DOCSLIB.ORG

Mitochondrial Functionality in Male Fertility: from Spermatogenesis to Fertilization

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Mitochondrial Functionality in Male Fertility: from Spermatogenesis to Fertilization antioxidants Review Mitochondrial Functionality in Male Fertility: From Spermatogenesis to Fertilization Yoo-Jin Park and Myung-Geol Pang * Department of Animal Science & Technology and BET Research Institute, Chung-Ang University, Anseong 17546, Gyeonggi-do, Korea; [email protected] * Correspondence: [email protected] Abstract: Mitochondria are structurally and functionally distinct organelles that produce adenosine triphosphate (ATP) through oxidative phosphorylation (OXPHOS), to provide energy to spermatozoa. They can also produce reactive oxidation species (ROS). While a moderate concentration of ROS is critical for tyrosine phosphorylation in cholesterol efflux, sperm–egg interaction, and fertilization, excessive ROS generation is associated with male infertility. Moreover, mitochondria participate in diverse processes ranging from spermatogenesis to fertilization to regulate male fertility. This review aimed to summarize the roles of mitochondria in male fertility depending on the sperm developmen- tal stage (from male reproductive tract to female reproductive tract). Moreover, mitochondria are also involved in testosterone production, regulation of proton secretion into the lumen to maintain an acidic condition in the epididymis, and sperm DNA condensation during epididymal maturation. We also established the new signaling pathway using previous proteomic data associated with male fertility, to understand the overall role of mitochondria in male fertility. The pathway revealed that male infertility is associated with a loss of mitochondrial proteins in spermatozoa, which induces low sperm motility, reduces OXPHOS activity, and results in male infertility. Keywords: mitochondria; oxidative phosphorylation; spermatozoa; male infertility; testis; epididymis; capacitation; fertilization Citation: Park, Y.-J.; Pang, M.-G. Mitochondrial Functionality in Male Fertility: From Spermatogenesis to 1. Introduction Fertilization. Antioxidants 2021, 10, 98. https://doi.org/10.3390/ Mitochondria are structurally and functionally distinct organelles that have double antiox10010098 membranes and produce more than 90% of energy in eukaryotic cells, through oxidative phosphorylation (OXPHOS) [1,2]. The inner mitochondrial membrane, namely cristae, Received: 26 November 2020 is a highly folded and specialized compartment that increases the membrane surface for Accepted: 9 January 2021 storage of 94% of the OXPHOS complexes and adenosine triphosphate (ATP) synthase, and Published: 12 January 2021 85% of the total cytochrome c that is important for the OXPHOS system [3,4]. To produce ATP through OXPHOS, four respiratory enzyme complexes (complex I–IV) work in concert Publisher’s Note: MDPI stays neu- to create the electrochemical proton gradient in the mitochondrial inner membrane [5,6]. tral with regard to jurisdictional clai- The tricarboxylic acid (TCA) cycle, a series of enzymatic reactions that generate acetyl ms in published maps and institutio- coenzyme A (acetyl-CoA) by catabolism of carbohydrates, fats, and proteins, takes place in nal affiliations. the mitochondrial matrix. Acetyl Co-A is oxidized to generate carbon dioxide (CO2) and reducing agents, including nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FADH2). NADH and FADH2 work as fuel for the respiratory chain, through the transfer of electrons to the mitochondrial respiratory chain for the initiation of OXPHOS Copyright: © 2021 by the authors. Li- censee MDPI, Basel, Switzerland. (Figure1)[ 7]. The flux of electrons derived from the oxidation of substrates through various This article is an open access article redox carriers of the inner membrane electron transport chain (ETC) ultimately terminates distributed under the terms and con- in a four-electron reduction of molecular oxygen to water [8]. However, reactive oxidation − ditions of the Creative Commons At- species (ROS), such as superoxide (O2 ) and hydrogen peroxide (H2O2), are produced tribution (CC BY) license (https:// by the incomplete reduction of oxygen by an electron [9,10]. ROS work as important sec- creativecommons.org/licenses/by/ ondary messengers that regulate intracellular pathways through the oxidative activation of 4.0/). proteins, receptors, kinases, phosphatases, caspases, ion channels, and transcription factors Antioxidants 2021, 10, 98. https://doi.org/10.3390/antiox10010098 https://www.mdpi.com/journal/antioxidants Antioxidants 2021, 10, x FOR PEER REVIEW 2 of 27 Antioxidants 2021, 10, 98 2 of 24 [9,10]. ROS work as important secondary messengers that regulate intracellular pathways through the oxidative activation of proteins, receptors, kinases, phosphatases, caspases, ion channels, and transcription factors in normal conditions, and an imbalance between ROSin and normal the antioxidant conditions, defense and an imbalancesystem induces between oxidative ROS and stress the [11,12]. antioxidant Oxidative defense stress system frominduces ROS generation oxidative can stress cause [11, 12various]. Oxidative diseas stresses, including from ROS type generation II diabetes, can chronic cause various in- flammation,diseases, ischemia, including neurodegenerative type II diabetes, chronic disease, inflammation, and male infertility ischemia, [11,13–15]. neurodegenerative disease, and male infertility [11,13–15]. FigureFigure 1. Schematic 1. Schematic image imageof OXPHOS of OXPHOS and glycolysis and glycolysis in spermatozoa. in spermatozoa. Pyruvate is Pyruvate produced is by produced glycolysisby glycolysis and is then and converted is then converted to lactate in to th lactatee principal in the piece principal of spermatozoa piece of spermatozoaby lactate dehy- by lactate drogenase.dehydrogenase. Then, pyruvate Then, in pyruvate the principal in the piece principal is transported piece is transported inside into insidethe mitochondria into the mitochondria and it is usedand as it fuel is used for asTCA fuel cycles, for TCA which cycles, produce which the produce NADH. the OXPHOS NADH. OXPHOStake place take in mitochondria place in mitochondria of spermof midpiece. sperm midpiece. Oxidation Oxidation of NADH ofNADH in the elec in thetron electron transport transport chain pr chainoduce produce the ATP the molecules ATP molecules by OXPHOS.by OXPHOS. In spermatozoa,In spermatozoa, approximately approximately 80 mitochondr 80 mitochondriaia are present are present in the midpiece in the midpiece [16]. The [16]. mitochondria’sThe mitochondria’s roles are rolesnot limited are not to limitedproviding to providingenergy in sperm energy cells; in sperm they have cells; diverse they have functions,diverse including functions, the including production the productionof steroid hormones of steroid in hormones the testis, in and the testis,regulation and regula-of cell proliferationtion of cell proliferation as well as cell as death well asfor cell maintaining death for male maintaining fertility [17–20]. male fertility ROS generated [17–20]. ROS fromgenerated mitochondria from are mitochondria a double-edged are asword: double-edged they are crucial sword: for they tyrosine are crucial phosphoryla- for tyrosine tion phosphorylationin spermatozoa, cholesterol in spermatozoa, efflux, cholesterol and sperm–egg efflux, interaction, and sperm–egg but also interaction, play a role but in also pathologicalplay a role processes in pathological through processes oxidative through stress [21]. oxidative Approximately stress [21]. 30–80% Approximately of male 30–80%in- fertilityof male cases infertility are associated cases arewith associated ROS-mediated with ROS-mediated damage to spermatozoa damage to [22]. spermatozoa Exposure [22]. to ROSExposure induces to damage ROS induces of structural damage and of structural functional and components functional of components cells such as of proteins, cells such as membrane,proteins, and membrane, DNA in spermatozoa, and DNA in spermatozoa,which affects sperm which motility, affects sperm its ability motility, to penetrate its ability to oocytes,penetrate and embryonic oocytes, and development embryonic [22–24]. development A previous [22–24 study]. A previous reported study that lower reported mi- that tochondriallower mitochondrial respiratory activities respiratory is closely activities associated is closely with associated not only with the not reduced only the sperm reduced motility,sperm i.e., motility, asthenozoospermia, i.e., asthenozoospermia, but also lower but also viability lower and viability concentration and concentration of spermato- of sper- zoa matozoa[25]. Moreover, [25]. Moreover, lower mitochondrial lower mitochondrial membrane membrane potential potentialdue to the due abnormal to the abnormal mor- phologymorphology of the axoneme of the axoneme is a major is acause major of cause low sperm of low motility sperm motilityand severe and asthenozoo- severe astheno- spermiazoospermia [26]. Recently, [26]. Recently, comparative comparative proteomi proteomicc studies studiesbetween between sperm samples sperm samples from in- from fertileinfertile patients patients with asthenozoospermia with asthenozoospermia and fe andrtile fertile men menrevealed revealed that thatproteins proteins associated associated withwith mitochondrial mitochondrial OXPHOS, OXPHOS, and andTCA TCA cycles, cycles, and andmetabolism metabolism of pyruvate of pyruvate are downreg- are downreg- ulatedulated in spermatozoa in spermatozoa from from asthenozoospermia asthenozoospermia patients patients [27,28]. [27, 28These]. These previous previous reports reports
Load more

Recommended publications
  • Expression and Localization of Myosin VI in Developing Mouse Spermatids
    Histochem Cell Biol DOI 10.1007/s00418-017-1579-z ORIGINAL PAPER Expression and localization of myosin VI in developing mouse spermatids Przemysław Zakrzewski1 · Robert Lenartowski2 · Maria Jolanta Re˛dowicz3 · Kathryn G. Miller4 · Marta Lenartowska1 Accepted: 4 May 2017 © The Author(s) 2017. This article is an open access publication Abstract Myosin VI (MVI) is a versatile actin-based and Sertoli cell actin hoops. Since this is the frst report of motor protein that has been implicated in a variety of dif- MVI expression and localization during mouse spermio- ferent cellular processes, including endo- and exocytic genesis and MVI partners in developing sperm have not yet vesicle traffcking, Golgi morphology, and actin structure been identifed, we discuss some probable roles for MVI stabilization. A role for MVI in crucial actin-based pro- in this process. During early stages, MVI is hypothesized cesses involved in sperm maturation was demonstrated in to play a role in Golgi morphology and function as well Drosophila. Because of the prominence and importance of as in actin dynamics regulation important for attachment actin structures in mammalian spermiogenesis, we inves- of developing acrosome to the nuclear envelope. Next, tigated whether MVI was associated with actin-mediated the protein might also play anchoring roles to help gener- maturation events in mammals. Both immunofuorescence ate forces needed for spermatid head elongation. Moreo- and ultrastructural analyses using immunogold labeling ver, association of MVI with actin that accumulates in showed that MVI was strongly linked with key structures the Sertoli cell ectoplasmic specialization and other actin involved in sperm development and maturation.
    [Show full text]
  • Dissert 1 Title
    GENETIC AND BIOCHEMICAL ANALYSIS OF ESSENTIAL ENZYMES IN TRIACYLGLYCEROL SYNTHESIS IN ARABIDOPSIS By KIMBERLY LYNN COTTON A dissertation submitted in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY WASHINGTON STATE UNIVERSITY Molecular Plant Sciences DECEMBER 2015 © Copyright by KIMBERLY LYNN COTTON, 2015 All Rights Reserved To the Faculty of Washington State University: The members of the Committee appointed to examine the dissertation of KIMBERLY LYNN COTTON find it satisfactory and recommend that it be accepted. ____________________________________________ John A. Browse, Ph.D., Chair _____________________________________________ Michael M. Neff, Ph.D. _____________________________________________ Thomas W. Okita, Ph.D. _____________________________________________ John J. Wyrick, Ph.D. ii ACKNOWLEDGMENTS To my PI John Browse, thank you for your support, guidance and patience over the years. To my committee, John Browse, Michael Neff, Thomas Okita, and John Wyrick, thank you for your insightful discussions and suggestions. To my collaborators Jay Shockey, Anushobha Regmi, Neil Adhikari, John Browse, and Phil Bates, thanks for the great work and great times. To Browse Lab members past and present, thanks for the help, guidance and wonderful memories. To the undergraduate students who have helped me over the years, thank you for all your help in making this project possible; and with a special thank you to Arlene, Tesa, Breana, and “no-longer-undergrads” David and Barbara Cotton for their particular dedication to this project. And finally, a huge thank you to my family and friends who have loved and supported me throughout this venture. Go Cougs! iii GENETIC AND BIOCHEMICAL ANALYSIS OF ESSENTIAL ENZYMES IN TRIACYLGLYCEROL SYNTHESIS IN ARABIDOPSIS Abstract by Kimberly Lynn Cotton, Ph.D.
    [Show full text]
  • 82870547.Pdf
    ORIGINAL RESEARCH published: 06 March 2017 doi: 10.3389/fpls.2017.00291 Integrated Transcriptome and Metabolic Analyses Reveals Novel Insights into Free Amino Acid Metabolism in Huangjinya Tea Cultivar Qunfeng Zhang 1, 2, Meiya Liu 1, 2 and Jianyun Ruan 1, 2* 1 Tea Research Institute, Chinese Academy of Agricultural Sciences, Hangzhou, China, 2 Key Laboratory for Plant Biology and Resource Application of Tea, The Ministry of Agriculture, Hangzhou, China The chlorotic tea variety Huangjinya, a natural mutant, contains enhanced levels of free amino acids in its leaves, which improves the drinking quality of its brewed tea. Consequently, this chlorotic mutant has a higher economic value than the non-chlorotic Edited by: varieties. However, the molecular mechanisms behind the increased levels of free amino Basil J. Nikolau, Iowa State University, USA acids in this mutant are mostly unknown, as are the possible effects of this mutation Reviewed by: on the overall metabolome and biosynthetic pathways in tea leaves. To gain further John A. Morgan, insight into the effects of chlorosis on the global metabolome and biosynthetic pathways Purdue University, USA Vinay Kumar, in this mutant, Huangjinya plants were grown under normal and reduced sunlight, Central University of Punjab, India resulting in chlorotic and non-chlorotic leaves, respectively; their leaves were analyzed *Correspondence: using transcriptomics as well as targeted and untargeted metabolomics. Approximately Jianyun Ruan 5,000 genes (8.5% of the total analyzed) and ca. 300 metabolites (14.5% of the total [email protected] detected) were significantly differentially regulated, thus indicating the occurrence of Specialty section: marked effects of light on the biosynthetic pathways in this mutant plant.
    [Show full text]
  • Identification of Candidate Substrates of the Leucine Rich Repeat Kinase 2 by Mass Spectrometry
    Identification of candidate substrates of the leucine rich repeat kinase 2 by mass spectrometry- based phosphoproteomics William Carl Edelman A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy University of Washington 2016 Reading Committee: Judit Villén, Chair Leo Pallanck Deborah Nickerson Program Authorized to Offer Degree: Department of Genome Sciences 2 © Copyright 2016 William Carl Edelman 3 University of Washington Abstract Identification of candidate substrates of the leucine rich repeat kinase 2 by mass spectrometry-based phosphoproteomics William Carl Edelman Chair of the Supervisory Committee: Assistant Professor, Judit Villén Department of Genome Sciences Mutations in the kinase domain of the leucine rich repeat kinase (LRRK2) have been implicated in heritable forms of Parkinson’s disease (PD). Specifically, a glycine to serine mutation (G2019S) has demonstrated hyperactive autophosphorylation, neuronal toxicity, and locomotor deficits in the fruit fly Drosophila melanogaster— all of which are related to its pathogenicity in PD. My dissertation focuses on identifying novel substrates of LRRK2 through analysis of proteome-wide changes in protein abundance as well as identifying changes in phosphorylation of proteins in vitro and in the in vivo fruit fly model. Using mass spectrometry, I provide quantitative information on thousands of proteins and phosphorylation sites. In vitro kinase assays on peptides derived from fly heads or a neuroblastoma cell line provide evidence for direct substrates of LRRK2, while the in vivo experiment in flies expressing LRRK2 identifies both direct and indirect phosphorylation substrates of the kinase. Herein, I present evidence for novel, LRRK2-mediated phosphorylation sites in the Drosophila melanogaster and the neuroblastoma models of PD.
    [Show full text]
  • ATP-Citrate Lyase Has an Essential Role in Cytosolic Acetyl-Coa Production in Arabidopsis Beth Leann Fatland Iowa State University
    Iowa State University Capstones, Theses and Retrospective Theses and Dissertations Dissertations 2002 ATP-citrate lyase has an essential role in cytosolic acetyl-CoA production in Arabidopsis Beth LeAnn Fatland Iowa State University Follow this and additional works at: https://lib.dr.iastate.edu/rtd Part of the Molecular Biology Commons, and the Plant Sciences Commons Recommended Citation Fatland, Beth LeAnn, "ATP-citrate lyase has an essential role in cytosolic acetyl-CoA production in Arabidopsis " (2002). Retrospective Theses and Dissertations. 1218. https://lib.dr.iastate.edu/rtd/1218 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 Retrospective Theses and Dissertations by an authorized administrator of Iowa State University Digital Repository. For more information, please contact [email protected]. ATP-citrate lyase has an essential role in cytosolic acetyl-CoA production in Arabidopsis by Beth LeAnn Fatland A dissertation submitted to the graduate faculty in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY Major: Plant Physiology Program of Study Committee: Eve Syrkin Wurtele (Major Professor) James Colbert Harry Homer Basil Nikolau Martin Spalding Iowa State University Ames, Iowa 2002 UMI Number: 3158393 INFORMATION TO USERS The quality of this reproduction is dependent upon the quality of the copy submitted. Broken or indistinct print, colored or poor quality illustrations and photographs, print bleed-through, substandard margins, and improper alignment can adversely affect reproduction. In the unlikely event that the author did not send a complete manuscript and there are missing pages, these will be noted.
    [Show full text]
  • C-Terminal HSP90 Inhibitors Block the HSP90:HIF-1Α Interaction and Inhibit the Cellular Hypoxic Response
    bioRxiv preprint doi: https://doi.org/10.1101/521989; this version posted January 24, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. C-terminal HSP90 Inhibitors Block the HSP90:HIF-1α Interaction and Inhibit the Cellular Hypoxic Response Nalin Katariaa, Bernadette Kerra, Samantha S. Zaiterb, Shelli McAlpineb and Kristina M Cooka† a. University of Sydney, Faculty of Medicine and Health, Charles Perkins Centre, Sydney, Australia. b. School of Chemistry, University of New South Wales, Sydney, Australia. † To whom correspondence should be addressed: Kristina M Cook, Charles Perkins Centre, University of Sydney, Sydney, NSW, Australia 2006; [email protected]; Tel: +61 286274858. Hypoxia Inducible Factor (HIF) is a transcription factor cancer cells known as a heat shock response (HSR)12. The activated by low oxygen, which is common in solid compounds also have poor selectivity for HSP9013,14. tumours. HIF controls the expression of genes involved in C-terminus inhibitors of HSP90 (SM molecules)11 act in a angiogenesis, chemotherapy resistance and metastasis. The selective manner, and in contrast to the N-terminus chaperone HSP90 (Heat Shock Protein 90) stabilizes the inhibitors, they do not induce a heat shock response13,15. subunit HIF-1α and prevents degradation. Previously Although the SM molecules block all co-chaperones that identified HSP90 inhibitors bind to the N-terminal pocket bind to the C-terminus of HSP90 and inhibit HSP90 of HSP90 which blocks binding to HIF-1α, and produces function16,17, there is no data discussing whether C- HIF-1α degradation.
    [Show full text]
  • A Computational Approach for Defining a Signature of Β-Cell Golgi Stress in Diabetes Mellitus
    Page 1 of 781 Diabetes A Computational Approach for Defining a Signature of β-Cell Golgi Stress in Diabetes Mellitus Robert N. Bone1,6,7, Olufunmilola Oyebamiji2, Sayali Talware2, Sharmila Selvaraj2, Preethi Krishnan3,6, Farooq Syed1,6,7, Huanmei Wu2, Carmella Evans-Molina 1,3,4,5,6,7,8* Departments of 1Pediatrics, 3Medicine, 4Anatomy, Cell Biology & Physiology, 5Biochemistry & Molecular Biology, the 6Center for Diabetes & Metabolic Diseases, and the 7Herman B. Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN 46202; 2Department of BioHealth Informatics, Indiana University-Purdue University Indianapolis, Indianapolis, IN, 46202; 8Roudebush VA Medical Center, Indianapolis, IN 46202. *Corresponding Author(s): Carmella Evans-Molina, MD, PhD ([email protected]) Indiana University School of Medicine, 635 Barnhill Drive, MS 2031A, Indianapolis, IN 46202, Telephone: (317) 274-4145, Fax (317) 274-4107 Running Title: Golgi Stress Response in Diabetes Word Count: 4358 Number of Figures: 6 Keywords: Golgi apparatus stress, Islets, β cell, Type 1 diabetes, Type 2 diabetes 1 Diabetes Publish Ahead of Print, published online August 20, 2020 Diabetes Page 2 of 781 ABSTRACT The Golgi apparatus (GA) is an important site of insulin processing and granule maturation, but whether GA organelle dysfunction and GA stress are present in the diabetic β-cell has not been tested. We utilized an informatics-based approach to develop a transcriptional signature of β-cell GA stress using existing RNA sequencing and microarray datasets generated using human islets from donors with diabetes and islets where type 1(T1D) and type 2 diabetes (T2D) had been modeled ex vivo. To narrow our results to GA-specific genes, we applied a filter set of 1,030 genes accepted as GA associated.
    [Show full text]
  • Electronic Supplementary Material (ESI) for Molecular Biosystems
    Electronic Supplementary Material (ESI) for Molecular BioSystems. This journal is © The Royal Society of Chemistry 2015 Table S5 Mass data of proteins identified with label free shotgun proteomics a #Alt. Proteins; b Scores; c #Peptides; d SC [%]; e RMS90 [ppm]; f Rank; g Median(Controls:GLP1); h #(Controls:GLP1); i CV [%](Controls:GLP1); l Median(Controls:Palmitate); m #(Controls:Palmitate) ; n CV [%](Controls:Palmitate); o Median(Controls:GLP1 + Palmitate); p #(Controls:GLP1 + Palmitate); q CV [%](Controls:GLP1 + Palmitate) MW OK Accession Protein pI a b c d e f g h i l m n o p q [kDa] 78 kDa glucose-regulated protein OS=Rattus 1672.6 true GRP78_RAT 72.3 4.9 1 22 39 1.48 1 1.16 6 20.53 1.18 9 9.56 1.24 10 13.52 norvegicus GN=Hspa5 PE=1 SV=1 (M:1672.6) Endoplasmin OS=Rattus norvegicus 1253.0 true ENPL_RAT 92.7 4.6 1 23 29 3.34 2 0.85 8 33.34 1.19 6 39.58 1.18 10 28.11 GN=Hsp90b1 PE=1 SV=2 (M:1253.0) Stress-70 protein, mitochondrial OS=Rattus 1138.8 true GRP75_RAT 73.8 5.9 1 17 28.7 1.18 3 1.21 5 7.62 0.78 1 1.1 8 8.04 norvegicus GN=Hspa9 PE=1 SV=3 (M:1138.8) Protein disulfide-isomerase A3 OS=Rattus 1035.4 true PDIA3_RAT 56.6 5.8 1 16 35.2 2.84 4 0.94 3 17.22 1.15 4 26.62 1.25 4 7.86 norvegicus GN=Pdia3 PE=1 SV=2 (M:1035.4) Aconitate hydratase, mitochondrial 983.8 true ACON_RAT OS=Rattus norvegicus GN=Aco2 PE=1 85.4 8.7 1 19 31.4 2.72 5 0.89 2 23.6 1.21 2 9.25 0.89 3 32.87 (M:983.8) SV=2 60 kDa heat shock protein, mitochondrial 906.9 true CH60_RAT OS=Rattus norvegicus GN=Hspd1 PE=1 60.9 5.8 1 13 32.5 3.26 6 1.05 5 13.15 1.01 2 32.54 1.01
    [Show full text]
  • Datasheet: VPA00673 Product Details
    Datasheet: VPA00673 Description: RABBIT ANTI ATP5D Specificity: ATP5D Format: Purified Product Type: PrecisionAb™ Polyclonal Isotype: Polyclonal IgG Quantity: 100 µl Product Details Applications This product has been reported to work in the following applications. This information is derived from testing within our laboratories, peer-reviewed publications or personal communications from the originators. Please refer to references indicated for further information. For general protocol recommendations, please visit www.bio-rad-antibodies.com/protocols. Yes No Not Determined Suggested Dilution Western Blotting 1/1000 PrecisionAb antibodies have been extensively validated for the western blot application. The antibody has been validated at the suggested dilution. Where this product has not been tested for use in a particular technique this does not necessarily exclude its use in such procedures. Further optimization may be required dependant on sample type. Target Species Human Species Cross Reacts with: Mouse Reactivity N.B. Antibody reactivity and working conditions may vary between species. Product Form Purified IgG - liquid Preparation Rabbit polyclonal antibody purified by affinity chromatography Buffer Solution Phosphate buffered saline Preservative 0.09% Sodium Azide Stabilisers 2% Sucrose Immunogen Synthetic peptide encompassing part of the middle region of human ATP5D External Database Links UniProt: P30049 Related reagents Entrez Gene: 513 ATP5D Related reagents Specificity Rabbit anti Human ATP5D antibody recognizes ATP synthase subunit delta, mitochondrial, also known as ATP synthase subunit delta mitochondrial, F-ATPase delta subunit, mitochondrial ATP Page 1 of 2 synthase complex delta-subunit precusor, or mitochondrial ATP synthase, delta subunit. ATP5D gene encodes a subunit of mitochondrial ATP synthase. Mitochondrial ATP synthase catalyzes ATP synthesis, utilizing an electrochemical gradient of protons across the inner membrane during oxidative phosphorylation.
    [Show full text]
  • Transcriptional Control of Tissue-Resident Memory T Cell Generation
    Transcriptional control of tissue-resident memory T cell generation Filip Cvetkovski Submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the Graduate School of Arts and Sciences COLUMBIA UNIVERSITY 2019 © 2019 Filip Cvetkovski All rights reserved ABSTRACT Transcriptional control of tissue-resident memory T cell generation Filip Cvetkovski Tissue-resident memory T cells (TRM) are a non-circulating subset of memory that are maintained at sites of pathogen entry and mediate optimal protection against reinfection. Lung TRM can be generated in response to respiratory infection or vaccination, however, the molecular pathways involved in CD4+TRM establishment have not been defined. Here, we performed transcriptional profiling of influenza-specific lung CD4+TRM following influenza infection to identify pathways implicated in CD4+TRM generation and homeostasis. Lung CD4+TRM displayed a unique transcriptional profile distinct from spleen memory, including up-regulation of a gene network induced by the transcription factor IRF4, a known regulator of effector T cell differentiation. In addition, the gene expression profile of lung CD4+TRM was enriched in gene sets previously described in tissue-resident regulatory T cells. Up-regulation of immunomodulatory molecules such as CTLA-4, PD-1, and ICOS, suggested a potential regulatory role for CD4+TRM in tissues. Using loss-of-function genetic experiments in mice, we demonstrate that IRF4 is required for the generation of lung-localized pathogen-specific effector CD4+T cells during acute influenza infection. Influenza-specific IRF4−/− T cells failed to fully express CD44, and maintained high levels of CD62L compared to wild type, suggesting a defect in complete differentiation into lung-tropic effector T cells.
    [Show full text]
  • SUPPLEMENTARY TABLES and FIGURE LEGENDS Supplementary
    SUPPLEMENTARY TABLES AND FIGURE LEGENDS Supplementary Figure 1. Quantitation of MYC levels in vivo and in vitro. a) MYC levels in cell lines 6814, 6816, 5720, 966, and 6780 (corresponding to first half of Figure 1a in main text). MYC is normalized to tubulin. b) MYC quantitations (normalized to tubulin) for cell lines Daudi, Raji, Jujoye, KRA, KRB, GM, and 6780 corresponding to second half of Figure 1a. c) In vivo MYC quantitations, for mice treated with 0-0.5 ug/ml doxycycline in their drinking water. MYC is normalized to tubulin. d) Quantitation of changing MYC levels during in vitro titration, normalized to tubulin. e) Levels of Odc (normalized to tubulin) follow MYC levels in titration series. Supplementary Figure 2. Evaluation of doxycycline concentration in the plasma of mice treated with doxycycline in their drinking water. Luciferase expressing CHO cells (Tet- off) (Clonethech Inc) that is responsive to doxycycline by turning off luciferase expression was treated with different concentrations of doxycycline in culture. A standard curve (blue line) correlating luciferase activity (y-axis) with treatment of doxycycline (x- axis) was generated for the CHO cell in culture. Plasma from mice treated with different concentrations of doxycycline in their drinking water was separated and added to the media of the CHO cells. Luciferase activity was measured and plotted on the standard curve (see legend box). The actual concentration of doxycycline in the plasma was extrapolated for the luciferase activity measured. The doxycycline concentration 0.2 ng/ml measured in the plasma of mice correlates with 0.05 μg/ml doxycycline treatment in the drinking water of mice, the in vivo threshold for tumor regression.
    [Show full text]
  • Construction of Prognostic Microrna Signature for Human Invasive Breast Cancer by Integrated Analysis
    Journal name: OncoTargets and Therapy Article Designation: Original Research Year: 2019 Volume: 12 OncoTargets and Therapy Dovepress Running head verso: Shi et al Running head recto: Shi et al open access to scientific and medical research DOI: 189265 Open Access Full Text Article ORIGINAL RESEARCH Construction of prognostic microRNA signature for human invasive breast cancer by integrated analysis This article was published in the following Dove Medical Press journal: OncoTargets and Therapy Wei Shi* Background: Despite the advances in early detection and treatment methods, breast cancer Fang Dong* still has a high mortality rate, even in those patients predicted to have a good prognosis. The Yujia Jiang purpose of this study is to identify a microRNA signature that could better predict prognosis in Linlin Lu breast cancer and add new insights to the current classification criteria. Changwen Wang Materials and methods: We downloaded microRNA sequencing data along with correspond- Jie Tan ing clinicopathological data from The Cancer Genome Atlas (TCGA). Of 1,098 breast cancer patients identified, 253 patients with fully characterized microRNA profiles were selected for Wen Yang analysis. A three-microRNA signature was generated in the training set. Subsequently, the per- Hui Guo formance of the signature was confirmed in a validation set. After construction of the signature, Jie Ming* we conducted additional experiments, including flow cytometry and the Cell Counting Kit-8 Tao Huang* assay, to illustrate the correlation of this microRNA signature with breast cancer cell cycle, Department of Breast and Thyroid apoptosis, and proliferation. Surgery, Union Hospital, Tongji Results: Three microRNAs (hsa-mir-31, hsa-mir-16-2, and hsa-mir-484) were identified to Medical College, Huazhong University of Science and Technology, Wuhan be significantly and independently correlated with patient prognosis, and performed with good 430022, China stability.
    [Show full text]