Cellular Differentiation

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Int. J. Med. Sci. 2011, 8 http://www.medsci.org 88 IInntteerrnnaattiioonnaall JJoouurrnnaall ooff MMeeddiiccaall SScciieenncceess 2011; 8(2):88-96 © Ivyspring International Publisher. All rights reserved. Research Paper Parvovirus B19 Nonstructural Protein-Induced Damage of Cellular DNA and Resultant Apoptosis Brian D. Poole1,2, Violetta Kivovich1,3,4,5, Leona Gilbert4,5 and Stanley J. Naides1, 5,6  1. Huck Institute for Life Sciences, Pennsylvania State University College of Medicine/Milton S. Hershey Medical Center, Hershey, PA, USA 2. Current: Department of Microbiology and Molecular Biology, Brigham Young University, Provo, UT, USA 3. MD/PhD Program, Pennsylvania State University College of Medicine/Milton S. Hershey Medical Center, Hershey, PA, USA 4. Chemical Biology Division, Department of Biological and Environmental Science, University of Jyväskylä, Jyväskylä, Finland 5. Division of Rheumatology, Department of Medicine, Pennsylvania State University College of Medicine/Milton S. Hershey Medical Center, Hershey, PA, USA 6. Current: Quest Diagnostics Nichols Institute, San Juan Capistrano, CA, USA  Corresponding author: Stanley J. Naides, MD, Immunology, Quest Diagnostics Nichols Institute, 33608 Ortega Highway, San Juan Capistrano, CA 92690. Tel. 949 728-4578; fax 949 728-7852; email: stanley.j.naides@questdiagnostics.com Received: 2010.12.02; Accepted: 2011.01.13; Published: 2011.01.15 Abstract Parvovirus B19 is a widespread virus with diverse clinical presentations. The viral non-structural protein, NS1, binds to and cleaves the viral genome, and induces apoptosis when transfected into nonpermissive cells, such as hepatocytes. We hypothesized that the cyto-toxicity of NS1 in such cells results from chromosomal DNA damage caused by the DNA-nicking and DNA-attaching activities of NS1. Upon testing this hypothesis, we found that NS1 covalently binds to cellular DNA and is modified by PARP, an enzyme involved in repairing single-s t r a n d e d D N A n i c k s . W e f u r t h e r m o r e d i s c o v e r e d t h a t t h e D N A n i c k r e p a i r pathway initiated by poly(ADPribose)polymerase and the DNA repair pathways initiated by ATM/ATR are necessary for efficient apoptosis resulting from NS1 expression. Key words: Parvovirus B19, DNA damage and repair, fulminant liver failure, apoptosis, autoan -tibody, systemic lupus erythematosus Introduction Parvovirus B19 (B19) is a common virus with multiple clinical presentations. Infection in children is typically seen as erythema infectiosum, or fifth dis-ease (1), while adults often experience arthropathy lasting up to several months (2). Autoantibodies are often found subsequent to B19 infection, and are as-sociated with arthropathy (3-5). In patients with chronic hemolytic anemias, such as sickle cell disease or hereditary spherocytosis, the destruction of the e r y t h r o i d p r e c u r s o r p o o l b y B 1 9 l e a d s t o a p l a s t i c c r i s i s (6). B19 infection is implicated in hepatitis non-A-E acute fulminant liver failure (7-16). Although these are the best-described clinical illnesses caused by B19, the virus has been implicated in a wide spectrum of other illnesses (17). B19 infects a variety of cell types, but predomi-nantly replicates in erythroid precursors (18). Infec-tion of other cell types results in a limited, non-replicative state with overexpression of the viral nonstructural protein, NS1, and little expression of genes for the Cellular Differentiation Cellular Differentiation Bởi: OpenStaxCollege How does a complex organism such as a human develop from a single cell—a fertilized egg—into the vast array of cell types such as nerve cells, muscle cells, and epithelial cells that characterize the adult? Throughout development and adulthood, the process of cellular differentiation leads cells to assume their final morphology and physiology Differentiation is the process by which unspecialized cells become specialized to carry out distinct functions Stem Cells A stem cell is an unspecialized cell that can divide without limit as needed and can, under specific conditions, differentiate into specialized cells Stem cells are divided into several categories according to their potential to differentiate The first embryonic cells that arise from the division of the zygote are the ultimate stem cells; these stems cells are described as totipotent because they have the potential to differentiate into any of the cells needed to enable an organism to grow and develop The embryonic cells that develop from totipotent stem cells and are precursors to the fundamental tissue layers of the embryo are classified as pluripotent A pluripotent stem cell is one that has the potential to differentiate into any type of human tissue but cannot support the full development of an organism These cells then become slightly more specialized, and are referred to as multipotent cells A multipotent stem cell has the potential to differentiate into different types of cells within a given cell lineage or small number of lineages, such as a red blood cell or white blood cell Finally, multipotent cells can become further specialized oligopotent cells An oligopotent stem cell is limited to becoming one of a few different cell types In contrast, a unipotent cell is fully specialized and can only reproduce to generate more of its own specific cell type Stem cells are unique in that they can also continually divide and regenerate new stem cells instead of further specializing There are different stem cells present at different 1/6 Cellular Differentiation stages of a human’s life They include the embryonic stem cells of the embryo, fetal stem cells of the fetus, and adult stem cells in the adult One type of adult stem cell is the epithelial stem cell, which gives rise to the keratinocytes in the multiple layers of epithelial cells in the epidermis of skin Adult bone marrow has three distinct types of stem cells: hematopoietic stem cells, which give rise to red blood cells, white blood cells, and platelets ([link]); endothelial stem cells, which give rise to the endothelial cell types that line blood and lymph vessels; and mesenchymal stem cells, which give rise to the different types of muscle cells Hematopoiesis The process of hematopoiesis involves the differentiation of multipotent cells into blood and immune cells The multipotent hematopoietic stem cells give rise to many different cell types, including the cells of the immune system and red blood cells Differentiation When a cell differentiates (becomes more specialized), it may undertake major changes in its size, shape, metabolic activity, and overall function Because all cells in the body, beginning with the fertilized egg, contain the same DNA, how the different cell types 2/6 Cellular Differentiation come to be so different? The answer is analogous to a movie script The different actors in a movie all read from the same script, however, they are each only reading their own part of the script Similarly, all cells contain the same full complement of DNA, but each type of cell only “reads” the portions of DNA that are relevant to its own function In biology, this is referred to as the unique genetic expression of each cell In order for a cell to differentiate into its specialized form and function, it need only manipulate those genes (and thus those proteins) that will be expressed, and not those that will remain silent The primary mechanism by which genes are turned “on” or “off” is through transcription factors A transcription factor is one of a class of proteins that bind to specific genes on the DNA molecule and either promote or inhibit their transcription ([link]) Transcription Factors Regulate Gene Expression While each body cell contains the organism’s entire genome, different cells regulate gene expression with the use of various transcription factors Transcription factors are proteins that affect the binding of RNA polymerase to a particular gene on the DNA molecule Everyday Connection Stem Cell Research Stem cell research aims to find ways to use stem cells to regenerate and repair cellular damage Over time, most adult cells undergo the wear and tear of aging and lose their ability to divide and repair themselves Stem cells not display a particular morphology or function Adult stem cells, which exist as a small subset of cells in most tissues, keep dividing and can differentiate into a number of specialized ...Differential expression of endogenous sialidases of human monocytes during cellular differentiation into macrophages Nicholas M. Stamatos 1,2 , Feng Liang 3 , Xinli Nan 1 , Karine Landry 3 , Alan S. Cross 2 , Lai-Xi Wang 1 and Alexey V. Pshezhetsky 3 1 Institute of Human Virology, University of Maryland, Baltimore, MD, USA 2 Division of Infectious Diseases, Department of Medicine, University of Maryland Medical Center, Baltimore, MD, USA 3Ho ˆ pital Sainte-Justine and De ´ partement de Pe ´ diatrie, Universite ´ de Montre ´ al, Montre ´ al, Quebec, Canada Sialic acid is present on glycoproteins and glycolipids that are widely distributed throughout nature. Removal of sialic acid from these glycoconjugates on the surface of mammalian cells changes the functional capacity of the cells [1–8]. Sialidases comprise a family of enzymes that remove terminal sialyl residues from glycoconju- gates. Four genetically distinct forms of mammalian sialidase have been characterized, each with a predom- Keywords differentiation; glycoconjugates; human monocytes; sialidases; sialic acid Correspondence N. M. Stamatos, 725 West Lombard St., Institute of Human Virology, University of Maryland Medical System, Baltimore, MD 21201, USA Fax: +1 410 7064619 Tel: +1 410 7062645 E-mail: stamatos@umbi.umd.edu (Received 20 October 2004, revised 11 March 2005, accepted 22 March 2005) doi:10.1111/j.1742-4658.2005.04679.x Sialidases are enzymes that influence cellular activity by removing terminal sialic acid from glycolipids and glycoproteins. Four genetically distinct sia- lidases have been identified in mammalian cells. In this study, we demon- strate that three of these sialidases, lysosomal Neu1 and Neu4 and plasma membrane-associated Neu3, are expressed in human monocytes. When measured using the artificial substrate 2¢-(4-methylumbelliferyl)-a-d-N- acetylneuraminic acid (4-MU-NANA), sialidase activity of monocytes increased up to 14-fold per milligram of total protein after cells had differ- entiated into macrophages. In these same cells, the specific activity of other cellular proteins (e.g. b-galactosidase, cathepsin A and alkaline phospha- tase) increased only two- to fourfold during differentiation of monocytes. Sialidase activity measured with 4-MU-NANA resulted from increased expression of Neu1, as removal of Neu1 from the cell lysate by immuno- precipitation eliminated more than 99% of detectable sialidase activity. When exogenous mixed bovine gangliosides were used as substrates, there was a twofold increase in sialidase activity per milligram of total protein in monocyte-derived macrophages in comparison to monocytes. The increased activity measured with mixed gangliosides was not affected by removal of Neu1, suggesting that the expression of a sialidase other than Neu1 was present in macrophages. The amount of Neu1 and Neu3 RNAs detected by real time RT-PCR increased as monocytes differentiated into macro- phages, whereas the amount of Neu4 RNA decreased. No RNA encoding the cytosolic sialidase (Neu2) was detected in monocytes or macrophages. Western blot analysis using specific antibodies showed that the amount of Neu1 and Neu3 proteins increased during monocyte differentiation. Thus, the differentiation of monocytes into macrophages is associated with regu- lation of the expression of at least three distinct cellular sialidases, with specific up-regulation of the enzyme activity of only Neu1. Abbreviations LAMP-2, lysosome-associated membrane protein; 4-MU-NANA, 2¢-(4-methylumbelliferyl)-a- D-N-acetylneuraminic acid; PMN, polymorphonuclear leukocyte. FEBS Journal 272 (2005) BioMed Central Page 1 of 16 (page number not for citation purposes) Retrovirology Open Access Research Induction of the HIV-1 Tat co-factor cyclin T1 during monocyte differentiation is required for the regulated expression of a large portion of cellular mRNAs Wendong Yu 1 , Yan Wang 1 , Chad A Shaw 2 , Xiao-Feng Qin 3 and Andrew P Rice* 1 Address: 1 Department of Molecular Virology and Microbiology, Baylor College of Medicine, Houston, Texas 77030, USA, 2 Department of Human and Molecular Genetics, Baylor College of Medicine, Houston, Texas, USA and 3 Center for Cancer Immunology Research, Department of Immunology, The University of Texas M.D. Anderson Cancer Center, Houston, Texas, USA Email: Wendong Yu - wy132177@bcm.tmc.edu; Yan Wang - yw135452@bcm.tmc.edu; Chad A Shaw - cashaw@bcm.tmc.edu; Xiao- Feng Qin - fqin@mdanderson.org; Andrew P Rice* - arice@bcm.tmc.edu * Corresponding author Abstract Background: P-TEFb, a general RNA polymerase II elongation factor, is composed of CDK9 (cyclin-dependent kinase 9) as a catalytic unit and either cyclin T1, T2 or K as a regulatory subunit. The cyclin T1/P-TEFb complex is targeted by HIV to mediate Tat transactivation. Cyclin T1 protein expression is induced during early macrophage differentiation, suggesting a role in regulation of mRNA expression during the differentiation process. To study the functional significance of cyclin T1 induction during differentiation, we utilized the human Mono Mac 6 (MM6) monocytic cell line. Results: We found that cyclin T1 protein expression is induced by a post-transcriptional mechanism following PMA treatment of MM6 cells, similar to its induction in primary monocytes and macrophages. Also in agreement with findings in primary cells, cyclin T2a is present at relatively high levels in MM6 cells and is not induced by PMA. Although the knock-down of cyclin T1 in MM6 cells by shRNA inhibited HIV-1 Tat transactivation, MM6 cell growth was not affected by the depletion of cyclin T1. Using DNA microarray technology, we found that more than 20% of genes induced by PMA require cyclin T1 for their normal level of induction, and approximately 15% of genes repressed by PMA require cyclin T1 for their normal level of repression. Gene ontology analysis indicates that many of these cyclin T1-dependent genes are related to immune response and signal transduction. Conclusion: These results suggest that cyclin T1 serves a critical role in the program of macrophage differentiation, and this raises questions about the feasibility of cyclin T1 serving as an antiviral therapeutic target. Background Mammalian RNA polymerase II transcription (RNAP II) is a complex and coordinated process and its regulation is involved in many important cellular events such as differ- entiation, activation, and stress response. While the regu- lation of transcription initiation has been an actively Published: 09 June 2006 Retrovirology 2006, 3:32 doi:10.1186/1742-4690-3-32 Received: 27 April 2006 Accepted: 09 June 2006 This article is available from: http://www.retrovirology.com/content/3/1/32 © 2006 Yu et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0 ), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Retrovirology 2006, 3:32 http://www.retrovirology.com/content/3/1/32 Page 2 of 16 (page number not for citation purposes) studied area for decades, the regulation of transcription elongation has not been as actively investigated until recent years when a number of transcription elongation factors have been identified [1]. One factor of particular interest to transcriptional elongation is P-TEFb, a protein kinase that appears to regulate expression of a large por- tion of mammalian genes [2,3]. P-TEFb is believed to acti- vate transcriptional elongation through phosphorylation Genome Biology 2009, 10:R37 Open Access 2009Fraseret al.Volume 10, Issue 4, Article R37 Software Chromatin conformation signatures of cellular differentiation James Fraser * , Mathieu Rousseau † , Solomon Shenker * , Maria A Ferraiuolo * , Yoshihide Hayashizaki ‡ , Mathieu Blanchette † and Josée Dostie * Addresses: * Department of Biochemistry and McGill Cancer Center, McGill University, 3655 Promenade Sir-William-Osler, Montréal, H3G1Y6, Canada. † McGill Centre for Bioinformatics, McGill University, 3775 University, Montréal, H3A 2B4, Canada. ‡ RIKEN Omics Science Center, RIKEN Yokohama Institute, 1-7-22 Suehiro-cho Tsurumi-ku, Yokohama, 230-0045, Japan. Correspondence: Josée Dostie. Email: josee.dostie@mcgill.ca © 2009 Fraser et al.; licensee BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Chromatin conformation signatures<p>A suite of computer programs to identify genome-wide chromatin conformation signatures with 5C technology is reported.</p> Abstract One of the major genomics challenges is to better understand how correct gene expression is orchestrated. Recent studies have shown how spatial chromatin organization is critical in the regulation of gene expression. Here, we developed a suite of computer programs to identify chromatin conformation signatures with 5C technology http://Dostielab.biochem.mcgill.ca. We identified dynamic HoxA cluster chromatin conformation signatures associated with cellular differentiation. Genome-wide chromatin conformation signature identification might uniquely identify disease-associated states and represent an entirely novel class of human disease biomarkers. Rationale Cell specialization is the defining hallmark of metazoans and results from differentiation of precursor cells. Differentiation is characterized by growth arrest of proliferating cells fol- lowed by expression of specific phenotypic traits. This process is essential throughout development and for adult tissue maintenance. For example, improper cellular differentiation in adult tissues can lead to human diseases such as leukemia [1,2]. For this reason, identifying mechanisms involved in dif- ferentiation is not only essential to understand biology, but also to develop effective strategies for prevention, diagnosis and treatment of cancer. Suzuki et al. recently defined the underlying transcription network of differentiation in the THP-1 leukemia cell line [3]. Using several powerful genom- ics approaches, this study challenges the traditional views that transcriptional activators acting as master regulators mediate differentiation. Instead, differentiation is shown to require the concerted up- and down-regulation of numerous transcription factors. This study provides the first integrated picture of the interplay between transcription factors, proxi- mal promoter activity, and RNA transcripts required for dif- ferentiation of human leukemia cells. Although extremely powerful, several observations indicate that implementation of new technologies will be required to gain a full appreciation of how cells differentiate. First, gene expression is controlled by a complex array of regulatory DNA elements. Each gene may be controlled by multiple elements and each element may control multiple genes [4]. Second, the functional organization of genes and elements is not linear along chromosomes. For example, a given element may regu- late distant genes or genes located on other chromosomes without affecting the ones adjacent to it [4,5]. Third, gene reg- ulation is known to involve both local and long-range chro- Published: 19 April 2009 Genome Biology 2009, 10:R37 (doi:10.1186/gb-2009-10-4-r37) Received: 24 October 2008 Revised: 22 December 2008 Accepted: 19 April 2009 The MOLECULAR AND CELLULAR FUNCTIONS OF THE ALTERNATIVELY SPLICED ISOFORMS OF GDNF RECEPTOR COMPLEX IN NEURONAL DIFFERENTIATION ZHOU LIHAN B.Sc. (Hons.), NUS A THESIS SUBMITTED FOR THE DEGREE OF DOCTOR OF PHILOSOPHY DEPARTMENT OF BIOCHEMISTRY NATIONAL UNIVERSITY OF SINGAPORE 2012 i DECLARATION I hereby declare that this thesis is my original work and it has been written by me in its entirety. I have duly acknowledged all the sources of information which have been used in the thesis. This thesis has also not been submitted for any degree in any university previously. ZHOU LIHAN Dec 2012 ii ACKNOWLEDGEMENT “Tell me and I forget, teach me and I may remember, involve me and I learn.” ― Benjamin Franklin Neither this thesis, nor the man I am today, would be possible without the heroic effort of Professor Too Heng-Phon, whose philosophy of mentoring is a true embodiment of the quote. Professor Too never fails to captivate, inspire and involve his students in the pursuit of scientific excellence. Working alongside with him on the bench is one of the most daunting tasks any fresh graduate can face, but also a routine one would dearly miss when leaving his lab. Professor Too and his philosophy is truly the reason that I, and the many before me, continue to pursue the fun and challenges in the arena of science. I am also blessed to have Professor Tang Bor Luen and Professor Low Chian Ming as my thesis advisors. Special thanks for Professor Tang Bor Luen, who has been a wonderful advisor since my undergraduate days. It was my privilege to have worked with so many dynamic and intelligent lab members over the years. My heartfelt gratitude to Dr Yoong Li Foong and Dr Wan Guoqiang, whose constant assistance and assurance helped me to survive, grow and excel in the lab. Special thanks to Zou Ruiyang and Sarah Ho Yoon Khei for being such wonderful colleagues in our pursuit of the microRNA dream. I am also grateful to Jeremy Lim Qing’ En, Dr Zhou Kang, Sha Lanjie, Seow Kok Huei, Simon Zhang Congqiang, Chen Xixian, Cheng He, Wong Long Hui and Chin Meiyi for all the stimulating discussions, fun and laughter throughout the years. iii This thesis, is dedicated to my parents, grandparents and my wife, who tolerated my years of absence from their lives, and supported me with unrelenting kindness, understanding and love. You are truly the safe harbour a man can ever wish for. “For every fact there is an infinity of hypotheses.” ― Robert M. Pirsig I would also like to dedicate this thesis to those who find inspiration and use in its findings and analyses. It has been a truly enjoyable and rewarding experience making the observations, generating the hypotheses and uncovering the evidences. It is my greatest hope that these will be useful in spurring even more thoughts and hypotheses. iv Table of Contents ACKNOWLEDGEMENT III SUMMARY IX LIST OF FIGURES AND TABLES XII LIST OF ABBREVIATIONS XV CHAPTER INTRODUCTION 16 1.1 Motivations of the study 16 1.2 Organization of the thesis 17 1.3 List of related publications (published, submitted and in preparation) 18 1.4 List of Invention Disclosures 20 1.5 List of Awards 20 1.6 Conference Presentation 21 CHAPTER LITERATURE REVIEW 22 2.1 GDNF family of ligands (GFLs) 22 2.2 GDNF family of receptors (GFRs) and co-receptors 25 2.3 Alternatively spliced isoforms of GDNF receptors 28 2.4 GFL-GFRα-RET signaling and function 30 2.5 Conclusion 31 CHAPTER CYCLIC AMP SIGNALING THROUGH PKA BUT NOT EPAC IS ESSENTIAL FOR NEURTURIN-INDUCED BIPHASIC ERK1/2 ACTIVATION AND NEURITE OUTGROWTHS THROUGH GFRΑ2 ISOFORMS 33 Section 3.1 Introduction 33 Section 3.2 Results 3.2.1 NTN induced CREB phosphorylation, biphasic ERK1/2 activation and neurite outgrowth through selected GFRα isoforms 3.2.2 Cyclic AMP and Protein Kinase A signaling is involved in NTN-induced neurite outgrowth 34 34 37 v 3.2.3 De novo transcription and translation is required for late phase of ERK1/2 activation and ... different body tissues 4/6 Cellular Differentiation In contrast, adult stem cells isolated from a patient are not seen as foreign by the body, but they have a limited range of differentiation Some... body, beginning with the fertilized egg, contain the same DNA, how the different cell types 2/6 Cellular Differentiation come to be so different? The answer is analogous to a movie script The different... pluripotent stem cells (iPSCs) from mouse and human adult stem cells These cells are genetically 3/6 Cellular Differentiation reprogrammed multipotent adult cells that function like embryonic stem cells;