Epigenomics and Epigenetics Effector bottleneck: Microbial reprogramming of parasitized host cell transcription by epigenetic remodeling of chromatin structure. Sara H Sinclair, Kristen E Rennoll-Bankert and J Stephen Dumler Journal Name: Frontiers in Genetics ISSN: 1664-8021 Article type: Review Article Received on: 17 Jun 2014 Accepted on: 26 Jul 2014 Provisional PDF published on: 26 Jul 2014 www.frontiersin.org: www.frontiersin.org Citation: Sinclair SH, Rennoll-bankert KE and Dumler JS(2014) Effector bottleneck: Microbial reprogramming of parasitized host cell transcription by epigenetic remodeling of chromatin structure.. Front. Genet. 5:274. doi:10.3389/fgene.2014.00274 Copyright statement: © 2014 Sinclair, Rennoll-bankert and Dumler. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. This Provisional PDF corresponds to the article as it appeared upon acceptance, after rigorous peer-review. Fully formatted PDF and full text (HTML) versions will be made available soon. 1 Effector bottleneck: Microbial reprogramming of parasitized host 2 cell transcription by epigenetic remodeling of chromatin structure. 3 4 Running title: Epigenetic reprogramming by Anaplasma phagocytophilum 5 6 Sara H. Sinclair1,2,3,4, Kristen E. Rennoll-Bankert2,3, J. Stephen Dumler1,2,3,4* 7 8 1Graduate Program in Cellular and Molecular Medicine, The Johns Hopkins University School 9 of Medicine, Baltimore MD, USA 10 2Departments of Microbiology and Immunology, University of Maryland Baltimore, School of 11 Medicine, Baltimore MD, USA 12 3Department of Pathology, The Johns Hopkins University School of Medicine, Baltimore MD, 13 USA 14 4Department of Pathology, University of Maryland Baltimore, School of Medicine, Baltimore 15 MD, USA 16 17 *Correspondence: J. Stephen Dumler, M.D., Departments of Pathology and Microbiology & 18 Immunology, University of Maryland School of Medicine, 685 W. Baltimore St., Health 19 Sciences Facility-1 Room 322D, Baltimore, MD USA 21201 20 [email protected] 21 1 22 Abstract 23 Obligate intracellular pathogenic bacteria evolved to manipulate their host cells with a limited 24 range of proteins constrained by their compact genomes. The harsh environment of a phagocytic 25 defense cell is one that challenges the majority of commensal and pathogenic bacteria; yet, these 26 are the obligatory vertebrate homes for important pathogenic species in the Anaplasmataceae 27 family. Survival requires that the parasite fundamentally alter the native functions of the cell to 28 allow its entry, intracellular replication, and transmission to a hematophagous arthropod. The 29 small genomic repertoires encode several eukaryotic-like proteins, including Ankyrin A (AnkA) 30 of Anaplasma phagocytophilum and Ank200 and TRP proteins of Ehrlichia chaffeensis that 31 localize to the host cell nucleus and directly bind DNA. As a model, A. phagocytophilum AnkA 32 appears to directly alter host cell gene expression by recruiting chromatin modifying enzymes 33 such as histone deacetylases and methyltransferases or by acting directly on transcription in cis. 34 While cis binding could feasibly alter limited ranges of genes and cellular functions, the complex 35 and dramatic alterations in transcription observed with infection are difficult to explain on the 36 basis of individually targeted genes. We hypothesize that nucleomodulins can act broadly, even 37 genome-wide, to affect entire chromosomal neighborhoods and topologically-associating 38 chromatin domains by recruiting chromatin remodeling complexes or by altering the folding 39 patterns of chromatin that bring distant regulatory regions together to coordinate control of 40 transcriptional reprogramming. This review focuses on the A. phagocytophilum nucleomodulin 41 AnkA, how it impacts host cell transcriptional responses, and current investigations that seek to 42 determine how these multifunctional eukaryotic-like proteins facilitate epigenetic alterations and 43 cellular reprogramming at the chromosomal level. 44 Key words 45 epigenetics, nucleomodulin, DNA methylation, histone deacetylase, chromatin, Anaplasma 46 phagocytophilum 2 47 Introduction 48 In order to infect mammalian hosts, bacterial pathogens evolved an array of mechanisms that 49 serve to create an environment conducive for survival, replication, and spread. While many 50 bacterial species survive in an extracellular environment, intracellular pathogens must be capable 51 of both entering their host cells undetected and altering the cellular milieu in order to replicate. 52 Traditionally, the ability of bacterial-derived proteins to induce disruptions of cell signaling or 53 major cellular processes such as NF-κB, MAPK, and JAK/STAT pathways, are the predominant 54 focus of host-pathogen interaction studies (Brodsky and Medzhitov 2009). Recently, there has 55 been an increasing interest in the ability of these intracellular pathogens to direct alterations in 56 host cell gene expression that promote survival and replication (Silmon de Monerri and Kim 57 2014; Bierne et al.2012; Paschos and Allday 2010). It is now well recognized that bacterial 58 pathogens can reprogram host gene expression either directly or indirectly, by altering the 59 accessibility of gene promoters via epigenetic modifications. 60 Eukaryotic DNA is highly organized and gene expression tightly regulated by an orchestrated 61 network of proteins, RNAs and other modulators. Histone octamers, or nucleosomes, organize 62 DNA by acting as a bobbin on which the DNA winds. The charge of the histone proteins can be 63 covalently modified in order to more tightly or loosely associate with DNA. Histone acetylation, 64 which imparts a negative charge, is predominantly associated with an open configuration where 65 promoters are easily accessed by RNA polymerases. Histone methylation or phosphorylation 66 which impart a positive charge, cause DNA to more tightly associate with histone proteins, 67 reducing promoter accessibility to transcription activating machinery. The process of modulating 68 open and closed chromatin is further complicated by i) the residue(s) of which histone protein(s) 69 is/are modified, ii) methylation of cytosine residues in DNA, and iii) non-coding RNAs, and by 70 the manner in which these mechanisms are intertwined. It therefore is no surprise that bacterial- 71 derived proteins have evolved to interfere with host gene expression that improves bacterial 72 fitness. 73 Over the last decade, examples of secreted bacterial effector proteins, ranging from those 74 produced by Listeria monocytogenes, Chlamydia trachomatis, Shigella flexneri, and others, were 75 found to target the host cell nucleus (Silmon de Monerri and Kim 2014; Bierne et al.2012; 76 Paschos and Allday 2010). These discoveries demonstrate the relationship of altered 77 transcription and function of infected host cells and microbial survival, and in some cases, 78 pathogenicity. S. flexneri, a bacterial pathogen that can cause dysentery, prevents NF-κB from 79 binding its target gene promoters by altering the phosphorylation state of histone H3 at serine 10. 80 The bacterium does so by secreting outer membrane protein F (OspF) that de-phosphorylates 81 MAPKs in the nucleus resulting in a lack of histone H3 phosphorylation at serine 10 at a number 82 of NF-κB-dependent genes (Arbibe et al. 2007). By altering NF-κB target gene transcription, S. 83 flexneri suppresses the host cell inflammatory response promoting microbial survival and 84 transmission (Arbibe et al. 2007). L. monocytogenes nuclear targeted protein A (LntA), blocks 85 binding of heterochromatin inducing protein BAHD1 at interferon-stimulated genes resulting in 86 upregulated expression (Rohde 2011). Nuclear effector E (NUE) of C. trachomatis and RomA of 87 L. pneumophila have methyltransferase activity and induce methylation of eukaryotic histones 88 and altered host cell gene expression (Rolando et al. 2013; Pennini et al. 2010). Plant pathogens 89 in the genus Xanthomonas, as well as Ralstonia solanacearum, Burkholderia rhizoxinica and an 90 endosymbiont of the plant pathogenic fungus Rhizopus microsporus all possess genes encoding 91 transcription activator-like (TAL) protein effectors that when expressed, bind plant DNA and 3 92 alter transcription and disease susceptibility (Bogdanove 2014; Sugio et al. 2007). These 93 examples are part of an expanding array of bacterial-derived proteins termed nucleomodulins 94 that target host cell chromatin or chromatin-linked pathways to alter transcription, typically at 95 one or a few host genes. To date, the only prokaryotic nucleomodulins shown to directly bind 96 mammalian DNA and influence surrounding chromatin are from the Anaplasmataceae family. 97 Ankyrin A (AnkA) of Anaplasma phagocytophilum, as well as Ank200 and several tandem- 98 repeat containing proteins (TRPs) from Ehrlichia chaffeensis, have been shown to enter the 99 nucleus and bind DNA, and interact with host epigenetic machinery or alter nearby histone 100 octamers (Luo and McBride 2012; Zhu et al. 2011; Luo et al. 2011; Dunphy et al. 2013; Garcia- 101 Garcia et al. 2009a). 102 Targeting of individual genes or binding