DOCSLIB.ORG

A Pex13 Knockout in Germ Cells Induces A

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
A Pex13 Knockout in Germ Cells Induces A A PEX13 KNOCKOUT IN GERM CELLS INDUCES A SPERMATOGENIC ARREST INAUGURAL DISSERTATION to obtain the academic degree D o c t o r r e r u m n a t u r a l i u m (Dr. rer. nat.) by ANN-KRISTIN BRAUNS Submitted to the Faculty of Mathematics, Informatics and Natural Sciences of the University of Hamburg Realised at the Department of Anatomy and Experimental Morphology of the University Medical Centre Hamburg-Eppendorf July 2017 in Hamburg First Supervisor and Committee Member: Prof. Dr. med Georg Lüers Second Supervisor and Committee Member: Prof. Dr. Thorsten Burmester Committee Members: Prof. Dr. med. Udo Schumacher Prof. Dr. Jörg Ganzhorn Dr. Sabine Lüthje Dr. Daniel Wicklein Date of the oral examination: 15th December 2017 DECLARATION ON OATH “I hereby declare, on oath, that I have written the present dissertation by my own and have not used other than the acknowledged resources and aids. I have appropriately acknowledged and referenced all text passages that are derived literally from or are based on the content of published or unpublished work of others, and all information that relates to verbal communications.” Hamburg, 17th July 2017 Ann-Kristin Brauns CONTENT LIST OF ABBREVIATIONS ................................................................................................................ I ABSTRACT ......................................................................................................................................... IV 1 INTRODUCTION ......................................................................................................................... 1 1.1 Peroxisomes ............................................................................................................................ 2 1.1.1 Peroxisome biogenesis .................................................................................................... 3 1.1.2 Peroxisomal protein synthesis and their import .............................................................. 4 1.1.3 Peroxisomal function ....................................................................................................... 4 1.1.3.1 Metabolite transport .................................................................................................... 5 1.1.3.2 Fatty acids (FA) ........................................................................................................... 5 1.1.3.3 Peroxisomal β-oxidation ............................................................................................. 6 1.1.3.4 Reactive oxygen species (ROS) .................................................................................. 7 1.1.4 Peroxisomal disorders ..................................................................................................... 8 1.1.4.1 Peroxisomal biogenesis deficiencies (PBD) ................................................................ 8 1.1.4.2 Peroxisomal single-enzyme/ transporter deficiencies ................................................. 9 1.1.5 Peroxisomal dysfunction and male fertility ..................................................................... 9 1.2 Infertility ............................................................................................................................... 10 1.3 The male reproductive system ............................................................................................... 11 1.3.1 Spermatogenesis ............................................................................................................ 12 1.3.1.1 Germ cell differentiation and spermiogenesis ........................................................... 12 1.3.1.2 Endocrine regulation via the hypothalamic-pituitary-gonadal axis ........................... 15 1.3.2 The Sertoli cell .............................................................................................................. 16 1.3.2.1 Blood-testis barrier (BTB) ........................................................................................ 17 1.3.2.2 Phagocytosis of residual bodies ................................................................................ 18 1.3.3 Lipid composition in all cells of the seminiferous tubule ............................................. 18 1.4 Mouse models of male infertility .......................................................................................... 19 1.5 Objectives .............................................................................................................................. 19 2 MATERIALS AND METHODS................................................................................................ 21 2.1 Mouse strains used for the study ........................................................................................... 21 2.1.1 C57Bl/6J mice ............................................................................................................... 21 2.1.2 GFP-PTS1 transgenic mice ........................................................................................... 21 2.1.3 Pex13loxP transgenic mice ........................................................................................... 22 2.1.4 Stra8-Cre transgenic mice ............................................................................................. 22 2.1.5 Prm-Cre transgenic mice ............................................................................................... 22 2.2 Backcrossing of mouse strains .............................................................................................. 22 2.2.1 Breeding strategy to generate Stra8-Cre or Prm-Cre mediated Pex13 KO mice .......... 23 2.3 Genotyping with polymerase chain reaction (PCR) .............................................................. 24 2.4 Laser-capture microdissection (LCM) .................................................................................. 26 2.5 Morphological analyses ......................................................................................................... 26 2.5.1 Preparation of testis biopsies ......................................................................................... 26 2.5.2 Fixation and processing of testes for cryopreservation ................................................. 27 2.5.3 Fixation and processing of tissue for electron microscopy ........................................... 27 2.5.4 Haematoxylin and eosin staining (HE) ......................................................................... 27 2.5.5 Immunohistochemistry (IHC) ....................................................................................... 28 2.5.6 Immunofluorescence (IF) .............................................................................................. 28 2.5.7 Oil Red O staining (ORO) ............................................................................................. 29 2.5.8 TUNEL assay ................................................................................................................ 29 2.5.9 Evans Blue..................................................................................................................... 29 2.6 Preparation of a testicular single cell suspension .................................................................. 30 2.6.1 Separation of cells by velocity sedimentation ............................................................... 30 2.6.2 Flow cytometric analysis and cell sorting ..................................................................... 32 2.6.2.1 Preparation of FACS purified cells for electron microscopy .................................... 32 2.6.2.2 RNA isolation and cDNA synthesis of purified fixed germ cells ............................. 32 2.6.2.3 QRT-PCR and primers used to characterize purified germ cells of WT mice .......... 32 2.6.3 Counterflow centrifugal elutriation (CCE) .................................................................... 34 2.6.4 Cytospin and morphological analyses of sorted cells by fluorescent microscopy ........ 34 2.7 Total RNA isolation from testis biopsies .............................................................................. 34 2.7.1 QRT-PCR to characterize the peroxisomal compartment in testes biopsies ................. 35 2.7.2 QRT-PCR and primers used to characterize the BTB and fatty acid synthesis in testes biopsies ....................................................................................................................................... 36 2.8 Testes biopsies for lipid analysis ........................................................................................... 37 2.8.1 Gas chromatography-mass spectrometry (GC-MS) ...................................................... 37 2.9 Serum collection for steroid measurements .......................................................................... 38 2.9.1 ELISA assay .................................................................................................................. 38 2.10 Primary and secondary antibodies ......................................................................................... 39 2.11 Buffer solutions ..................................................................................................................... 41 2.12 Reagents, chemicals and kits ................................................................................................. 42 2.13 Equipment and materials ....................................................................................................... 44 3 RESULTS....................................................................................................................................
Load more

Recommended publications
  • PEX2 Is the E3 Ubiquitin Ligase Required for Pexophagy During Starvation
    JCB: Article PEX2 is the E3 ubiquitin ligase required for pexophagy during starvation Graeme Sargent,1,6 Tim van Zutphen,7 Tatiana Shatseva,6 Ling Zhang,3 Valeria Di Giovanni,3 Robert Bandsma,2,3,4,5 and Peter Kijun Kim1,6 1Cell Biology Department, 2Department of Paediatric Laboratory Medicine, 3Physiology and Experimental Medicine Program, Research Institute, 4Division of Gastroenterology, Hepatology and Nutrition, and 5Centre for Global Child Health, The Hospital for Sick Children, Toronto, ON M5G 1X8, Canada 6Biochemistry Department, University of Toronto, Toronto, ON M5S 1A8, Canada 7Department of Pediatrics, Center for Liver, Digestive and Metabolic Diseases, University of Groningen, University Medical Center Groningen, 9700 AD Groningen, Netherlands Peroxisomes are metabolic organelles necessary for anabolic and catabolic lipid reactions whose numbers are highly dynamic based on the metabolic need of the cells. One mechanism to regulate peroxisome numbers is through an auto- phagic process called pexophagy. In mammalian cells, ubiquitination of peroxisomal membrane proteins signals pexo- phagy; however, the E3 ligase responsible for mediating ubiquitination is not known. Here, we report that the peroxisomal E3 ubiquitin ligase peroxin 2 (PEX2) is the causative agent for mammalian pexophagy. Expression of PEX2 leads to Downloaded from gross ubiquitination of peroxisomes and degradation of peroxisomes in an NBR1-dependent autophagic process. We identify PEX5 and PMP70 as substrates of PEX2 that are ubiquitinated during amino acid starvation. We also find that PEX2 expression is up-regulated during both amino acid starvation and rapamycin treatment, suggesting that the mTORC1 pathway regulates pexophagy by regulating PEX2 expression levels. Finally, we validate our findings in vivo using an animal model.
    [Show full text]
  • Peroxisomal Disorders and Their Mouse Models Point to Essential Roles of Peroxisomes for Retinal Integrity
    International Journal of Molecular Sciences Review Peroxisomal Disorders and Their Mouse Models Point to Essential Roles of Peroxisomes for Retinal Integrity Yannick Das, Daniëlle Swinkels and Myriam Baes * Lab of Cell Metabolism, Department of Pharmaceutical and Pharmacological Sciences, KU Leuven, 3000 Leuven, Belgium; [email protected] (Y.D.); [email protected] (D.S.) * Correspondence: [email protected] Abstract: Peroxisomes are multifunctional organelles, well known for their role in cellular lipid homeostasis. Their importance is highlighted by the life-threatening diseases caused by peroxisomal dysfunction. Importantly, most patients suffering from peroxisomal biogenesis disorders, even those with a milder disease course, present with a number of ocular symptoms, including retinopathy. Patients with a selective defect in either peroxisomal α- or β-oxidation or ether lipid synthesis also suffer from vision problems. In this review, we thoroughly discuss the ophthalmological pathology in peroxisomal disorder patients and, where possible, the corresponding animal models, with a special emphasis on the retina. In addition, we attempt to link the observed retinal phenotype to the underlying biochemical alterations. It appears that the retinal pathology is highly variable and the lack of histopathological descriptions in patients hampers the translation of the findings in the mouse models. Furthermore, it becomes clear that there are still large gaps in the current knowledge on the contribution of the different metabolic disturbances to the retinopathy, but branched chain fatty acid accumulation and impaired retinal PUFA homeostasis are likely important factors. Citation: Das, Y.; Swinkels, D.; Baes, Keywords: peroxisome; Zellweger; metabolism; fatty acid; retina M. Peroxisomal Disorders and Their Mouse Models Point to Essential Roles of Peroxisomes for Retinal Integrity.
    [Show full text]
  • Supplementary Table S4. FGA Co-Expressed Gene List in LUAD
    Supplementary Table S4. FGA co-expressed gene list in LUAD tumors Symbol R Locus Description FGG 0.919 4q28 fibrinogen gamma chain FGL1 0.635 8p22 fibrinogen-like 1 SLC7A2 0.536 8p22 solute carrier family 7 (cationic amino acid transporter, y+ system), member 2 DUSP4 0.521 8p12-p11 dual specificity phosphatase 4 HAL 0.51 12q22-q24.1histidine ammonia-lyase PDE4D 0.499 5q12 phosphodiesterase 4D, cAMP-specific FURIN 0.497 15q26.1 furin (paired basic amino acid cleaving enzyme) CPS1 0.49 2q35 carbamoyl-phosphate synthase 1, mitochondrial TESC 0.478 12q24.22 tescalcin INHA 0.465 2q35 inhibin, alpha S100P 0.461 4p16 S100 calcium binding protein P VPS37A 0.447 8p22 vacuolar protein sorting 37 homolog A (S. cerevisiae) SLC16A14 0.447 2q36.3 solute carrier family 16, member 14 PPARGC1A 0.443 4p15.1 peroxisome proliferator-activated receptor gamma, coactivator 1 alpha SIK1 0.435 21q22.3 salt-inducible kinase 1 IRS2 0.434 13q34 insulin receptor substrate 2 RND1 0.433 12q12 Rho family GTPase 1 HGD 0.433 3q13.33 homogentisate 1,2-dioxygenase PTP4A1 0.432 6q12 protein tyrosine phosphatase type IVA, member 1 C8orf4 0.428 8p11.2 chromosome 8 open reading frame 4 DDC 0.427 7p12.2 dopa decarboxylase (aromatic L-amino acid decarboxylase) TACC2 0.427 10q26 transforming, acidic coiled-coil containing protein 2 MUC13 0.422 3q21.2 mucin 13, cell surface associated C5 0.412 9q33-q34 complement component 5 NR4A2 0.412 2q22-q23 nuclear receptor subfamily 4, group A, member 2 EYS 0.411 6q12 eyes shut homolog (Drosophila) GPX2 0.406 14q24.1 glutathione peroxidase
    [Show full text]
  • E3 Ubiquitin Ligase SP1 Regulates Peroxisome Biogenesis in Arabidopsis
    E3 ubiquitin ligase SP1 regulates peroxisome PNAS PLUS biogenesis in Arabidopsis Ronghui Pana, John Satkovicha, and Jianping Hua,b,1 aDepartment of Energy Plant Research Laboratory, Michigan State University, East Lansing, MI 48824; and bPlant Biology Department, Michigan State University, East Lansing, MI 48824 Edited by Natasha V. Raikhel, Center for Plant Cell Biology, Riverside, CA, and approved September 30, 2016 (received for review August 17, 2016) Peroxisomes are ubiquitous eukaryotic organelles that play pivotal signal type 1) and N-terminal PTS2 sequences, respectively roles in a suite of metabolic processes and often act coordinately (15, 16). In Arabidopsis, PEX5 is also required for PTS2 protein with other organelles, such as chloroplasts and mitochondria. Peroxi- import (16). Two membrane proteins, PEX13 and PEX14, form somes import proteins to the peroxisome matrix by peroxins (PEX the docking site for PEX5 and PEX7 (17, 18). After receptor proteins), but how the function of the PEX proteins is regulated is docking, cargo proteins translocate into the matrix before re- poorly understood. In this study, we identified the Arabidopsis RING ceptors are recycled to the cytosol (19–21). These processes re- (really interesting new gene) type E3 ubiquitin ligase SP1 [suppressor quire the RING (really interesting new gene)-finger peroxins of plastid protein import locus 1 (ppi1) 1] as a peroxisome membrane PEX2,PEX10,andPEX12(22–25), the ATPases PEX1 and protein with a regulatory role in peroxisome protein import. SP1 PEX6 and their membrane tether APEM9 (aberrant peroxisome interacts physically with the two components of the peroxisome morphology 9) and the ubiquitin-conjugating enzyme PEX4 and protein docking complex PEX13–PEX14 and the (RING)-finger per- its membrane anchor PEX22 (26–29).
    [Show full text]
  • 2014A Veenhuis Microbial Cell
    Microreview www.microbialcell.com De novo peroxisome biogenesis revisited Marten Veenhuis and Ida J. van der Klei* Molecular Cell Biology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, The Netherlands. * Corresponding Author: Ida J. van der Klei, P.O. Box 11103; 9700 CC Groningen, The Netherlands; Tel: +31 50 363 2179/2400; Fax: +31 50 363 2348; E-mail: [email protected] We describe an alternative peroxisome formation made in pex19 cells of both species although the pex19 pathway in yeast pex3 and pex19 cells, which relies on vesicles differed from those present in pex3 cells in that the existence of small peroxisomal remnants that are they contained, besides Pex13, Pex14 and Pex8, also Pex3. present in these cells. This groundbreaking result chal- Pex13 and Pex14 are key components of the matrix protein lenges current models prescribing that peroxisomes receptor docking complex. In the membrane remnants derive de novo from the ER. Our data also has major small amounts of matrix protein were present suggesting implications for the sorting pathway of specific peroxi- that Pex13 and Pex14 were correctly inserted and func- somal membrane proteins (PMPs). We propose a novel tional as receptor docking site. The low matrix content may be explained in that the proteins of the receptor recycling sorting pathway for the PMPs Pex13 and Pex14 that is system (including the RING finger proteins) were not pre- independent of the known Pex3/Pex19 machinery. sent on these structures, thereby preventing recycling of the PTS1 receptor Pex5. Indeed, Pex5 was found associated Peroxisomes are crucial, multifunctional organelles the with the vesicles, whereas other PMPs, e.g.
    [Show full text]
  • Microduplication in the 2P16.1P15 Chromosomal Region Linked To
    Lovrecic et al. Molecular Cytogenetics (2018) 11:39 https://doi.org/10.1186/s13039-018-0388-y CASE REPORT Open Access Microduplication in the 2p16.1p15 chromosomal region linked to developmental delay and intellectual disability Luca Lovrecic1* , Chiara Gnan2, Federica Baldan3, Alessandra Franzoni2, Sara Bertok4, Giuseppe Damante5, Bertrand Isidor6 and Borut Peterlin1 Abstract Background: Several patients with the 2p16.1p15 microdeletion syndrome have been reported. However, microduplication in the 2p16.1p15 chromosomal region has only been reported in one case, and milder clinical features were present compared to those attributed to 2p16.1p15 microdeletion syndrome. Some additional cases were deposited in DECIPHER database. Case presentation: In this report we describe four further cases of 2p16.1p15 microduplication in four unrelated probands. They presented with mild gross motor delay, delayed speech and language development, and mild dysmorphic features. In addition, two probands have macrocephaly and one a congenital heart anomaly. Newly described cases share several phenotype characteristics with those detailed in one previously reported microduplication case. Conclusion: The common features among patients are developmental delay, speech delay, mild to moderate intellectual disability and unspecific dysmorphic features. Two patients have bilateral clinodactyly of the 5th finger and two have bilateral 2nd-3rd toes syndactyly. Interestingly, as opposed to the deletion phenotype with some cases of microcephaly, 2 patients are reported
    [Show full text]
  • Supp Table 6.Pdf
    Supplementary Table 6. Processes associated to the 2037 SCL candidate target genes ID Symbol Entrez Gene Name Process NM_178114 AMIGO2 adhesion molecule with Ig-like domain 2 adhesion NM_033474 ARVCF armadillo repeat gene deletes in velocardiofacial syndrome adhesion NM_027060 BTBD9 BTB (POZ) domain containing 9 adhesion NM_001039149 CD226 CD226 molecule adhesion NM_010581 CD47 CD47 molecule adhesion NM_023370 CDH23 cadherin-like 23 adhesion NM_207298 CERCAM cerebral endothelial cell adhesion molecule adhesion NM_021719 CLDN15 claudin 15 adhesion NM_009902 CLDN3 claudin 3 adhesion NM_008779 CNTN3 contactin 3 (plasmacytoma associated) adhesion NM_015734 COL5A1 collagen, type V, alpha 1 adhesion NM_007803 CTTN cortactin adhesion NM_009142 CX3CL1 chemokine (C-X3-C motif) ligand 1 adhesion NM_031174 DSCAM Down syndrome cell adhesion molecule adhesion NM_145158 EMILIN2 elastin microfibril interfacer 2 adhesion NM_001081286 FAT1 FAT tumor suppressor homolog 1 (Drosophila) adhesion NM_001080814 FAT3 FAT tumor suppressor homolog 3 (Drosophila) adhesion NM_153795 FERMT3 fermitin family homolog 3 (Drosophila) adhesion NM_010494 ICAM2 intercellular adhesion molecule 2 adhesion NM_023892 ICAM4 (includes EG:3386) intercellular adhesion molecule 4 (Landsteiner-Wiener blood group)adhesion NM_001001979 MEGF10 multiple EGF-like-domains 10 adhesion NM_172522 MEGF11 multiple EGF-like-domains 11 adhesion NM_010739 MUC13 mucin 13, cell surface associated adhesion NM_013610 NINJ1 ninjurin 1 adhesion NM_016718 NINJ2 ninjurin 2 adhesion NM_172932 NLGN3 neuroligin
    [Show full text]
  • Perkinelmer Genomics to Request the Saliva Swab Collection Kit for Patients That Cannot Provide a Blood Sample As Whole Blood Is the Preferred Sample
    Zellweger Spectrum Disorder Panel Test Code D4611 Test Summary This panel analyzes 15 genes that have been associated with Zellweger Spectrum Disorders. Turn-Around-Time (TAT)* 3 - 5 weeks Acceptable Sample Types Whole Blood (EDTA) (Preferred sample type) DNA, Isolated Dried Blood Spots Saliva Acceptable Billing Types Self (patient) Payment Institutional Billing Commercial Insurance Indications for Testing This panel may be appropriate for individuals with signs and symptoms of a peroxisomal biogenesis disfuction and/or Zellweger syndrome spectrum disorder and/or a family history of these conditions. Test Description This panel analyzes 15 genes that have been associated with Zellweger Spectrum Disorders. Both sequencing and deletion/duplication (CNV) analysis will be performed on the coding regions of all genes included (unless otherwise marked). All analysis is performed utilizing Next Generation Sequencing (NGS) technology. CNV analysis is designed to detect the majority of deletions and duplications of three exons or greater in size. Smaller CNV events may also be detected and reported, but additional follow-up testing is recommended if a smaller CNV is suspected. All variants are classified according to ACMG guidelines. Condition Description Zellweger syndrome spectrum disorders include Zellweger syndrome (ZS), neonatal adrenoleukodystrophy (NALD), an intermediate form and infantile Refsum disease (IRD). Symptoms may include hypotonia, feeding problems, hearing and vision loss, seizures, distinctive facial characteristics and skeletal abnormalities. Zellweger spectrum disorder is estimated to occur in 1/50,000 individuals. Genes ACOX1, AMACR, HSD17B4, PEX1, PEX10, PEX12, PEX13, PEX14, PEX16, PEX19, PEX2, PEX26, PEX3, PEX5, PEX6 Test Methods and Limitations Sequencing is performed on genomic DNA using an Agilent targeted sequence capture method to enrich for the genes on this panel.
    [Show full text]
  • Genetic and Clinical Aspects of Zellweger Spectrum Patients with PEX1 Mutations H Rosewich, a Ohlenbusch, J Ga¨Rtner
    1of6 J Med Genet: first published as 10.1136/jmg.2005.033324 on 1 September 2005. Downloaded from ONLINE MUTATION REPORT Genetic and clinical aspects of Zellweger spectrum patients with PEX1 mutations H Rosewich, A Ohlenbusch, J Ga¨rtner ............................................................................................................................... J Med Genet 2005;42:e58 (http://www.jmedgenet.com/cgi/content/full/42/9/e58). doi: 10.1136/jmg.2005.033324 and can survive up to several decades. RCDP is clinically Objective: To analyse the PEX1 gene, the most common and genetically distinctive from the Zellweger syndrome cause for peroxisome biogenesis disorders (PBD), in a spectrum and includes classical RCDP as the prototype and consecutive series of patients with Zellweger spectrum. also milder variants. Patients with classical RCDP have Methods: Mutations were detected by different methods unique clinical symptoms including proximal shortening of including SSCP analyses as a screening technique on the the limbs (rhizomelia), cataracts, and profound psychomotor basis of genomic or cDNA, followed by direct sequencing of retardation. PCR fragments with an abnormal electrophoresis pattern. PBDs can be caused by defects in any of several processes Results: 33 patients were studied. Two common mutations, in organelle formation, including the synthesis of peroxisome c.2528GRA, G843D and c.2098_2098insT, I700YfsX42, membranes, the recognition of newly synthesised peroxiso- accounted for over 80% of all abnormal PEX1 alleles, mal matrix proteins, or any of the downstream steps in emphasising their diagnostic relevance. Most PEX1 mutations peroxisomal protein import. Progress over the last two were distributed over the two AAA cassettes with the two decades has led to the identification of 13 different human functional protein domains, D1 and D2, and the highly PEX genes involved in peroxisome biogenesis, explaining the conserved Walker motifs.
    [Show full text]
  • LRP1 Regulates Peroxisome Biogenesis and Cholesterol
    RESEARCH ARTICLE LRP1 regulates peroxisome biogenesis and cholesterol homeostasis in oligodendrocytes and is required for proper CNS myelin development and repair Jing-Ping Lin1, Yevgeniya A Mironova2, Peter Shrager3, Roman J Giger1,2,4,5* 1Department of Cell and Developmental Biology, University of Michigan School of Medicine, Ann Arbor, MI, United States; 2Cellular and Molecular Biology Graduate Program, University of Michigan Medical School, Ann Arbor, MI, United States; 3Department of Neuroscience, University of Rochester Medical Center, Rochester, NY, United States ; 4Department of Neurology, University of Michigan Medical School, Ann Arbor, MI, United States; 5Interdepartmental Neuroscience Graduate Program, University of Michigan Medical School, Ann Arbor, MI, United States Abstract Low-density lipoprotein receptor-related protein-1 (LRP1) is a large endocytic and signaling molecule broadly expressed by neurons and glia. In adult mice, global inducible (Lrp1flox/ flox;CAG-CreER) or oligodendrocyte (OL)-lineage specific ablation (Lrp1flox/flox;Pdgfra-CreER) of Lrp1 attenuates repair of damaged white matter. In oligodendrocyte progenitor cells (OPCs), Lrp1 is required for cholesterol homeostasis and differentiation into mature OLs. Lrp1-deficient OPC/ OLs show a strong increase in the sterol-regulatory element-binding protein-2 yet are unable to maintain normal cholesterol levels, suggesting more global metabolic deficits. Mechanistic studies revealed a decrease in peroxisomal biogenesis factor-2 and fewer peroxisomes in OL processes. / *For correspondence: Treatment of Lrp1À À OPCs with cholesterol or activation of peroxisome proliferator-activated [email protected] receptor-g with pioglitazone alone is not sufficient to promote differentiation; however, when combined, cholesterol and pioglitazone enhance OPC differentiation into mature OLs. Collectively, Competing interests: The our studies reveal a novel role for Lrp1 in peroxisome biogenesis, lipid homeostasis, and OPC authors declare that no competing interests exist.
    [Show full text]
  • Lysosomal Exocytosis Releases Pathogenic Α-Synuclein Species
    bioRxiv preprint doi: https://doi.org/10.1101/2021.04.10.439302; this version posted April 11, 2021. 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. Lysosomal Exocytosis Releases Pathogenic α-Synuclein Species from Neurons Ying Xue Xie1#, Nima N. Naseri2#, Jasmine Fels1, Parinati Kharel1, Yoonmi Na1, Jacqueline Burré1, Manu Sharma1* 1 Appel Institute for Alzheimer’s Research, and Brain & Mind Research Institute, Weill Cornell Medicine, New York, NY, USA. 2 Department of Chemistry, University of Pennsylvania, Philadelphia, PA, USA. # Equal contribution * Correspondence: [email protected] bioRxiv preprint doi: https://doi.org/10.1101/2021.04.10.439302; this version posted April 11, 2021. 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. SUMMARY Considerable evidence supports the release of pathogenic aggregates of the neuronal protein α-Synuclein (αSyn) into the extracellular space. While this release is proposed to instigate the neuron-to-neuron transmission and spread of αSyn pathology in synucleinopathies including Parkinson’s disease, the molecular-cellular mecha- nism(s) remain unclear. Here we show that pathogenic species of αSyn accumulate within neuronal lysosomes in mouse brains and primary neurons. We then find that neurons release these pathogenic αSyn species via SNARE- dependent lysosomal exocytosis; proposing a central mechanism for exocytosis of aggregated and degradation- resistant proteins from neurons. INTRODUCTION Cytoplasmic aggregates of the synaptic protein αSyn are a characteristic feature of multiple neurodegenerative diseases termed “synucleinopathies”, including Parkinson disease (PD).
    [Show full text]
  • Inheritest 500 PLUS
    Inheritest® 500 PLUS 525 genes Specimen ID: 00000000010 Container ID: H0651 Control ID: Acct #: LCA-BN Phone: SAMPLE REPORT, F-630049 Patient Details Specimen Details Physician Details DOB: 01/01/1991 Date Collected: 08/05/2019 12:00 (Local) Ordering: Age (yyy/mm/dd): 028/07/04 Date Received: 08/06/2019 Referring: Gender: Female Date Entered: 08/06/2019 ID: Patient ID: 00000000010 Date Reported: 08/21/2019 15:29 (Local) NPI: Ethnicity: Unknown Specimen Type: Blood Lab ID: MNEGA Indication: Carrier screening Genetic Counselor: None SUMMARY: POSITIVE POSITIVE RESULTS DISORDER (GENE) RESULTS INTERPRETATION Spinal muscular atrophy AT RISK AT RISK to be a silent carrier (2+0). For ethnic-specific risk (SMN1) 2 copies of SMN1; positive for revisions see Methods/Limitations. Genetic counseling is NMID: NM_000344 c.*3+80T>G SNP recommended. Risk: AT INCREASED RISK FOR AFFECTED PREGNANCY. See Additional Clinical Information. NEGATIVE RESULTS DISORDER (GENE) RESULTS INTERPRETATION Cystic fibrosis NEGATIVE This result reduces, but does not eliminate the risk to be a (CFTR) carrier. NMID: NM_000492 Risk: NOT at an increased risk for an affected pregnancy. Fragile X syndrome NEGATIVE: Not a carrier of a fragile X expansion. (FMR1) 29 and 36 repeats NMID: NM_002024 Risk: NOT at an increased risk for an affected pregnancy. ALL OTHER DISORDERS NEGATIVE This result reduces, but does not eliminate the risk to be a carrier. Risk: The individual is NOT at an increased risk for having a pregnancy that is affected with one of the disorders covered by this test. For partner's gene-specific risks, visit www.integratedgenetics.com.
    [Show full text]