DOCSLIB.ORG

Involvement of Protein Degradation, Calpain Autolysis and Protein

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Involvement of Protein Degradation, Calpain Autolysis and Protein Iowa State University Capstones, Theses and Graduate Theses and Dissertations Dissertations 2009 Involvement of protein degradation, calpain autolysis and protein nitrosylation in fresh meat quality during early postmortem refrigerated storage Wangang Zhang Iowa State University Follow this and additional works at: https://lib.dr.iastate.edu/etd Part of the Animal Sciences Commons Recommended Citation Zhang, Wangang, "Involvement of protein degradation, calpain autolysis and protein nitrosylation in fresh meat quality during early postmortem refrigerated storage" (2009). Graduate Theses and Dissertations. 10623. https://lib.dr.iastate.edu/etd/10623 This Dissertation is brought to you for free and open access by the Iowa State University Capstones, Theses and Dissertations at Iowa State University Digital Repository. It has been accepted for inclusion in Graduate Theses and Dissertations by an authorized administrator of Iowa State University Digital Repository. For more information, please contact [email protected]. i Involvement of protein degradation, calpain autolysis and protein nitrosylation in fresh meat quality during early postmortem refrigerated storage by Wangang Zhang A dissertation submitted to the graduate faculty in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY Major: Meat Science Program of Study Committee: Elisabeth Huff-Lonergan, Major Professor Steven Lonergan Joseph Sebranek Patricia Murphy Richard Robson Iowa State University Ames, Iowa 2009 Copyright © Wangang Zhang, 2009. All rights reserved. ii TABLE OF CONTENTS ABSTRACT GENERAL INTRODUCTION 1 INTRODUCTION TO LITERATURE REVIEW 7 Structure of skeletal muscle 8 Myofibrillar proteins 9 Sarcoplasmic proteins 31 Stromal proteins 34 Muscle contraction 37 Fresh meat quality 39 Water holding capacity 39 Drip loss formation 44 Meat tenderness 46 Muscle fibers types and meat quality 50 Calpain family 53 Calpains 53 Calpain activity assay 59 Calpain autolysis 61 Calpastatin 64 Protein modification and meat quality 66 Protein oxidation 66 Protein nitrosylation 70 Summary 76 iii Literature cited 78 METHODS TO DETERMINE THE ACTIVITY OF CALPAIN PROTEINASE 116 Abstract 116 Introduction 116 Assays and discussion 119 Summary 128 Literature cited 130 CONTRIBUTION OF POSTMORTEM CHANGES OF INTEGRIN, DESMIN AND µ-CALPAIN TO VARIATION IN WATER HOLDING CAPACITY OF PORK 134 Abstract 134 Introduction 135 Materials and methods 138 Results and discussion 142 Conclusions 150 Acknowledgement 150 Literature cited 151 Tables and Figures 155 S-NITROSOGLUTATHIONE AFFECTS THE ACTIVITY AND AUTOLYSIS OF µ- AND m-CALPAIN 161 Abstract 161 Introduction 162 iv Materials and methods 163 Results 170 Discussion 177 Conclusions 184 Literature cited 186 Tables and Figures 190 COMPARISON OF PROTEIN NITROSYLATION BETWEEN POSTMORTEM BEEF LONGISIMMUS DORSI AND PSOAS MAJOR 196 Abstract 196 Introduction 196 Materials and methods 198 Results and discussion 203 Conclusions 208 Literature cited 210 Tables and Figures 214 GENERAL SUMMARY 220 ACKNOWLEDGEMENTS 224 v ABSTRACT Fresh meat quality including meat tenderness and water holding capacity are important for consumer acceptance and industry profitability in the US. Exploring the basic mechanism that regulates the development of meat quality attributes is critical for delivering consistently high quality fresh meat to consumers. The overall hypothesis of this study was that biochemical factors that influence proteolysis during the antemortem or the early postmortem period can contribute to variations in fresh meat quality. The objective was to determine the extent to which protein modifications including protein degradation, protein nitrosylation and calpain autolysis affect the development of fresh meat quality. The first experiment was designed to test the hypothesis that protein degradation and calpain activation could influence water holding capacity of fresh pork. Intensity of intact desmin was positively correlated with moisture loss, while intensity of intact integrin was negatively correlated with moisture loss during postmortem storage. Significant correlations were found between different calpain autolysis products and intensity of intact desmin and moisture loss. These results support the hypothesis that proteolysis of desmin contributes to greater water holding capacity. The second experiment hypothesized that the properties of calpains could be regulated by a nitric oxide donor S- nitrosoglutathione. S-Nitrosoglutathione could further nitrosylate µ-calpain in both the absence and the presence of calcium especially at pH 6.5. The combination of S- nitrosoglutathione and calcium affected the activity of m- and µ-calpain and regulated the rate of µ-calpain autolysis. In the third experiment, the hypothesis that the muscle fiber type could be related to the levels of protein nitrosylation in postmortem beef was tested. vi Beef longissimus dorsi muscle showed higher percentage of type IIA/X and lower percentage of type I myosin heavy chain isoforms than psoas major muscle. The intensity of protein nitrosylation in longissimus dorsi muscle was greater compared to psoas major muscle in samples prepared 1 d postmortem. In conclusion, biochemical and biophysical reactions during preslaughter and postmortem storage can mediate the development of fresh meat quality during postmortem aging. 1 GENERAL INTRODUCTION Lack of consistency in tenderness and water holding capacity are two major problems facing the meat industry. Variations in these fresh meat attributes largely influence the satisfaction of consumers and ultimately the profitability of meat producers (Barbut et al., 2008; George, Tatum, Belk and Smith, 1999; Stetzer and McKeith, 2003). For example, 24% of US retail beef loin steaks have been categorized with values lower than slightly tender. At least 50% of pork in the US market has been found to have high drip or purge loss (Kauffman, Cassens, Scherer and Meeker, 1992). Thus, it is of particular importance to explore basic biochemical processes that govern meat quality including meat tenderization and water holding capacity. Skeletal muscles have complex structures to allow them to function. Muscle proteins on average account for 18-19% of muscle weight (Aberle, Forrest, Gerrard and Mills, 2001). In postmortem muscle, proteolysis of myofibrillar and myofibril-associated proteins can change the structure of skeletal muscle and thus influence meat quality during postmortem storage (Anderson and Parrish, 1989; Davey and Gilbert, 1966, 1969; Davis, Sebranek, Huff- Lonergan and Lonergan, 2004; Goll, Henderson and Kline, 1964; Huff-Lonergan and Lonergan, 1999, 2005, 2007; Koohmaraie, 1992; Kristensen and Purslow, 2001; Melody, Lonergan, Rowe, Huiatt, Mayes and Huff-Lonergan, 2004; Rowe, Huff-Lonergan and Lonergan, 2001; Taylor, Geesink, Thompson, Koohmaraie and Goll, 1995). Integrin plays a role in the adhesion between cell cytoskeleton and cell extracellular matrix. During postmortem aging, degradation of integrin has been found to be associated with the formation of drip channels and increased drip loss in pork (Lawson, 2004; Straadt, Rasmussen, Young, 2 and Bertram, 2008; Zhang, Lonergan, Gardner and Huff-Lonergan, 2006). Another protein, desmin is one of the intermediate filament proteins that connect adjacent myofibrils and also links the cell membrane to myofibrils. Proteolysis of desmin can remove connections between adjacent myofibrils and thus could prevent myofibril shrinkage to be transferred to the level of the entire muscle cells (Offer and Trinick, 1983). Several studies have shown that desmin degradation is associated with improved tenderness and better water holding capacity (Huff-Lonergan, Mitsuhashi, Beekman, Parrish, Olson and Robson, 1996; Kristensen and Purslow, 2001; Melody et al., 2004; Rowe et al., 2001; Zhang et al., 2006). Calpains are a family of protease enzymes that have been shown to be involved with many cellular processes. However, the model of its activation and control is still poorly understood in vivo. One of the obstacles is lack of appropriate methods to monitor the status of calpain. In postmortem muscle, the calpain system is the major factor for meat tenderization during postmortem aging (Goll, Taylor, Christiansen and Thomson, 1992; Huff-Lonergan et al., 1996). Therefore, finding some sensitive and continuous activity assays for calpain is critical to control variations of fresh meat quality. Currently, 14 members of cysteine proteases have been found in calpain family (Huang and Wang, 2001). The calpain system was originally composed of two cysteine proteases m- and µ-calpain and their inhibitor calpastatin (Goll, Thompson, Li, Wei and Cong, 2003). Both calpains need calcium for their proteolytic activation. Calcium is also required for the interaction between ubiquitous calpains and their inhibitor calpastatin (Cottin, Vidalenc and Duscastaing, 1981). At the same time, m- and µ-calpain can autolyze in the presence of calcium (Suzuki, Tsuji, Ishiura, Kimura, Kubota and Imahori, 1981a). Autolysis can decrease the calcium 3 concentration required for proteolytic activation of m- and µ-calpain (Suzuki, Tsuji, Kubota, Kimura and Imahori, 1981b). However, complete autolysis can destroy their integral structure and thus lead to loss of their proteolytic activity (Elce, Hegadorn and Arthur, 1997). In postmortem muscle, autolysis of µ-calpain has been shown to be parallel to its proteolytic activation (Saido et al., 1994; DeMartino, Huff and Croall, 1986). Thus, the extent
Load more

Recommended publications
  • Discovery of Endoplasmic Reticulum Calcium Stabilizers to Rescue ER-Stressed Podocytes in Nephrotic Syndrome
    Discovery of endoplasmic reticulum calcium stabilizers to rescue ER-stressed podocytes in nephrotic syndrome Sun-Ji Parka, Yeawon Kima, Shyh-Ming Yangb, Mark J. Hendersonb, Wei Yangc, Maria Lindahld, Fumihiko Uranoe, and Ying Maggie Chena,1 aDivision of Nephrology, Department of Medicine, Washington University School of Medicine, St. Louis, MO 63110; bNational Center for Advancing Translational Sciences, National Institutes of Health, Rockville, MD 20850; cDepartment of Genetics, Washington University School of Medicine, St. Louis, MO 63110; dInstitute of Biotechnology, University of Helsinki, Helsinki, Finland 00014; and eDivision of Endocrinology, Metabolism, and Lipid Research, Department of Medicine, Washington University School of Medicine, St. Louis, MO 63110 Edited by Martin R. Pollak, Beth Israel Deaconess Medical Center, Brookline, MA, and approved May 28, 2019 (received for review August 16, 2018) Emerging evidence has established primary nephrotic syndrome activating transcription factor 6 (ATF6), which act as proximal (NS), including focal segmental glomerulosclerosis (FSGS), as a sensors of ER stress. ER stress activates these sensors by inducing primary podocytopathy. Despite the underlying importance of phosphorylation and homodimerization of IRE1α and PERK/ podocyte endoplasmic reticulum (ER) stress in the pathogenesis of eukaryotic initiation factor 2α (eIF2α), as well as relocalization of NS, no treatment currently targets the podocyte ER. In our mono- ATF6 to the Golgi, where it is cleaved by S1P/S2P proteases from genic podocyte ER stress-induced NS/FSGS mouse model, the 90 kDa to the active 50-kDa ATF6 (8), leading to activation of podocyte type 2 ryanodine receptor (RyR2)/calcium release channel their respective downstream transcription factors, spliced XBP1 on the ER was phosphorylated, resulting in ER calcium leak and (XBP1s), ATF4, and p50ATF6 (8–10).
    [Show full text]
  • CCN3 and Calcium Signaling Alain Lombet1, Nathalie Planque2, Anne-Marie Bleau2, Chang Long Li2 and Bernard Perbal*2
    Cell Communication and Signaling BioMed Central Review Open Access CCN3 and calcium signaling Alain Lombet1, Nathalie Planque2, Anne-Marie Bleau2, Chang Long Li2 and Bernard Perbal*2 Address: 1CNRS UMR 8078, Hôpital Marie Lannelongue, 133, Avenue de la Résistance 92350 Le PLESSIS-ROBINSON, France and 2Laboratoire d'Oncologie Virale et Moléculaire, Tour 54, Case 7048, Université Paris 7-D.Diderot, 2 Place Jussieu 75005 PARIS, France Email: Alain Lombet - [email protected]; Nathalie Planque - [email protected]; Anne-Marie Bleau - [email protected]; Chang Long Li - [email protected]; Bernard Perbal* - [email protected] * Corresponding author Published: 15 August 2003 Received: 26 June 2003 Accepted: 15 August 2003 Cell Communication and Signaling 2003, 1:1 This article is available from: http://www.biosignaling.com/content/1/1/1 © 2003 Lombet et al; licensee BioMed Central Ltd. This is an Open Access article: verbatim copying and redistribution of this article are permitted in all media for any purpose, provided this notice is preserved along with the article's original URL. Abstract The CCN family of genes consists presently of six members in human (CCN1-6) also known as Cyr61 (Cystein rich 61), CTGF (Connective Tissue Growth Factor), NOV (Nephroblastoma Overexpressed gene), WISP-1, 2 and 3 (Wnt-1 Induced Secreted Proteins). Results obtained over the past decade have indicated that CCN proteins are matricellular proteins, which are involved in the regulation of various cellular functions, such as proliferation, differentiation, survival, adhesion and migration. The CCN proteins have recently emerged as regulatory factors involved in both internal and external cell signaling.
    [Show full text]
  • Proteomics of Serum Extracellular Vesicles Identifies a Novel COPD Biomarker, Fibulin-3 from Elastic Fibres
    ORIGINAL ARTICLE COPD Proteomics of serum extracellular vesicles identifies a novel COPD biomarker, fibulin-3 from elastic fibres Taro Koba 1, Yoshito Takeda1, Ryohei Narumi2, Takashi Shiromizu2, Yosui Nojima 3, Mari Ito3, Muneyoshi Kuroyama1, Yu Futami1, Takayuki Takimoto4, Takanori Matsuki1, Ryuya Edahiro1, Satoshi Nojima5, Yoshitomo Hayama1, Kiyoharu Fukushima1, Haruhiko Hirata1, Shohei Koyama1, Kota Iwahori1, Izumi Nagatomo1, Mayumi Suzuki1, Yuya Shirai1, Teruaki Murakami1, Kaori Nakanishi 1, Takeshi Nakatani1, Yasuhiko Suga1, Kotaro Miyake1, Takayuki Shiroyama1, Hiroshi Kida 1, Takako Sasaki6, Koji Ueda7, Kenji Mizuguchi3, Jun Adachi2, Takeshi Tomonaga2 and Atsushi Kumanogoh1 ABSTRACT There is an unmet need for novel biomarkers in the diagnosis of multifactorial COPD. We applied next-generation proteomics to serum extracellular vesicles (EVs) to discover novel COPD biomarkers. EVs from 10 patients with COPD and six healthy controls were analysed by tandem mass tag-based non-targeted proteomics, and those from elastase-treated mouse models of emphysema were also analysed by non-targeted proteomics. For validation, EVs from 23 patients with COPD and 20 healthy controls were validated by targeted proteomics. Using non-targeted proteomics, we identified 406 proteins, 34 of which were significantly upregulated in patients with COPD. Of note, the EV protein signature from patients with COPD reflected inflammation and remodelling. We also identified 63 upregulated candidates from 1956 proteins by analysing EVs isolated from mouse models. Combining human and mouse biomarker candidates, we validated 45 proteins by targeted proteomics, selected reaction monitoring. Notably, levels of fibulin-3, tripeptidyl- peptidase 2, fibulin-1, and soluble scavenger receptor cysteine-rich domain-containing protein were significantly higher in patients with COPD.
    [Show full text]
  • Calpain Inhibitor and Ibudilast Rescue Β Cell Functions in a Cellular Model of Wolfram Syndrome
    Calpain inhibitor and ibudilast rescue β cell functions in a cellular model of Wolfram syndrome Lien D. Nguyena,b,1, Tom T. Fischera,c,1, Damien Abreud,e, Alfredo Arroyoa, Fumihiko Uranod,f, and Barbara E. Ehrlicha,b,2 aDepartment of Pharmacology, Yale University, New Haven, CT 06520; bInterdepartmental Neuroscience Program, Yale University, New Haven, CT 06520; cInstitute of Pharmacology, University of Heidelberg, 69117 Heidelberg, Germany; dDepartment of Medicine, Division of Endocrinology, Metabolism, and Lipid Research, Washington University School of Medicine, St. Louis, MO 63110; eMedical Scientist Training Program, Washington University School of Medicine, St. Louis, MO 63110; and fDepartment of Pathology and Immunology, Washington University School of Medicine, St. Louis, MO 63110 Edited by Melanie H. Cobb, University of Texas Southwestern Medical Center, Dallas, TX, and approved June 1, 2020 (received for review April 18, 2020) Wolfram syndrome is a rare multisystem disease characterized by diseases, such as Alzheimer’s disease (22), cancer progression childhood-onset diabetes mellitus and progressive neurodegener- (23), and diabetes mellitus (24, 25). ation. Most cases are attributed to pathogenic variants in a single Here we show that KO of WFS1 in rat insulinoma (INS1) cells gene, Wolfram syndrome 1 (WFS1). There currently is no disease- led to elevated resting cytosolic calcium, reduced stimulus- modifying treatment for Wolfram syndrome, as the molecular con- evoked calcium signaling, and, consequently, hypersusceptibility sequences of the loss of WFS1 remain elusive. Because diabetes to hyperglycemia and decreased glucose-stimulated insulin se- mellitus is the first diagnosed symptom of Wolfram syndrome, we cretion. Overexpression of WFS1 or WFS1’s interacting partner aimed to further examine the functions of WFS1 in pancreatic β neuronal calcium sensor-1 (NCS1) reversed the deficits observed cells in the context of hyperglycemia.
    [Show full text]
  • Ryanodine Receptor Regulation During Exercise Tobias Kohla, Gunnar Weningera, Ran Zalkb, Philip Eatonc, and Stephan E
    COMMENTARY COMMENTARY Intensity matters: Ryanodine receptor regulation during exercise Tobias Kohla, Gunnar Weningera, Ran Zalkb, Philip Eatonc, and Stephan E. Lehnarta,d,1 aHeart Research Center Göttingen, Clinic of Cardiology & Pulmonology, University Medical Center & Georg-August-University, 37099 Göttingen, Germany; bThe National Institute for Biotechnology in the Negev, Ben-Gurion University of the Negev, Beer-Sheva 84105, Israel; cKing’s College London, Cardiovascular Division, The British Heart Foundation Centre of Excellence, The Rayne Institute, St Thomas’ Hospital, London SE17EH, United Kingdom; and dGerman Centre for Cardiovascular Research site Götttingen, 37099 Göttingen, Germany Skeletal muscles provide fascinating examples which were correlated with depressed muscle how humans have evolved and exercise. While function (3). humans developed superior cognition, Previous studies determined that calpain metabolome evolution studies indicate an activation cleaves RyR1 channels into stably accelerated parallel decline in muscle ener- associated ∼375- and ∼150-kDa fragments getic capacity and strength (1). New forms of sustaining subconductance channel open gat- locomotion including exceptional endurance ing in vitro (6). Recombinant, N-terminally were adapted ∼2 million years ago (2). Now- truncated RyR1 channels reconstituted in lipid adays, we generally assume healthy aging and bilayers exhibited CICR-like functions (7). disease prevention depend on regular Notably, 24 h after HIIT exercise recreation- exercise. Contrasting with evolutionary ally active humans showed significant RyR1 adaptation, high-intensity interval training fragmentation (re)producing a ∼375-kDa (HIIT) represents an ultrashort form of ex- fragment in muscle biopsies similar to cal- ercise and a promising intervention for dis- pain-treated RyR1 (3, 6). Calpain is known ease prevention. However, the molecular to cleave RyR1 protomers at residues 1383– mechanisms are incompletely understood.
    [Show full text]
  • Meat Tenderness and the Calpain Proteolytic System in Longissimus Muscle of Young Bulls and Steers1
    Meat Tenderness and the Calpain Proteolytic System in Longissimus Muscle of Young Bulls and Steers1 J. B. Morgan**2, T. L. Wheeler?, M. Koohmaraie', J. W. Savell", and J. D. CrousetJ +Roman L. Hruska U.S. Meat Animal Research Center, ARS, USDA, Clay Center, NE 68933-0166 and *Department of Animal Science, Texas A&M University, College Station 77843-2471 ABSTRACT: The objectives of this study were to than meat from steers; however, sensory panelists examine the effects of castration on the calpain were unable (P > .05) to detect differences in proteinase system ( p-calpain, m-calpain, and cal- tenderness or other sensory traits between bulls and pastatin) activities and meat tenderness. Six each, steers. Activities of p- and m-calpain were not affected MARC 111 bulls and steers were slaughtered at (P > .05) by castration; however, calpastatin was approximately 12 mo of age. Longissimus muscle higher (P c .05) in muscles from the bull carcasses. samples were obtained for determining myofibril fragmentation index, Warner-Bratzler shear force, and Lower ( P c .05) myofibril fragmentation index values sensory panel evaluation at 1, 7, and 14 d postmortem, indicate that less proteolysis occurred in muscle from and p- and m-calpain and calpastatin activities at 24 h bulls than in muscle from steers during the first 7 d postmortem. Bulls produced leaner carcasses with postmortem. Greater calpastatin 24-h activity may be lower (Pc .05) quality grades than did steers. Meat associated with the increased shear force of meat from from bulls had higher ( P c .05) shear force values bulls.
    [Show full text]
  • Calmodulin-Androgen Receptor (AR) Interaction: Calcium- Dependent, Calpain-Mediated Breakdown of AR in Lncapprostatecancer Cells
    Research Article Calmodulin-Androgen Receptor (AR) Interaction: Calcium- Dependent, Calpain-Mediated Breakdown of AR in LNCaPProstateCancer Cells Ronald P. Pelley,1 Kannagi Chinnakannu,1 Shalini Murthy,1 Faith M. Strickland,2 Mani Menon,1 Q. Ping Dou,3 Evelyn R. Barrack,1 and G. Prem-Veer Reddy1,3 1Vattikuti Urology Institute and 2Department of Dermatology, Henry Ford Hospital; 3Karmanos Cancer Institute and Department of Pathology, Wayne State University School of Medicine, Detroit, Michigan Abstract Introduction Chemotherapy of prostate cancer targets androgen receptor Adenocarcinoma of the prostate is the most frequently (AR) by androgen ablation or antiandrogens, but unfortu- diagnosed cancer and second leading cause of cancer deaths in nately, it is not curative. Our attack on prostate cancer American men (1). Although androgen ablation is the most envisions the proteolytic elimination of AR, which requires a common therapy for disseminated prostate cancer, it is palliative fuller understanding of AR turnover. We showed previously in nature and most patients eventually succumb to hormone- that calmodulin (CaM) binds to AR with important con- refractory disease resistant to chemotherapy. Whether normal or sequences for AR stability and function. To examine the mutated, androgen receptor (AR) is required for growth in both involvement of Ca2+/CaM in the proteolytic breakdown of AR, androgen-sensitive and androgen-insensitive prostate cancer (2). we analyzed LNCaP cell extracts that bind to a CaM affinity Therefore, it is of paramount importance to dissect the various column for the presence of low molecular weight forms of AR ways in which AR is regulated to not simply inactivate but to (intact AR size, f114 kDa).
    [Show full text]
  • Calpain: a Molecule to Induce AIF-Mediated Necroptosis in RGC-5
    Shang et al. BMC Neuroscience 2014, 15:63 http://www.biomedcentral.com/1471-2202/15/63 RESEARCH ARTICLE Open Access Calpain: a molecule to induce AIF-mediated necroptosis in RGC-5 following elevated hydrostatic pressure Lei Shang1, Ju-Fang Huang1, Wei Ding1, Shuang Chen1, Li-Xiang Xue2, Ruo-Fei Ma3 and Kun Xiong1* Abstract Background: RIP3 (Receptor-interacting protein 3) pathway was mainly described as the molecular mechanism of necroptosis (programmed necrosis). But recently, non-RIP3 pathways were found to mediate necroptosis. We deliberate to investigate the effect of calpain, a molecule to induce necroptosis as reported (Cell Death Differ 19:245–256, 2012), in RGC-5 following elevated hydrostatic pressure. Results: First, we identified the existence of necroptosis of RGC-5 after insult by using necrostatin-1 (Nec-1, necroptosis inhibitor) detected by flow cytometry. Immunofluorescence staining and western blot were used to detect the expression of calpain. Western blot analysis was carried out to describe the truncated AIF (tAIF) expression with or without pretreatment of ALLN (calpain activity inhibitor). Following elevated hydrostatic pressure, necroptotic cells pretreated with or without ALLN was stained by Annexin V/PI, The activity of calpain was also examined to confirm the inhibition effect of ALLN. The results showed that after cell injury there was an upregulation of calpain expression. Upon adding ALLN, the calpain activity was inhibited, and tAIF production was reduced upon injury along with the decreased number of necroptosis cells. Conclusion: Our study found that calpain may induce necroptosis via tAIF-modulation in RGC-5 following elevated hydrostatic pressure. Keywords: Retinal ganglion cells-5, Calpain, Elevated hydrostatic pressure, tAIF, Necroptosis Background During cerebral hypoxic-ischemia, there’sanoverloadof Calpains are calcium-activated neutral protease, which intracellular calcium which activates calpains, as a result belongs to the family of cytosolic cysteine proteinases.
    [Show full text]
  • Calpain Activation Contributes to Hyperglycaemia-Induced Apoptosis in Cardiomyocytes
    Cardiovascular Research (2009) 84, 100–110 doi:10.1093/cvr/cvp189 Calpain activation contributes to hyperglycaemia-induced apoptosis in cardiomyocytes Ying Li1,2, Yanwen Li3, Qingping Feng1,2,4, Malcolm Arnold1,2,4, and Tianqing Peng1,2,5* 1Critical Illness Research, Lawson Health Research Institute, University of Western Ontario, VRL 6th Floor, A6-140, 800 Commissioners Road, London, ON, Canada N6A 4G5; 2Department of Medicine, University of Western Ontario, London, ON, Canada N6A 4G5; 3Department of Microbiology, Imperial College London, Flowers Building, Armstrong Road, London SW7 2AZ, UK; 4Department of Physiology and Pharmacology, University of Western Ontario, London, ON, Canada N6A 4G5; and 5Department of Pathology, University of Western Ontario, London, ON, Canada N6A 4G5 Received 22 June 2008; revised 11 May 2009; accepted 29 May 2009; online publish-ahead-of-print 8 June 2009 Time for primary review: 23 days KEYWORDS Aims Cardiomyocyte apoptosis contributes to cardiac complications of diabetes. The aim of this study Cardiomyocyte; was to investigate the role of calpain in cardiomyocyte apoptosis induced by hyperglycaemia. Apoptosis; Methods and results In cultured adult rat ventricular cardiomyocytes, high glucose (33 mM) increased Calpain; calpain activity and induced apoptosis, concomitant with the impairment of Naþ/Kþ ATPase activity. NADPH oxidase; These effects of high glucose on cardiomyocytes were abolished by various pharmacological calpain þ þ Na /K ATPase; inhibitors, knockdown of calpain-1 but not calpain-2 using siRNA, or over-expression of calpastatin, a Downloaded from High glucose specific endogenous calpain inhibitor. The effect of calpain inhibition on cardiomyocyte apoptosis was abrogated by ouabain, a selective inhibitor of Naþ/Kþ ATPase.
    [Show full text]
  • The Role of Phospholamban and Sarcolipin in Skeletal Muscle Disease
    The Role of Phospholamban and Sarcolipin in Skeletal Muscle Disease by Val Andrew M. Fajardo A thesis presented to the University of Waterloo in fulfillment of the thesis requirement for the degree of Doctor of Philosophy in Kinesiology Waterloo, Ontario, Canada, 2015 ©Val Andrew M. Fajardo AUTHOR’S DECLARATION This thesis consists of material all of which I authored or co-authored: see Statement of Contributions included in the thesis. This is a true copy of the thesis, including any required final revisions, as accepted by my examiners. I understand that my thesis may be made electronically available to the public. Val Andrew M. Fajardo ii Statement of Contributions Chapters 2, 3, and 4 of this thesis are presented as manuscripts published or currently in submission, and all manuscripts were a joint effort across several co-authors. The contributions from every author for each of the chapters are listed below. Chapter 2 – Thesis Study I: Phospholamban overexpression in mice causes a centronuclear myopathy-like phenotype Val A. Fajardo1§, Eric Bombardier1§, Elliott McMillan1, Khanh Tran1, Brennan J. Wadsworth1, Daniel Gamu1, Andrew Hopf, Chris Vigna1, Ian C. Smith1, Catherine Bellissimo1, Robin N. Michel2, Mark A. Tarnopolsky3,4,5, Joe Quadrilatero1, A. Russell Tupling1* From the 1Department of Kinesiology, University of Waterloo, Waterloo, Ontario, Canada, 2Department of Exercise Science, Concordia University, Montreal, Quebec, Canada, 3Departement of Kinesiology, 4Department of Pediatrics, and 5Department of Medicine, McMaster University, Hamilton, Ontario, Canada. A.R.T., V.A.F., and E.B. conceived the study idea. V.A.F. coordinated Western blot experiments and with K.T.
    [Show full text]
  • The Crystal Structure of Calcium-Free Human M-Calpain Suggests an Electrostatic Switch Mechanism for Activation by Calcium
    The crystal structure of calcium-free human m-calpain suggests an electrostatic switch mechanism for activation by calcium Stefan Strobl*†‡, Carlos Fernandez-Catalan*†, Marianne Braun†, Robert Huber†, Hajime Masumoto§, Kazuhiro Nakagawa§, Akihiro Irie§, Hiroyuki Sorimachi§¶, Gleb Bourenkowʈ, Hans Bartunikʈ, Koichi Suzuki§, and Wolfram Bode†** †Max-Planck-Institute of Biochemistry, Am Klopferspitz 18a, D 82 152 Planegg-Martinsried, Germany; §Institute of Molecular and Cellular Biosciences, University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan; and ʈArbeitsgruppe Proteindynamik Max-Planck-Gesellschaft Arbeitsgruppen für Strukfurelle Molekularbiologie, c͞o Deutsches Elektronen Synchrotron, D-22603 Hamburg, Germany Contributed by Robert Huber, November 16, 1999 Calpains (calcium-dependent cytoplasmic cysteine proteinases) are functioning of calpains, the structures of full-length calpain must implicated in processes such as cytoskeleton remodeling and signal to be known. We (10) and others (11) have communicated transduction. The 2.3-Å crystal structure of full-length het- crystals of full-length human and partially truncated rat m- -erodimeric [80-kDa (dI-dIV) ؉ 30-kDa (dV؉dVI)] human m-calpain calpain, respectively. In the following, we describe the funda crystallized in the absence of calcium reveals an oval disc-like mental properties of full-length human m-calpain and discuss the shape, with the papain-like catalytic domain dII and the two possible mechanisms of calcium activation. calmodulin-like domains dIV؉dVI occupying opposite poles, and the tumor necrosis factor ␣-like ␤-sandwich domain dIII and the Materials and Methods -N-terminal segments dI؉dV located between. Compared with Full-length human m-calpain containing an N-terminal GlyArg -papain, the two subdomains dIIa؉dIIb of the catalytic unit are ArgAspArgSer L-chain elongation was overexpressed in a bac rotated against one another by 50°, disrupting the active site and ulovirus expression system and was purified (12).
    [Show full text]
  • Phosphorylation-Dependent Interactome of Ryanodine Receptor Type 2 in the Heart
    proteomes Article Phosphorylation-Dependent Interactome of Ryanodine Receptor Type 2 in the Heart David Y. Chiang 1,† , Satadru Lahiri 2,3,† , Guoliang Wang 2, Jason Karch 2,3, Meng C. Wang 4,5,6, Sung Y. Jung 7 , Albert J. R. Heck 8,9, Arjen Scholten 8,9 and Xander H. T. Wehrens 2,3,10,11,12,13,* 1 Cardiovascular Division, Department of Medicine, Harvard Medical School, Brigham and Women’s Hospital, Boston, MA 02115, USA; [email protected] 2 Cardiovascular Research Institute, Department of Molecular Physiology & Biophysics, Baylor College of Medicine, Houston, TX 77030, USA; [email protected] (S.L.); [email protected] (G.W.); [email protected] (J.K.) 3 Department of Molecular Physiology & Biophysics, Baylor College of Medicine, Houston, TX 77030, USA 4 Huffington Center on Aging, Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX 77030, USA; [email protected] 5 Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX 77030, USA 6 Howard Hughes Medical Institute, Baylor College of Medicine, Houston, TX 77030, USA 7 Department of Biochemistry, Baylor College of Medicine, Houston, TX 77030, USA; [email protected] 8 Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Utrecht University, 3584 Utrecht, The Netherlands; [email protected] (A.J.R.H.); [email protected] (A.S.) 9 Netherlands Proteomics Centre, 3584 Utrecht, The Netherlands 10 Department of Medicine (Cardiology), Baylor College of Medicine, Houston, TX 77030, USA 11 Department of Neuroscience, Baylor College of Medicine, Houston, TX 77030, USA 12 Department of Pediatrics (Cardiology), Baylor College of Medicine, Houston, TX 77030, USA 13 Center for Space Medicine, Baylor College of Medicine, Houston, TX 77030, USA Citation: Chiang, D.Y.; Lahiri, S.; * Correspondence: [email protected]; Tel.: +1-713-798-4261 Wang, G.; Karch, J.; Wang, M.C.; Jung, † Equal contribution.
    [Show full text]