Mouse Model of Huntington’S Disease

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Proteasomal-Dependent Aggregate Reversal and Absence of Cell Death in a Conditional Mouse Model of Huntington’S Disease The Journal of Neuroscience, November 15, 2001, 21(22):8772–8781 Proteasomal-Dependent Aggregate Reversal and Absence of Cell Death in a Conditional Mouse Model of Huntington’s Disease Ester Martı´n-Aparicio,1 Ai Yamamoto,2 Fe´ lix Herna´ ndez,1 Rene´ Hen,2 Jesu´ s Avila,1 and Jose´ J. Lucas1 1Centro de Biologı´a Molecular “Severo Ochoa,” Consejo Superior de Investigaciones Cientı´ficas–Universidad Auto´ noma de Madrid, 28049 Madrid, Spain, and 2Center for Neurobiology and Behavior, Columbia University, New York, New York 10032 Neuronal intranuclear inclusions are a histopathological hall- aggregate formation, and5doftransgene suppression led to mark of Huntington’s disease. Nevertheless, the precise mech- aggregate disappearance. In mice, full reversal of aggregates anism by which they are formed and their relevance to neuronal and intranuclear mutant huntingtin was more rapid than re- cell death and/or dysfunction remains unclear. We recently ported previously and preceded the motor recovery by several generated a conditional mouse model of Huntington’s disease weeks. Furthermore, the proteasome inhibitor lactacystin inhib- (HD94) in which silencing expression of mutated huntingtin led ited the aggregate clearance observed in culture, thus indicat- to the disappearance of intranuclear aggregates and ameliora- ing that aggregate formation is a balance between the rate of tion of the behavioral phenotype. Here, we analyze primary huntingtin synthesis and its degradation by the proteasome. striatal neuronal cultures from HD94 mice to explore the dy- Finally, neither expression of the mutant huntingtin nor aggre- namics of aggregate formation and reversal, the possible gates compromised the viability of HD94 cultures. This corre- mechanisms involved, and the correlation between aggregates lated with the lack of cell death in symptomatic HD94 mice, and neuronal death. In parallel, we examine symptomatic adult thus demonstrating that neuronal dysfunction, and not cell loss, HD94 mice in similar studies and explored the relationship triggered by mutant huntingtin underlies symptomatology. between aggregate clearance and behavioral reversal. We re- Key words: Huntington’s disease; aggregates; conditional port that, in culture, aggregate formation and reversal were transgenic mouse model; fast reversal; proteasome; absence of rapid processes, such that2doftransgene expression led to cell death Huntington’s disease (HD) is a progressive, autosomal dominant, respective proteins. These triplet-repeat disorders share an inter- neurodegenerative disorder (Wexler et al., 1987). Patients suffer esting commonality: the presence of intraneuronal aggregates from motor dysfunction, cognitive decline, and psychological dis- (Ross, 1997; Nakamura et al., 2001). turbances over 10–15 years until death (Ambrose et al., 1994). The relevance of aggregate formation to the etiology of HD is The neuropathology is extremely restricted, with atrophy occur- unclear. Aggregates have been implicated as the trigger for neu- ring in the striatum and to a lesser extent in the cortex. rodegeneration, because both aggregates and pathogenicity are The mutation leading to HD is an expansion of CAG repeats caused by the same polyQ length threshold (Scherzinger et al., near the 5Ј end of the IT15 gene (The Huntington’s Disease 1997). Furthermore, in a transgenic mouse model, these aggre- Collaborative Research Group, 1993; Rubinsztein et al., 1994; gates precede the onset of symptomatology (Davies et al., 1997). Gusella and MacDonald, 1996). Although unaffected individuals Inclusions are also a feature in other non-polyQ disorders, such as Ͼ have 35 or fewer repeats, repeat lengths of 40 invariably lead to Alzheimer’s disease and Parkinson’s disease, thus suggesting that HD. The repeat sequence is translated into a polyglutamine protein aggregation might cause neurodegeneration through a stretch (polyQ) near the N terminus of the IT15 gene product general mechanism (Price et al., 1998; Tobin and Signer, 2000). huntingtin (htt). Eight other autosomal dominant neurological Moreover, proteins prone to aggregate, such as mutated hunting- diseases are also caused by a polyQ expansion mutation in their tin and cystic fibrosis proteins, have been reported to inhibit the ubiquitin proteasome system (UPS) (Bence et al., 2001). This Received July 2, 2001; revised Aug. 23, 2001; accepted Sept. 5, 2001. This work was supported by grants from Asociacio´n Espan˜ola de Corea de inhibition in turn altered the half-life of proteins involved in Huntington (ACHE), by the United States–Spain Commission for Scientific and apoptosis and cell survival (Jana et al., 2001). Nevertheless, Technological Cooperation, by the Hereditary Disease Foundation (HDF), and by whereas some studies using transfected cells support aggregate- an institutional grant from Fundacio´n Ramo´n Areces. E.M.-A. and A.Y. are recip- ients of predoctoral fellowships from ACHE and Fundacio´n Ferrer and from HDF, induced toxicity (Waelter et al., 2001), others dissociate inclusion respectively. We thank Drs. Javier Dı´az-Nido, Francisco Wandosell, Mar Pe´rez, and formation and cell survival (Kim et al., 1999) and even suggest Jose´ Gonza´lez-Castan˜o for helpful discussion and comments; Drs. Eric Kandel that intranuclear inclusions may reflect a protective mechanism and Mark Mayford for the use of the CaMKII ␣-tTA mice; Drs. Erich E. Wanker and Xiao-Jiang Li for kindly providing CAG53b and EM48 antibodies, respectively; against soluble htt-induced toxicity (Saudou et al., 1998). Drs. Victoria Arango and Mark Underwood for stereology; and Dr. Gine´s Morata Using a conditional animal model of HD that reversibly ex- for microscope and photography facility. We are also grateful to Carlos Sa´nchez, Raquel Cuadros, Helena Garuti, and Elena Langa for technical assistance. pressed a mutated htt fragment selectively in the forebrain Correspondence should be addressed to Jose´ J. Lucas, Centro de Biologı´a (HD94) (Yamamoto et al., 2000), we found that abolishing trans- Molecular “Severo Ochoa,” Consejo Superior de Investigaciones Cientı´ficas–Uni- gene expression in symptomatic mice leads to amelioration of the versidad Auto´noma de Madrid, Facultad Ciencias, Universidad Auto´noma de Ma- drid, Cantoblanco, 28049 Madrid, Spain. E-mail: [email protected]. behavioral phenotype and disappearance of inclusions. This dem- Copyright © 2001 Society for Neuroscience 0270-6474/01/218772-10$15.00/0 onstrated that inclusions are dynamic structures and that a con- Martı´n-Aparicio et al. • HD Aggregate Clearance Is Proteasome Dependent J. Neurosci., November 15, 2001, 21(22):8772–8781 8773 tinuous influx of the protein is required for disease progression. ␤-Galactosidase activity assays In this initial characterization, however, the minimum time the LacZ staining. Cultures were fixed for 20 min in 4% PFA in 5 mM cell required to clear the aggregates versus the time the organism MgCl2–PBS. Slides were incubated for 1 hr at 30°C in an X-gal solution: required to recover its motor behavior was not established, thus 1 mg/ml X-gal (Boehringer Mannheim, Indianapolis, IN), 5 mM potas- sium ferrocyanide, 5 mM potassium ferricyanide, and 2 mM MgCl2 in hindering the search for possible mechanisms that underlie the PBS. After staining, cultures were coverslipped with Fluoromount. recovery process. In light of this, we have established primary O-nitrophenyl ␤-D-calactopyranoside colorimetric assay. Cultures were neuronal cultures from HD94 mice. With this system, we explore washed twice with 5 mM MgCl2 in PBS. Cells were lysed in 250 mM Tris the dynamics of aggregate formation and reversal, the mechanism buffer, pH 8.0, with 0.1% Triton X-100. Enzymatic assay was performed responsible for reversal, and the correlation between aggregate as described previously (Sambrook et al., 1989). formation and cell death. As we gained more insight from the culture system, we then returned to the animal model and reex- Survival assays amined the issues sought in vitro. Propidium iodide–calcein staining. Twenty-four hours after plating, cul- tures were shifted to media without serum, B27, or both. After 24, 48, or 72 hr, cell viability was assessed by propidium iodide–calcein staining MATERIALS AND METHODS (Mattson et al., 1995). Calcein-AM is taken up and cleaved by esterases present within living cells and leads to a yellowish-green fluorescence, Animals whereas propidium iodide (PI) is taken up by only dead cells and become HD94 mice were generated as described previously (Yamamoto et al., orange-red fluorescent. In brief, neurons were incubated for 30 min with 2000). Mice were bred at the Centro de Biolog´ıa Molecular “Severo 2 ␮M PI (Sigma) and 1 ␮M calcein-AM (Molecular Probes, Eugene, OR). Ochoa” (Madrid, Spain) and Center for Neurobiology and Behavior After several brief rinses with each medium, cells were visualized by (Columbia, NY). Four to five mice were housed per cage, with food and fluorescence microscopy using a Zeiss (Oberkochen, Germany) Axiovert water available ad libitum. Mice were maintained in a temperature- 135 microscope. Four fields (selected at random) were analyzed per well controlled environment on a 12 hr light/dark cycle, with light onset at (ϳ300 cells per field) in two independent experiments. Cell death was 7:00 A.M. expressed as percentage of PI-positive cells from the total number of cells. Primary culture Hoechst 33342 nuclear staining. Neurons were fixed in 4% PFA in 5 mM Primary cultures of neurons and glia were prepared according to modi- MgCl2-PBS and pretreated with 0.2% Triton X-100 in PBS. After that, fications of established procedures (Huettner and Baughman, 1986; cultures were incubated with Hoechst dye 33342 (2.5 ␮g/ml; Biologics) Dubinsky, 1989). Pups were killed at postnatal day 1, and tail tissue and visualized under a fluorescence microscope. The percentage of samples were taken for ulterior genotyping by PCR. Striatal tissue was condensed or fragmented nuclei was counted for each genotype and then dissected and dissociated individually from each pup with the culture condition at 20ϫ magnification. Data are reported as the mean Ϯ Papain Dissociation System (Worthington, Freehold, NJ). The rest of SEM (n ϭ 3).
Load more

Recommended publications
  • Classification of Cell Death
    Cell Death and Differentiation (2005) 12, 1463–1467 & 2005 Nature Publishing Group All rights reserved 1350-9047/05 $30.00 www.nature.com/cdd News and Commentary Classification of cell death: recommendations of the Nomenclature Committee on Cell Death G Kroemer*,1, WS El-Deiry2, P Golstein3, ME Peter4, D Vaux5, with or without, caspase activation and that ‘autophagic cell P Vandenabeele6, B Zhivotovsky7, MV Blagosklonny8, death’ represents a type of cell death with (but not necessarily W Malorni9, RA Knight10, M Piacentini11, S Nagata12 and through) autophagic vacuolization. This article details the G Melino10,13 2005 recommendations of NCCD. Over time, molecular definitions are expected to emerge for those forms of cell 1 CNRS-UMR8125, Institut Gustave Roussy, 39 rue Camille-Desmoulins, death that remain descriptive. F-94805 Villejuif, France 2 University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA Preface 3 Centre d’Immunologie INSERM/CNRS/Universite de la Mediterranee de Marseille-Luminy, Case 906, Avenue de Luminy, 13288 Marseille Cedex 9, It is obvious that clear definitions of objects that are only France shadows in Plato’s cage are difficult to be achieved. Cell death 4 The Ben May Institute for Cancer Research, University of Chicago, 924 E 57th and the different subroutines leading to cell death do not Street, Chicago, IL 60637, USA 5 escape this rule. Even worse, the notion of death is strongly Sir William Dunn School of Pathology, University of Oxford, Oxford OX1 3RE, influenced by religious and cultural beliefs, which may UK 6 Molecular Signalling and Cell Death Unit, Department for Molecular Biomedical subliminally influence the scientific view of cell death.
    [Show full text]
  • Analysis of Different Evolutionary Strategies to Prevent Protein Aggregation
    Universitat Autònoma de Barcelona Departament de Bioquímica i Biologia Molecular and Institut de Biotecnologia i Biomedicina Analysis of Different Evolutionary Strategies to Prevent Protein Aggregation Ricardo Graña Montes Bellaterra, December 2014 Universitat Autònoma de Barcelona Departament de Bioquímica i Biologia Molecular and Institut de Biotecnologia i Biomedicina Analysis of Different Evolutionary Strategies to Prevent Protein Aggregation Doctoral thesis submitted by Ricardo Graña Montes in candidacy for the degree of Ph.D. in Biochemistry, Molecular Biology and Biomedicine from the Universitat Autònoma de Barcelona. The work described herein has been performed at the Department de Bioquímica i Biologia Molecular and at the Institut de Biotecnologia i Biomedicina, under the supervision of Prof. Salvador Ventura Zamora. Ricardo Graña Montes Prof. Salvador Ventura Zamora Bellaterra, December 2014 SUMMARY IN ENGLISH In the last 15 years, the study of protein aggregation has evolved from a mostly neglected topic of protein chemistry to a highly dynamic research area which has expanded its implications through different fields including biochemistry, biotechnology, nanotechnology and biomedicine. The analysis of protein aggregation has attracted a particular interest in the biomedical and biotechnological areas. Because, on one side, the formation of insoluble protein deposits is associated to an increasing number of human disorders, many of which present fatal pathological consequences. And on the other hand, aggregation is a frequent shortcoming in the recombinant expression of proteins at the industrial level, such as in the production of proteinaceous therapeutic agents like antibodies. Consequently, the survey of mechanisms to prevent protein aggregation is currently the focus of deep investigation with the aim to develop preventive or therapeutic methods for the intervention of these depositional disorders and to enhance the yield in the biotechnological production of proteins.
    [Show full text]
  • Large-Scale Analysis of Macromolecular Crowding Effects on Protein Aggregation Using a Reconstituted Cell-Free Translation System
    DATA REPORT published: 08 October 2015 doi: 10.3389/fmicb.2015.01113 Large-scale analysis of macromolecular crowding effects on protein aggregation using a reconstituted cell-free translation system Tatsuya Niwa 1†, Ryota Sugimoto1† , Lisa Watanabe1, Shugo Nakamura2, Takuya Ueda3 and Hideki Taguchi1* 1 Department of Biomolecular Engineering, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Yokohama, Japan, 2 Department of Biotechnology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Tokyo, Japan, 3 Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Chiba, Japan Edited by: Proteins must fold into their native structures in the crowded cellular environment, to Salvador Ventura, perform their functions. Although such macromolecular crowding has been considered Universitat Autònoma de Barcelona, to affect the folding properties of proteins, large-scale experimental data have so far Spain Reviewed by: been lacking. Here, we individually translated 142 Escherichia coli cytoplasmic proteins Dong-Woo Lee, using a reconstituted cell-free translation system in the presence of macromolecular Kyungpook National University, South crowding reagents (MCRs), Ficoll 70 or dextran 70, and evaluated the aggregation Korea Pierre Genevaux, propensities of 142 proteins. The results showed that the MCR effects varied depending Centre National de la Recherche on the proteins, although the degree of these effects was modest. Statistical analyses Scientifique, France suggested that structural parameters were involved in the effects of the MCRs. Our *Correspondence: dataset provides a valuable resource to understand protein folding and aggregation Hideki Taguchi [email protected] inside cells. †These authors have contributed Keywords: protein aggregation, protein folding, cell-free translation system, macromolecular crowding, large- scale analysis equally to this work.
    [Show full text]
  • Prion-Like Proteins in Phase Separation and Their Link to Disease
    biomolecules Review Prion-Like Proteins in Phase Separation and Their Link to Disease Macy L. Sprunger and Meredith E. Jackrel * Department of Chemistry, Washington University, St. Louis, MO 63130, USA; [email protected] * Correspondence: [email protected] Abstract: Aberrant protein folding underpins many neurodegenerative diseases as well as certain myopathies and cancers. Protein misfolding can be driven by the presence of distinctive prion and prion-like regions within certain proteins. These prion and prion-like regions have also been found to drive liquid-liquid phase separation. Liquid-liquid phase separation is thought to be an important physiological process, but one that is prone to malfunction. Thus, aberrant liquid-to-solid phase transitions may drive protein aggregation and fibrillization, which could give rise to pathological inclusions. Here, we review prions and prion-like proteins, their roles in phase separation and disease, as well as potential therapeutic approaches to counter aberrant phase transitions. Keywords: prions; prion-like domains; amyloid; protein misfolding; liquid-liquid phase separation; aberrant phase transitions; chaperones 1. Introduction Protein folding is essential for life, as proteins serve structural roles, catalyze enzy- matic reactions, and transport materials through membranes and cells, among many other Citation: Sprunger, M.L.; Jackrel, functions [1]. Most globular proteins adopt a complex three-dimensional folded struc- M.E. Prion-Like Proteins in Phase ture that is well-defined and associated with specific functions. In contrast, intrinsically Separation and Their Link to Disease. disordered proteins (IDPs) do not adopt well-defined structures. Rather, IDPs occupy a Biomolecules 2021, 11, 1014. https:// highly dynamic ensemble of conformations, and these dynamic structural changes are doi.org/10.3390/biom11071014 key to the function of these proteins [2].
    [Show full text]
  • The Biochemistry of Cell Death Cell Death: Apoptosis and Other Means to an End, Second Edition by Douglas R
    Zampieri et al. Cell Death and Disease (2020) 11:259 https://doi.org/10.1038/s41419-020-2465-5 Cell Death & Disease BOOK REVIEW Open Access The biochemistry of cell death Cell Death: Apoptosis and Other Means to an End, Second Edition by Douglas R. Green, St. Jude Children’s Research Hospital, Cold Spring Harbor Laboratory Press, New York, 2018 Carlotta Zampieri 1, Carlo Ganini 1 and Gerry Melino 1 Can we impress you with a mind-blowing revelation? Douglas Green is one of the worldwide leading experts “Cells are not eternal in our bodies, but rather they on apoptosis and cell death. He has dealt with the role of encounter death!”1. cell death in the regulation of cancer and immune If someone pronounces this sentence at a biology class, response, studying the molecular events that drive the students will laugh. Why? Because cell death has been process, focusing on the induction-activation of apoptosis studied since the 60s of the last century. When we speak in T cells and the role of Myc, death receptors and Bcl-2 about apoptosis nowadays, we regard it as a well-known in this context. His deep understanding of the field truth. Anyway, this truth did not catch the attention of allowed him to write the first edition of this book in a scientists until the 1980s, where the interest in the field clear and straightforward manner, enriched by extremely exploded, leading to a dramatic increase of publications. informative figures. His talent as a biologist, as well as a Cell death passed “from neglect to hysteria” in only a few communicator, has been condensed in this second edi- years, citing Martin Raff2, one of the founders of the field.
    [Show full text]
  • Dual Effects of Thyroid Hormone on Neurons and Neurogenesis
    Lin et al. Cell Death and Disease (2020) 11:671 https://doi.org/10.1038/s41419-020-02836-9 Cell Death & Disease ARTICLE Open Access Dual effects of thyroid hormone on neurons and neurogenesis in traumatic brain injury Chao Lin1,2, Nan Li3, Hanxiao Chang1,2,Yuqishen1,2,ZhengLi1,2,Wuwei1,2,HuaChen1,2,HuaLu1,2,JingJi 1,2 and Ning Liu1,2 Abstract Thyroid hormone (TH) plays a crucial role in neurodevelopment, but its function and specific mechanisms remain unclear after traumatic brain injury (TBI). Here we found that treatment with triiodothyronine (T3) ameliorated the progression of neurological deficits in mice subjected to TBI. The data showed that T3 reduced neural death and promoted the elimination of damaged mitochondria via mitophagy. However, T3 did not prevent TBI-induced cell death in phosphatase and tensin homolog (PTEN)-induced putative kinase 1 (Pink1) knockout mice suggesting the involvement of mitophagy. Moreover, we also found that T3 promoted neurogenesis via crosstalk between mature neurons and neural stem cells (NSCs) after TBI. In neuron cultures undergoing oxygen and glucose deprivation (OGD), conditioned neuron culture medium collected after T3 treatment enhanced the in vitro differentiation of NSCs into mature neurons, a process in which mitophagy was required. Taken together, these data suggested that T3 treatment could provide a therapeutic approach for TBI by preventing neuronal death via mitophagy and promoting neurogenesis via neuron–NSC crosstalk. 1234567890():,; 1234567890():,; 1234567890():,; 1234567890():,; Introduction treatments focus only on preventing complications or Traumatic brain injury (TBI) is considered to be a providing support in nature. leading cause of substantial mortality and long-term dis- Thyroid hormone (TH) is crucial for neural stem cell ability among young adults worldwide1.
    [Show full text]
  • Estimation of Postmortem Interval Based on Cell Death Progression in Biological Fluids
    Estimation of Postmortem Interval Based on Cell Death Progression in Biological Fluids Author: Mónica Cristina Francisco Tomé Dissertation submitted to the Faculty of Medicine of University of Porto for the Master degree in Forensic Sciences Supervisor: Prof. Doutor Agostinho Almiro de Almeida (Faculty of Pharmacy, University of Porto) Co-supervisors: Prof. Doutor Agostinho José Carvalho dos Santos (Faculty of Medicine, University of Porto) Doutora Daniela Sofia Almeida Ribeiro (Faculty of Pharmacy, University of Porto) Porto, September 2017 Acknowledgments My special acknowledgments to: My supervisor Professor Agostinho Almeida that accepted to work with me and provided me the indispensable help to finish my master degree. My co-supervisor Professor Agostinho Santos, without him I would not be doing my dissertation, he opened my eyes and I am glad that he did. My co-supervisor Doctor Daniela Ribeiro that welcomed me with open arms and was always with me. Doctor Rui Almeida who is a very professional person and who was always ready to help me in whatever it takes! To Professor Eduarda Fernandes who was always present during the development of this study and was always trying to find solutions to the problems. To all of my friends, especially to Cátia Pereira, Margarida Pereira, Miguel Pinto and Sofia Salsinha, that provided me the emotional strength to continue to fight and get my motivation. To my parents and brother, that always listened to me and gave me the emotional support while I was far away from home. I have to thank also to all of those who were present during this period of my life and never let me give up.
    [Show full text]
  • The Dynamic Tumor Ecosystem: How Cell Turnover and Trade-Offs Affect Cancer Evolution
    bioRxiv preprint doi: https://doi.org/10.1101/270900; this version posted February 26, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY 4.0 International license. The dynamic tumor ecosystem: how cell turnover and trade-offs affect cancer evolution Jill A. Gallaher1, Joel Brown1*, and Alexander R. A. Anderson1* 1 Department of Integrated Mathematical Oncology, H. Lee Moffitt Cancer Center, Tampa, FL USA *These authors contributed equally. ABSTRACT Tumors are not static masses of cells but rather dynamic ecosystems where cancer cells experience constant turnover and evolve fitness-enhancing phenotypes. Selection for different phenotypes may vary with 1) the tumor niche (edge or core), 2) cell turnover rates, 3) the nature of the tradeoff between traits (proliferation vs migration), and 4) whether deaths occur in response to demographic or environmental stochasticity. In an agent based, spatially-explicit model, we observe how two traits (proliferation rate and migration speed) evolve under different trade-off conditions with different turnover rates. Migration rate is favored over proliferation at the tumor’s edge and vice-versa for the interior. Increasing cell turnover rates only slightly slows the growth of the tumor, but accelerates the rate of evolution for both proliferation and migration. The absence of a tradeoff favors ever higher values for proliferation and migration. A convex tradeoff tends to favor proliferation over migration while often promoting the coexistence of a generalist and specialist phenotype.
    [Show full text]
  • Molecular and Structural Architecture of Polyq Aggregates in Yeast
    Molecular and structural architecture of polyQ PNAS PLUS aggregates in yeast Anselm Grubera,1, Daniel Hornburgb,1, Matthias Antoninc,1, Natalie Krahmerb, Javier Colladoa,d, Miroslava Schaffera, Greta Zubaitea, Christian Lüchtenborge, Timo Sachsenheimere, Britta Brüggere, Matthias Mannb, Wolfgang Baumeistera,2, F. Ulrich Hartlc,f,2, Mark S. Hippc,f,2, and Rubén Fernández-Busnadiegoa,2 aDepartment of Structural Molecular Biology, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany; bDepartment of Proteomics and Signal Transduction, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany; cDepartment of Cellular Biochemistry, Max Planck Institute of Biochemistry, 82152 Martinsried, Germany; dGraduate School of Quantitative Biosciences Munich, 81337 Munich, Germany; eHeidelberg University Biochemistry Center, 69120 Heidelberg, Germany; and fMunich Cluster for Systems Neurology (SyNergy), 80336 Munich, Germany Contributed by Wolfgang Baumeister, February 23, 2018 (sent for review October 16, 2017; reviewed by Jeffery W. Kelly and Sheena E. Radford) Huntington’s disease is caused by the expansion of a polyglutamine differences in the protein homeostasis network (17) are critical in (polyQ) tract in the N-terminal exon of huntingtin (HttEx1), but the shaping aggregate morphology and toxicity. Furthermore, ex- cellular mechanisms leading to neurodegeneration remain poorly pression of polyQ expansion proteins in yeast resulted in mor- understood. Here we present in situ structural studies by cryo- phological alterations of mitochondria and lipid droplets (LDs) electron tomography of an established yeast model system of polyQ and in reduced levels of proteins related to energy metabolism, toxicity. We find that expression of polyQ-expanded HttEx1 results some of which were sequestered by polyQ aggregates. in the formation of unstructured inclusion bodies and in some cases fibrillar aggregates.
    [Show full text]
  • Protein Aggregation
    Protein aggregation I. Amyloid assembly is an alternative to protein folding Proteins fold into native states as a result of specific interactions between amino acids. However, these interactions are not limited to intramolecular contacts - they may also occur between different molecules. Intermolecular amino acid interactions lead to formation of protein aggregates. In many cases, protein aggregates eventually are transformed into amyloid fibrils (Fig. 1). β-strand orientation Fibril axis Fibril axis Fig. 1a Generic structure of amyloid fibril is stabilized by hydrogen bonds (HBs, dashed lines), which are oriented parallel to the fibril axis and perpendicular to β−strands. This arrangement of peptides is called in-registry, because each residue in one chain is exactly matched by the same residues from the neighboring chains. Fig. 1b Electronic microphotograph of a typical amyloid fibril. As a rule amyloid fibrils appear as unbranched, long rod- or ribbon-like structures with the typical diameter of about 10 nm and the length reaching up to 1μm and more. Amyloid fibrils are exceptionally stable against chemical denaturants or temperature. Experiments show that they can withstand up to 8M urea or the temperatures as high as 100 °C (prion fibrils). From a macroscopic viewpoint, formation of amyloid fibrils is 1 essentially irreversible event. Once formed, fibrils cannot dissociate under physiological conditions. Experimental (in vitro) time scales of amyloid formation, which can be as large as days or weeks, far exceed typical folding scales of 1 msec or less. More than 20 protein sequences sharing no obvious sequence similarity are known to assemble into wild-type amyloid structures.
    [Show full text]
  • Scientific Justification of Cryonics Practice
    REJUVENATION RESEARCH Volume 11, Number 2, 2008 http://www.ncbi.nlm.nih.gov/pubmed/18321197 http://www.liebertonline.com/doi/abs/10.1089/rej.2008.0661 Scientific Justification of Cryonics Practice Benjamin P. Best* ABSTRACT Very low temperatures create conditions that can preserve tissue for centuries, possibly including the neurological basis of the human mind. Through a process called vitrification, brain tissue can be cooled to cryogenic temperatures without ice formation. Damage associated with this process is theoretically reversible in the same sense that rejuvenation is theoretically possible by specific foreseeable technology. Injury to the brain due to stopped blood flow is now known to result from a complex series of processes that take much longer to run to completion than the six minute limit of ordinary resuscitation technology. Reperfusion beyond the six minute limit primarily damages blood vessels rather than brain tissue. Apoptosis of neurons takes many hours. This creates a window of opportunity between legal death and irretrievable loss of life for human and animal subjects to be cryopreserved with possibility of future resuscitation. Under ideal conditions, the time interval between onset of clinical death and beginning of cryonics procedures can be reduced to less than a minute, but much longer delays could also be compatible with ultimate survival. Although the evidence that cryonics may work is indirect, indirect evidence is essential in many areas of science. If complex changes due to aging are reversible at some future date, then similarly complex changes due to stopped blood flow and cryopreservation may also be reversible, with life-saving results for anyone with medical needs that exceed current capabilities.
    [Show full text]
  • The Effect of Laughter on Stress and Natural Killer Cell Activity
    Nurul Husniyah binti Che Soh et al /J. Pharm. Sci. & Res. Vol. 12(12), 2020, 1496-1498 The Effect of Laughter on Stress and Natural Killer Cell Activity Nurul Husniyah binti Che Soh Graduate Student, Department of Physiology,Saveetha Dental College and Hospital, Saveetha Institute of Medical and Technical Sciences, Chennai, India. Dr. Jothipriya Assistant professor, Department Physiology, Saveetha Dental College, Saveetha Institute of Medical and Technical Sciences, Chennai. Abstract Aim: To find out the effect of laughter on stress and natural killer cell activity. Background and reason: Stress is a well-known slow killer, is rampant in our society, which impact everyone differently, but the final results are easy to observe and explain. Laughter triggers the release of a cocktail of happy chemicals, including NK cells, endorphins, serotonin, and growth hormone that are produced each time we laugh that boosts the immune responses. Laughter stimulates circulation and helps muscle relaxation which is releasing stress. Effect of laughter on stress and natural killer cells are to be reviewed as laughter significantly can reduce stress as well as improve the natural killer cells activity. Thorough literature research performed with present inclusion and exclusion criteria. Keywords: stress, natural killer cells, endorphins, apoptosis, enzymes INTRODUCTION Cousins. In a study, a group of heart attack patients were Laughter is the best way to reduce stress in people live, separated into two smaller groups; the first group was and can aid us to deal and survive a stressful lifestyle. situated under standard medical care whereas the second Since 20 years ago, it is frequently documented by group watched humorous video for about 30 minutes per psycho-neuro-immunological (PNI) research related to day.
    [Show full text]