Copyright Nicodemus Edrick Oey 2014 Abstract The brain is a highly plastic structure, capable of changing its responses to the external stimuli it receives. This evolutionarily conserved concept of neuroplasticity underlies all forms of memory, from sea slugs, flies, rodents, all the way to humans. While short-term memories that last for minutes are mediated by transient changes in neural connections, long-term memories that last for years require more persistent changes that involve the production of new proteins. Decades of research have shown that the molecular mechanisms responsible for the consolidation of long-term memory are unlikely to be mediated by changes to the static genomic DNA sequence. The last twenty years have seen the emergence of epigenetics as a highly sophisticated mechanism by which a neuron can dictate with remarkable specificity which genes should be expressed at precisely which time in response to activity. Neuronal activity-dependent gene transcription depends on the action of several enzymes that respond to activity to specifically regulate the expression of genes that effectuate downstream functions. The ability of epigenetic regulators to tag specific locations in the genome for the de-novo transcription of genes has proven to be essential to learning and memory, as indicated by the disastrous consequences of their absence in various clinical syndromes of mental retardation. The present work attempts to study and characterize the events that mediate long-term memory consolidation from an epigenetics standpoint, specifically in chromatin modification or “epigenetic tagging” of specific nucleosomes which seem to be involved in both early and late events of memory consolidation along the temporal axis of neural activity. Amongst the many epigenetic regulators important for memory function, the TIP60 protein, in particular, is of significant interest due to its involvement both in early events of neuronal activity-dependent gene induction, and also in late events consisting of epigenetic changes leading to long-lasting memory consolidation. Using a combination of biochemistry, super-resolution microscopy, chromatin immunoprecipitation, and mass spectrometry-based techniques, the first part of this thesis presents findings on the role of the Alzheimer’s Disease-associated epigenetic enzyme TIP60 and an X-linked Mental Retardation (XLMR)-associated protein PHF8 in the rapid neuronal activity-dependent transcription of ARC, a crucial regulator of memory consolidation. The second part of this thesis will explore the role of TIP60 in mediating the functions of ARC protein itself in the late epigenetic events that eventually result in memory consolidation. The last part of the thesis will be devoted to discussing the importance of epigenetic processes of chromatin modification in general neuronal functions such as development and survival, as well as specific functions such as memory consolidation. Finally, looking forward to the future, several of the potentially endless possibilities in neuroepigenetics such as clinical applications of epigenetic modifying therapy and ARC-modulating strategies in neuropsychiatric disorders will be offered. iv Dedication As you read the words written on this page, neuronal cells in your brain are firing in highly patterned electrical activities across their synapses in order to encode the information you read, some of which you may remember, being eventually stored in your memory. This remarkable ability of the brain which is composed of over 200 billion neurons and more than 100 trillion synapses is thanks to the amazing capabilities found in each unique neuron, which is able to change the genes it expresses at any given time in response to the pattern of activity it receives. Such is the dynamics of neuronal cells that networks of them are able to underlie our ability to see, to hear, to smell, to taste, to move, to feel, and most importantly, to think… I would like to dedicate these four years’ work to my parents, Hoat and Megan Oey, for bringing me to life, for raising me as a good son and brother, and for allowing me to travel 15349km to Singapore to start living my dream of being a Clinician-Scientist, a long, arduous journey which this PhD dissertation is a part of: Mom and Dad, I love you. I would like to thank my siblings for all their support: my sister Elrika and new brother-in-law Christopher Prest in Waterloo for allowing me to sing at their wedding, my younger sister Elvina Oey in New York for the invaluable Occupational Therapist and Psychologist point-of-view, my younger brother Edbert Oey for muscling me into the right path no matter how much I strayed, my younger cousin William Tanaka Oey for being the fun, kind-hearted man that he is and Carissa Oey for taking care of my parents when I should be the one doing it…. thank you to all my wonderful siblings: I love you guys. One incredibly special mention goes out to the love of my life, Christine Chan, PhD candidate, for correcting my drawings of neurons that look like “Strepsils™”, for those many hours of rehearsals before presentations, and for helping me clean my incredibly messy room: I love you, Hui Shan. I would like to thank the VanDongen Laboratory: Shaun Teo and Caroline Wee, for holding me by the hand when I was first trying to walk (aka run gels), Rajaram Ezhilarasan for the hard work and dedication, Niamh Higgins, Knvul Sheikh, Annabel Tan, Gokul Banumurthy our amazing RA’s who work day and night and on the weekends too, Ju Han the computer genius who taught me Chinese, Mark Dranias for the discussions on synaptic plasticity, Xiaoyu for investigating long-term memory in cultured neurons, and especially How Wing Leung, our post-doc, who has taught me everything I know. Finally, I would like to say to my lab mom and dad, Tony and Margon VanDongen, “Aren’t you glad I survived?” – thank you Margon for the best spaghetti I’ve ever tasted, for believing in me when no one else did, and for defending me when odds were against it, thank you Tony for mentoring me… you have taught me how to science, and that is something I will never forget for the rest of my life. I am forever in your debt. With that, I hope you don’t mind me saying that I am very much looking forward to the exciting research we have planned together on the horizon! v Contents Title Page . i Abstract Signature ii Copyright . iii Abstract . iv Dedication . v Table of Contents . vi List of Figures x List of Abbreviations xii Chapter Review of Literature . I. Introduction to Learning and Memory A. A short history . B. Classifications of memory . II. Molecular Mechanisms of Long-Term Memory Formation A. Overview: plasticity and activity B. Epigenetics as a mechanism of activity-dependent gene expression C. ARC: a master regulator of synaptic plasticity 11 D. TIP60: an effector of early and late neuroepigenetic events . 15 E. PHF8: a specialized neuronal transcriptional co-activator 16 III. The Timeline of Neuronal Activation 18 Chapter Early Epigenetic Events: the Characterization of a Chromatin-modifying Complex Composed of PHF8 and TIP60 that Alter H3K9acS10P to Enable Activity-dependent Transcription of Arc 20 I. Abstract 20 II. Introduction 21 III. Materials and Methods . 24 A. Plasmid Construction and Cloning 24 B. Hippocampal and Cortical Neuronal Cell Culture 25 C. Transfections and Neuronal Stimulations . 26 D. Conventional Immunofluorescence . 26 E. Proximity Ligation In-Situ Assay . 27 F. Widefield Microscopy, Calcium imaging, and Data Analysis . 28 G. Co-immunoprecipitation and Western Blotting 29 H. Immunoprecipitation followed by Mass Spectrometry . 30 vi I. Chromatin Immunoprecipitation (ChIP) and Triton X-Acetic AcidUrea histone gel electrophoresis 30 J. 3-dimensional Structured Illumination Microscopy 32 K. 3-dimensional Stochastic Optical Reconstruction Microscopy 32 IV. Results . 33 A. Transcriptional Activators PHF8 and TIP60 Colocalize in the Interchromatin Space 33 B. The Histone Demethylase PHF8 Physically Associates the Histone Acetyltransferase TIP60 37 C. PHF8 and TIP60 Form a Dual-Function Complex that Increases Histone Acetylation on H3K4me3-bearing Chromatin . 39 D. PHF8 Removes Transcriptionally Suppressive H3K9me2 and Associates with Transcriptionally Active H3K9ac 42 E. PHF8 and TIP60 are Activity-dependent and Co-regulate H3K9acS10P in Response to Neuronal Activity 43 F. The PHF8-TIP60 Complex Modulates activity-induced H3K9acS10P 46 G. The PHF8-TIP60 Interactome is Rich in Proteins Involved in Transcription and Includes the Neuronal Splicing Factor PSF 50 H. Super-resolution Microscopy Situates Endogenous PHF8, TIP60, and PSF Within 30nm of Each Other in the Activated Neuronal Nucleus . 53 V. Discussion 56 Chapter Late Epigenetic Events: the Interaction Between TIP60 and ARC Functions to Regulate H4K12ac, a Learning-induced Chromatin Modification Involved in Ageing-associated Memory Impairment 64 I. Abstract 64 II. Introduction 66 III. Materials and Methods . 68 A. Constructs and Cloning 68 B. Cell Culture . 69 C. Transfections and Stimulations . 70 D. Immunofluorescence 70 E. Imaging and Data Analysis . 71 F. 3-dimensional Structured Illumination Microscopy 72 G. Photo-activated Localization Microscopy (PALM) and Direct Stochastic Optical Reconstruction Microscopy (dSTORM) . 72 H. Immunoprecipitation and Western Blotting . 73 I. Induction of Arc gene expression by stimulation of neural network activity . 74 IV. Results . 75 vii V. A. ARC Protein Interacts with betaSpIVSigma5, PHF8, PML and TIP60 and Components of the TIP60 Chromatin Remodeling Complex 75 B. PML, TIP60, and ARC Form a Tight Complex in the Nucleus of Activated Neurons 77 C. TIP60 and ARC Overexpression Increases H4K12 Acetylation but not H3K9, H3K14, H2AK5, or H2BK5 Acetylation . 78 D. ARC, PML, and PHF8 Modulate TIP60’s Acetyltransferase Activity 80 E. Endogenous ARC Interacts with TIP60 in a Variety of Dynamic Nuclear Structures as Seen on Localization Microscopy 81 F. Endogenous ARC is Correlated with High TIP60 Nuclear Levels in Activated Neurons 84 G. ARC Recruits TIP60 to PML Bodies . 85 H. Activity-induced ARC Increases H4K12 Acetylation at a Timepoint that Correlates with Memory Consolidation in Neurons 86 I. The Enzymatically Inactive Mutant of TIP60 Fails to Induce H4K12 Acetylation in Hippocampal Neurons 88 J. ARC Associates at Single-Molecule Level with the LearningInduced Histone Mark H4K12ac . 89 Discussion 91 Chapter Integrating the Findings: the Elucidation of the Genes and Mechanisms that lead to Memory Consolidation 98 I. Introduction 98 II. Preliminary Results and Discussion A. In-vitro Neural Network Activity Leads to Specific Site-Directed Changes in Chromatin Modification 101 B. In-vivo Novel Environment Enrichment Leads to Specific Patterns of Chromatin Modification Partly Mediated by PHF8 and TIP60102 C. PHF8 and TIP60 are Activity-Dependent Chromatin-modifying Enzymes With Different Promoter Occupancy Profiles . 103 D. The Transcriptional Activator PHF8 is Found Within Nanometres of PTB-associated Splicing Factor and Nascent RNA 105 E. The Recruitment of PHF8 to Active Transcriptional Start Sites Precedes RNA Polymerase II Binding at the Arc and c-Fos Genomic Loci Following Neuronal Activation 106 F. Specific Regulation of Arc Gene Expression by ERK and p38 MAPK Signaling Pathways . 107 G. The Interactome of PHF8, TIP60, and ARC Give Novel Clues to the Processes that Lead Ultimately to Memory Consolidation 110 viii Chapter Conclusions and Future Directions: Towards Epigenetically Informed Translational and Clinical Trials . 117 Bibliography 125 Appendix A. Publications accepted or under review 138 ix List of Figures Figure 1: PHF8 and TIP60 colocalize and recruit each other in neuronal interchromatin space. . . 35 Figure 2: PHF8 and TIP60 physically associate to form a dual function chromatin-modifying complex . 38 Figure 3: PHF8 removes the repressive histone mark H3K9me2 and associates with the activating histone mark H3K9ac. 42 Figure 4: Neuronal activity reorganizes PHF8 and TIP60 in the nucleus and effectuate histone methylation and acetylation changes. . 45 Figure 5: PHF8 and TIP60 modulate neuronal activity-induced histone acetylation at H3K9acS10P and activation of the Arc gene 48 Figure 6: Knockdown of PHF8 impairs activity-dependent induction of H3K9acS10P and Arc and c-Fos expression . 49 Figure 7: PHF8, TIP60, and H3K9acS10P are specifically enriched in the transcriptional start site of the Arc gene. 50 Figure 8: Common interacting partners between PHF8 and TIP60 function primarily in transcription and mRNA processing. 52 Figure 9: Endogenous TIP60 is located within 30nm of PHF8 in the activated hippocampal neuronal nucleus . 54 Figure 10: PHF8 and TIP60 form a tripartite complex with the splicing factor PSF/SFPQ 55 Figure 11: Four-color immunofluorescence of a quaternary complex formed between ARC, PML, bSpectrin, and TIP60 77 Figure 12: ARC protein interacts with two members of the TIP60 chromatin remodeling complex: the transcriptional coactivator BRG1 and AMIDA . 77 Figure 13: Endogenous ARC is able to localize TIP60 to PML bodies . 79 Figure 14: ARC+TIP60 overexpression had a mild effect on global H4K12 acetylation 80 Figure 15: ARC has a positive modulatory effect on TIP60-mediated H4K12 acetylation. . 81 Figure 16: 3D Stimulated Emission Depletion Microscopy shows association of endogenous Arc and Tip60. 83 Figure 17: Dual-color super-resolution microcopy of Arc-mEOS2 and endogenous Tip60 in the activated neuronal nucleus. . 84 x bring up an important issue underlying all of neurobiological research: in revisiting the issue of a “black box” between what happens at the level of molecules and cells to what happens at the level of the brain and the whole organism, is there a unified mechanism or concept that can reconcile findings observed at the nucleus with those observed in human behavior? The answer to this question will likely redefine the way neuroscience is practiced in the laboratories as well as in the clinics. In summary, research that has stemmed from findings described in this thesis and work that is currently underway in the clinics are beginning to study, characterize, and possibly treat disorders that affect long-term memory formation at the most fundamental level: that of genes in the nucleus. Beyond the shadow of a doubt, it is now known that neuronal activity triggers the expression of important molecules that eventually result in memory consolidation, and future research delving deeper into these mechanisms will likely shed light on the molecular pathways that underlie the way we think, behave, and remember. 123 Figure 34 – Graphical Summary of Present Dissertation – Experience, in the form of environmental stimuli / learning, mediate changes in synaptic activity. Electrical and chemical synaptic activity is transduced to NMDA receptor opening and/or G-protein coupled receptor signaling, which intracellularly follows through a kinase signaling cascade. The important kinase pathways include MAPK and p38-mediated pathways that eventually activate epigenetic enzymes in the nucleus which are responsible for epigenetic chromatin modification that ultimately lead to changes in memory-related gene activation or suppression. 124 Bibliography 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21. 22. Salomon, J.A., et al. Healthy life expectancy for 187 countries, 1990-2010: a systematic analysis for the Global Burden Disease Study 2010. Lancet 380, 21442162 (2012). Fryns, J.P., et al. 9th international workshop on fragile X syndrome and X-linked mental retardation. Am J Med Genet 94, 345-360 (2000). Lashley, K.S. In search of the engram. in Physiological mechanisms in animal behavior. (Society's Symposium IV.) 454-482 (Academic Press, Oxford, England, 1950). Hebb, D.O. The Organization of Behavior: A Neuropsychological Theory, (Taylor & Francis, 2005). Atkinson, R.C. & Shiffrin, R.M. Human memory: A proposed system and its control processes. Psychology of learning and motivation 2, 89-195 (1968). Dick, A.O. Iconic memory and its relation to perceptual processing and other memory mechanisms. Perception & Psychophysics 16, 575-596 (1974). Coltheart, M. Iconic memory and visible persistence. Perception & Psychophysics 27, 183-228 (1980). Miller, G.A. The magical number seven, plus or minus two: some limits on our capacity for processing information. Psychological Review 63, 81-97 (1956). Dudai, Y. The neurobiology of consolidations, or, how stable is the engram? Annual review of psychology 55, 51-86 (2004). Ju, H., Xu, J.X., Chong, E. & VanDongen, A.M. Effects of synaptic connectivity on liquid state machine performance. Neural networks : the official journal of the International Neural Network Society 38, 39-51 (2013). Dranias, M.R., Ju, H., Rajaram, E. & VanDongen, A.M. Short-term memory in networks of dissociated cortical neurons. J Neurosci 33, 1940-1953 (2013). Squire, L.R. & Barondes, S.H. Actinomycin-D: effects on memory at different times after training. Nature 225, 649-650 (1970). Roberts, R.B. & Flexner, L.B. The biochemical basis of long-term memory. Q Rev Biophys 2, 135-173 (1969). Agranoff, B.W., Davis, R.E., Casola, L. & Lim, R. Actinomycin D blocks formation of memory of shock-avoidance in goldfish. Science 158, 1600-1601 (1967). Izquierdo, L.A., et al. Molecular pharmacological dissection of short- and long-term memory. Cell Mol Neurobiol 22, 269-287 (2002). Kandel, E.R. The molecular biology of memory storage: a dialog between genes and synapses. Biosci Rep 24, 475-522 (2004). Lee, Y.S., Bailey, C.H., Kandel, E.R. & Kaang, B.K. Transcriptional regulation of longterm memory in the marine snail Aplysia. Mol Brain 1, (2008). Yin, J.C., Del Vecchio, M., Zhou, H. & Tully, T. CREB as a memory modulator: induced expression of a dCREB2 activator isoform enhances long-term memory in Drosophila. Cell 81, 107-115 (1995). Frank, D.A. & Greenberg, M.E. CREB: a mediator of long-term memory from mollusks to mammals. Cell 79, 5-8 (1994). Josselyn, S.A. & Nguyen, P.V. CREB, synapses and memory disorders: past progress and future challenges. Curr Drug Targets CNS Neurol Disord 4, 481-497 (2005). De Luca, A. & Giuditta, A. Role of a transcription factor (CREB) in memory processes. Riv Biol 90, 371-384 (1997). Greer, P.L. & Greenberg, M.E. From synapse to nucleus: calcium-dependent gene transcription in the control of synapse development and function. Neuron 59, 846860 (2008). 125 23. 24. 25. 26. 27. 28. 29. 30. 31. 32. 33. 34. 35. 36. 37. 38. 39. 40. 41. 42. 43. Cohen, S. & Greenberg, M.E. Communication between the synapse and the nucleus in neuronal development, plasticity, and disease. Annu Rev Cell Dev Biol 24, 183-209 (2008). Deisseroth, K., Bito, H. & Tsien, R.W. Signaling from synapse to nucleus: postsynaptic CREB phosphorylation during multiple forms of hippocampal synaptic plasticity. Neuron 16, 89-101 (1996). Reik, W. Stability and flexibility of epigenetic gene regulation in mammalian development. Nature 447, 425-432 (2007). Bird, A. Perceptions of epigenetics. Nature 447, 396-398 (2007). Conaway, J.W. Introduction to theme "Chromatin, epigenetics, and transcription". Annu Rev Biochem 81, 61-64 (2012). Molfese, D.L. Advancing neuroscience through epigenetics: molecular mechanisms of learning and memory. Dev Neuropsychol 36, 810-827 (2011). Urdinguio, R.G., Sanchez-Mut, J.V. & Esteller, M. Epigenetic mechanisms in neurological diseases: genes, syndromes, and therapies. Lancet Neurol 8, 1056-1072 (2009). Tsankova, N., Renthal, W., Kumar, A. & Nestler, E.J. Epigenetic regulation in psychiatric disorders. Nat Rev Neurosci 8, 355-367 (2007). Ropers, H.H. & Hamel, B.C. X-linked mental retardation. Nat Rev Genet 6, 46-57 (2005). Petrij, F., et al. Rubinstein-Taybi syndrome caused by mutations in the transcriptional co-activator CBP. Nature 376, 348-351 (1995). Laumonnier, F., et al. Mutations in PHF8 are associated with X linked mental retardation and cleft lip/cleft palate. Journal of medical genetics 42, 780-786 (2005). Jones, K.J. & North, K.N. Developmental delay, expressive aphasia, hypotonia and dysmorphism in two brothers: an X-linked mental retardation syndrome? Clin Genet 54, 443-445 (1998). Ahmad, W., et al. Linkage mapping of a new syndromic form of X-linked mental retardation, MRXS7, associated with obesity. Eur J Hum Genet 7, 828-832 (1999). Kutsche, K., et al. Mutations in ARHGEF6, encoding a guanine nucleotide exchange factor for Rho GTPases, in patients with X-linked mental retardation. Nat Genet 26, 247-250 (2000). Lungarotti, M.S., et al. X-linked mental retardation, microcephaly, and growth delay associated with hereditary bullous dystrophy macular type: report of a second family. Am J Med Genet 51, 598-601 (1994). Allen, K.M., et al. PAK3 mutation in nonsyndromic X-linked mental retardation. Nat Genet 20, 25-30 (1998). Martinez, F., Gal, A., Palau, F. & Prieto, F. Localization of a gene for X-linked nonspecific mental retardation (MRX24) in Xp22.2-p22.3. Am J Med Genet 55, 387390 (1995). Mattaliano, M.D., Montana, E.S., Parisky, K.M., Littleton, J.T. & Griffith, L.C. The Drosophila ARC homolog regulates behavioral responses to starvation. Mol Cell Neurosci 36, 211-221 (2007). Miyashita, T., Kubik, S., Lewandowski, G. & Guzowski, J.F. Networks of neurons, networks of genes: an integrated view of memory consolidation. Neurobiol Learn Mem 89, 269-284 (2008). Ivanova, T.N., et al. Arc/Arg3.1 mRNA expression reveals a subcellular trace of prior sound exposure in adult primary auditory cortex. Neuroscience 181, 117-126 (2011). Ploski, J.E., et al. The activity-regulated cytoskeletal-associated protein (Arc/Arg3.1) is required for memory consolidation of pavlovian fear conditioning in the lateral amygdala. J Neurosci 28, 12383-12395 (2008). 126 44. 45. 46. 47. 48. 49. 50. 51. 52. 53. 54. 55. 56. 57. 58. 59. 60. 61. 62. Gusev, P.A., Cui, C., Alkon, D.L. & Gubin, A.N. Topography of Arc/Arg3.1 mRNA expression in the dorsal and ventral hippocampus induced by recent and remote spatial memory recall: dissociation of CA3 and CA1 activation. J Neurosci 25, 93849397 (2005). Inberg, S., Elkobi, A., Edri, E. & Rosenblum, K. Taste familiarity is inversely correlated with Arc/Arg3.1 hemispheric lateralization. J Neurosci 33, 11734-11743 (2013). Tagawa, Y., Kanold, P.O., Majdan, M. & Shatz, C.J. Multiple periods of functional ocular dominance plasticity in mouse visual cortex. Nat Neurosci 8, 380-388 (2005). Guzowski, J.F., et al. Mapping behaviorally relevant neural circuits with immediateearly gene expression. Curr Opin Neurobiol 15, 599-606 (2005). Plath, N., et al. Arc/Arg3.1 is essential for the consolidation of synaptic plasticity and memories. Neuron 52, 437-444 (2006). Moorman, S., Mello, C.V. & Bolhuis, J.J. From songs to synapses: molecular mechanisms of birdsong memory. Molecular mechanisms of auditory learning in songbirds involve immediate early genes, including zenk and arc, the ERK/MAPK pathway and synapsins. Bioessays 33, 377-385 (2011). Eriksson, T.M., et al. Emotional memory impairments in a genetic rat model of depression: involvement of 5-HT/MEK/Arc signaling in restoration. Mol Psychiatry 17, 173-184 (2012). Kerrigan, T.L. & Randall, A.D. A new player in the "synaptopathy" of Alzheimer's disease - arc/arg 3.1. Front Neurol 4, (2013). Messaoudi, E., et al. Sustained Arc/Arg3.1 synthesis controls long-term potentiation consolidation through regulation of local actin polymerization in the dentate gyrus in vivo. J Neurosci 27, 10445-10455 (2007). Guzowski, J.F., et al. Inhibition of activity-dependent arc protein expression in the rat hippocampus impairs the maintenance of long-term potentiation and the consolidation of long-term memory. J Neurosci 20, 3993-4001 (2000). Jakkamsetti, V., et al. Experience-induced Arc/Arg3.1 primes CA1 pyramidal neurons for metabotropic glutamate receptor-dependent long-term synaptic depression. Neuron 80, 72-79 (2013). Park, S., et al. Elongation factor and fragile X mental retardation protein control the dynamic translation of Arc/Arg3.1 essential for mGluR-LTD. Neuron 59, 70-83 (2008). Shepherd, J.D. & Bear, M.F. New views of Arc, a master regulator of synaptic plasticity. Nat Neurosci 14, 279-284 (2011). Korb, E. & Finkbeiner, S. Arc in synaptic plasticity: from gene to behavior. Trends Neurosci 34, 591-598 (2011). Steward, O. & Worley, P.F. Selective targeting of newly synthesized Arc mRNA to active synapses requires NMDA receptor activation. Neuron 30, 227-240 (2001). Waltereit, R., et al. Arg3.1/Arc mRNA induction by Ca2+ and cAMP requires protein kinase A and mitogen-activated protein kinase/extracellular regulated kinase activation. J Neurosci 21, 5484-5493 (2001). Wang, Y., Zheng, F., Zhou, X., Sun, Z. & Wang, H. Converging signal on ERK1/2 activity regulates group I mGluR-mediated Arc transcription. Neurosci Lett 460, 3640 (2009). Adams, J.P., Robinson, R.A., Hudgins, E.D., Wissink, E.M. & Dudek, S.M. NMDA receptor-independent control of transcription factors and gene expression. Neuroreport 20, 1429-1433 (2009). Dyrvig, M., et al. Epigenetic regulation of Arc and c-Fos in the hippocampus after acute electroconvulsive stimulation in the rat. Brain Res Bull 88, 507-513 (2012). 127 63. 64. 65. 66. 67. 68. 69. 70. 71. 72. 73. 74. 75. 76. 77. 78. 79. 80. Saha, R.N., et al. Rapid activity-induced transcription of Arc and other IEGs relies on poised RNA polymerase II. Nat Neurosci 14, 848-856 (2011). Dynes, J.L. & Steward, O. Arc mRNA docks precisely at the base of individual dendritic spines indicating the existence of a specialized microdomain for synapsespecific mRNA translation. J Comp Neurol 520, 3105-3119 (2012). Bloomer, W.A., VanDongen, H.M. & VanDongen, A.M. Arc/Arg3.1 translation is controlled by convergent N-methyl-D-aspartate and Gs-coupled receptor signaling pathways. J Biol Chem 283, 582-592 (2008). McIntyre, C.K., et al. Memory-influencing intra-basolateral amygdala drug infusions modulate expression of Arc protein in the hippocampus. Proc Natl Acad Sci U S A 102, 10718-10723 (2005). Rial Verde, E.M., Lee-Osbourne, J., Worley, P.F., Malinow, R. & Cline, H.T. Increased expression of the immediate-early gene arc/arg3.1 reduces AMPA receptormediated synaptic transmission. Neuron 52, 461-474 (2006). Bloomer, W.A., VanDongen, H.M. & VanDongen, A.M. Activity-regulated cytoskeleton-associated protein Arc/Arg3.1 binds to spectrin and associates with nuclear promyelocytic leukemia (PML) bodies. Brain Res 1153, 20-33 (2007). Korb, E., Wilkinson, C.L., Delgado, R.N., Lovero, K.L. & Finkbeiner, S. Arc in the nucleus regulates PML-dependent GluA1 transcription and homeostatic plasticity. Nat Neurosci 16, 874-883 (2013). Wee, C.L., et al. Nuclear Arc Interacts with the Histone Acetyltransferase Tip60 to Modify H4K12 Acetylation. eneuro, ENEURO. 0019-0014.2014 (2014). van der Burg, M., et al. Defective Artemis nuclease is characterized by coding joints with microhomology in long palindromic-nucleotide stretches. Eur J Immunol 37, 3522-3528 (2007). Gedeon, A.K., Nelson, J., Gecz, J. & Mulley, J.C. X-linked mild non-syndromic mental retardation with neuropsychiatric problems and the missense mutation A365E in PAK3. Am J Med Genet A 120A, 509-517 (2003). Tzschach, A., et al. Novel JARID1C/SMCX mutations in patients with X-linked mental retardation. Hum Mutat 27, 389 (2006). Dieckmann, P.M., Lucena, L.C., Dutra, L.A., Pedroso, J.L. & Barsottini, O.G. Marfanoid features and X-linked mental retardation associated with craniofacial abnormalities: the Lujan-Fryns syndrome. Arq Neuropsiquiatr 71, 68-69 (2013). Field, M., et al. Mutations in the BRWD3 gene cause X-linked mental retardation associated with macrocephaly. Am J Hum Genet 81, 367-374 (2007). Iwase, S., et al. The X-linked mental retardation gene SMCX/JARID1C defines a family of histone H3 lysine demethylases. Cell 128, 1077-1088 (2007). De Vos, B., Frints, S., Borghgraef, M. & Fryns, J.P. Cognitive and behavioral characteristics in affected males of a family with non-specific X-linked mental retardation and TM4 SF2-gene mutation. Genet Couns 13, 191-194 (2002). Leshinsky-Silver, E., et al. MEHMO (Mental retardation, Epileptic seizures, Hypogenitalism, Microcephaly, Obesity): a new X-linked mitochondrial disorder. Eur J Hum Genet 10, 226-230 (2002). Plenge, R.M., Stevenson, R.A., Lubs, H.A., Schwartz, C.E. & Willard, H.F. Skewed Xchromosome inactivation is a common feature of X-linked mental retardation disorders. Am J Hum Genet 71, 168-173 (2002). Lorbeck, M., Pirooznia, K., Sarthi, J., Zhu, X. & Elefant, F. Microarray analysis uncovers a role for Tip60 in nervous system function and general metabolism. PLoS One 6, e18412 (2011). 128 81. 82. 83. 84. 85. 86. 87. 88. 89. 90. 91. 92. 93. 94. 95. 96. 97. 98. 99. 100. Villard, L. & Fontes, M. Alpha-thalassemia/mental retardation syndrome, X-Linked (ATR-X, MIM #301040, ATR-X/XNP/XH2 gene MIM #300032). Eur J Hum Genet 10, 223-225 (2002). Klauck, S.M., et al. A mutation hot spot for nonspecific X-linked mental retardation in the MECP2 gene causes the PPM-X syndrome. Am J Hum Genet 70, 1034-1037 (2002). Zhang, S. & Krahe, R. Physical and transcript map of a 2-Mb region in Xp22.1 containing candidate genes for X-linked mental retardation and short stature. Genomics 79, 274-277 (2002). Sugie, H. [X-linked mental retardation, fragile X syndrome]. Ryoikibetsu Shokogun Shirizu, 830-832 (2001). Chelly, J. & Mandel, J.L. Monogenic causes of X-linked mental retardation. Nat Rev Genet 2, 669-680 (2001). Wu, Q., et al. PML3 Orchestrates the Nuclear Dynamics and Function of TIP60. J Biol Chem 284, 8747-8759 (2009). Grinkevich, L.N. [Investigation of histone H3 methylation during long term memory formation]. Ross Fiziol Zh Im I M Sechenova 98, 1111-1118 (2012). Gupta-Agarwal, S., Jarome, T.J., Fernandez, J. & Lubin, F.D. NMDA receptor- and ERK-dependent histone methylation changes in the lateral amygdala bidirectionally regulate fear memory formation. Learn Mem 21, 351-362 (2014). Gupta, S., et al. Histone methylation regulates memory formation. J Neurosci 30, 3589-3599 (2010). Santos, C., et al. A novel mutation in JARID1C gene associated with mental retardation. Eur J Hum Genet 14, 583-586 (2006). Jensen, L.R., et al. Mutations in the JARID1C gene, which is involved in transcriptional regulation and chromatin remodeling, cause X-linked mental retardation. Am J Hum Genet 76, 227-236 (2005). Feng, W., Yonezawa, M., Ye, J., Jenuwein, T. & Grummt, I. PHF8 activates transcription of rRNA genes through H3K4me3 binding and H3K9me1/2 demethylation. Nature structural & molecular biology 17, 445-450 (2010). Fortschegger, K., et al. PHF8 targets histone methylation and RNA polymerase II to activate transcription. Mol Cell Biol 30, 3286-3298 (2010). Kleine-Kohlbrecher, D., et al. A functional link between the histone demethylase PHF8 and the transcription factor ZNF711 in X-linked mental retardation. Mol Cell 38, 165-178 (2010). Asensio-Juan, E., Gallego, C. & Martinez-Balbas, M.A. The histone demethylase PHF8 is essential for cytoskeleton dynamics. Nucleic acids research 40, 9429-9440 (2012). Arteaga, M.F., et al. The histone demethylase PHF8 governs retinoic acid response in acute promyelocytic leukemia. Cancer cell 23, 376-389 (2013). Hardingham, G.E., Arnold, F.J. & Bading, H. Nuclear calcium signaling controls CREBmediated gene expression triggered by synaptic activity. Nat Neurosci 4, 261-267 (2001). Nithianantharajah, J. & Hannan, A.J. Enriched environments, experience-dependent plasticity and disorders of the nervous system. Nat Rev Neurosci 7, 697-709 (2006). West, A.E. & Greenberg, M.E. Neuronal activity-regulated gene transcription in synapse development and cognitive function. Cold Spring Harbor perspectives in biology 3(2011). Inoue, M., et al. Synaptic activity-responsive element (SARE): A unique genomic structure with an unusual sensitivity to neuronal activity. Commun Integr Biol 3, 443446 (2010). 129 101. 102. 103. 104. 105. 106. 107. 108. 109. 110. 111. 112. 113. 114. 115. 116. 117. 118. 119. 120. Day, J.J. & Sweatt, J.D. Epigenetic mechanisms in cognition. Neuron 70, 813-829 (2011). Ronan, J.L., Wu, W. & Crabtree, G.R. From neural development to cognition: unexpected roles for chromatin. Nature reviews. Genetics 14, 347-359 (2013). Lewis, P.N., Lukiw, W.J., De Boni, U. & McLachlan, D.R. Changes in chromatin structure associated with Alzheimer's disease. Journal of neurochemistry 37, 11931202 (1981). Lubin, F.D., Gupta, S., Parrish, R.R., Grissom, N.M. & Davis, R.L. Epigenetic Mechanisms: Critical Contributors to Long-Term Memory Formation. The Neuroscientist : a review journal bringing neurobiology, neurology and psychiatry (2011). Kim, T.K., et al. Widespread transcription at neuronal activity-regulated enhancers. Nature 465, 182-187 (2010). Hargreaves, D.C., Horng, T. & Medzhitov, R. Control of inducible gene expression by signal-dependent transcriptional elongation. Cell 138, 129-145 (2009). Koivisto, A.M., et al. Screening of mutations in the PHF8 gene and identification of a novel mutation in a Finnish family with XLMR and cleft lip/cleft palate. Clinical genetics 72, 145-149 (2007). Kim, K.K., Kim, Y.C., Adelstein, R.S. & Kawamoto, S. Fox-3 and PSF interact to activate neural cell-specific alternative splicing. Nucleic acids research 39, 3064-3078 (2011). Antunes-Martins, A., Mizuno, K., Irvine, E.E., Lepicard, E.M. & Giese, K.P. Sexdependent up-regulation of two splicing factors, Psf and Srp20, during hippocampal memory formation. Learn Mem 14, 693-702 (2007). Van de Ven, T.J., VanDongen, H.M. & VanDongen, A.M. The nonkinase phorbol ester receptor α1-chimerin binds the NMDA receptor NR2A subunit and regulates dendritic spine density. The Journal of neuroscience 25, 9488-9496 (2005). Ruiz, S., et al. Generation of a drug-inducible reporter system to study cell reprogramming in human cells. The Journal of biological chemistry 287, 4076740778 (2012). Ma, H., Liu, Q., Diamond, S.L. & Pierce, E.A. Mouse embryonic stem cells efficiently lipofected with nuclear localization peptide result in a high yield of chimeric mice and retain germline transmission potency. Methods 33, 113-120 (2004). Shechter, D., Dormann, H.L., Allis, C.D. & Hake, S.B. Extraction, purification and analysis of histones. Nature protocols 2, 1445-1457 (2007). Tycon, M.A., Daddysman, M.K. & Fecko, C.J. RNA Polymerase II Subunits Exhibit a Broad Distribution of Macromolecular Assembly States in the Interchromatin Space of Cell Nuclei. The journal of physical chemistry. B 118, 423-433 (2014). Politz, J.C., Tuft, R.A., Pederson, T. & Singer, R.H. Movement of nuclear poly(A) RNA throughout the interchromatin space in living cells. Current biology : CB 9, 285-291 (1999). Eskiw, C.H., Rapp, A., Carter, D.R. & Cook, P.R. RNA polymerase II activity is located on the surface of protein-rich transcription factories. Journal of cell science 121, 1999-2007 (2008). Fredriksson, S., et al. Protein detection using proximity-dependent DNA ligation assays. Nature biotechnology 20, 473-477 (2002). Rice, J.C. & Allis, C.D. Histone methylation versus histone acetylation: new insights into epigenetic regulation. Current opinion in cell biology 13, 263-273 (2001). Yu, L., et al. Structural insights into a novel histone demethylase PHF8. Cell research 20, 166-173 (2010). Kimura, A. & Horikoshi, M. Tip60 acetylates six lysines of a specific class in core histones in vitro. Genes Cells 3, 789-800 (1998). 130 121. 122. 123. 124. 125. 126. 127. 128. 129. 130. 131. 132. 133. 134. 135. 136. 137. 138. 139. 140. Loenarz, C., et al. PHF8, a gene associated with cleft lip/palate and mental retardation, encodes for an Nepsilon-dimethyl lysine demethylase. Hum Mol Genet 19, 217-222 (2010). Suganuma, T. & Workman, J.L. Features of the PHF8/KIAA1718 histone demethylase. Cell research 20, 861-862 (2010). Zhu, Z., et al. PHF8 is a histone H3K9me2 demethylase regulating rRNA synthesis. Cell research 20, 794-801 (2010). Kramer, J.M., et al. Epigenetic regulation of learning and memory by Drosophila EHMT/G9a. PLoS Biol 9, e1000569 (2011). Zlatanova, J. & Leuba, S.H. Chromatin Structure and Dynamics: State-of-the-art, (Elsevier, 2004). Ma, D.K., et al. Neuronal activity-induced Gadd45b promotes epigenetic DNA demethylation and adult neurogenesis. Science 323, 1074-1077 (2009). Martinowich, K., et al. DNA methylation-related chromatin remodeling in activitydependent BDNF gene regulation. Science 302, 890-893 (2003). Arnold, F.J., et al. Microelectrode array recordings of cultured hippocampal networks reveal a simple model for transcription and protein synthesis-dependent plasticity. J Physiol 564, 3-19 (2005). Chwang, W.B., Arthur, J.S., Schumacher, A. & Sweatt, J.D. The nuclear kinase mitogen- and stress-activated protein kinase regulates hippocampal chromatin remodeling in memory formation. J Neurosci 27, 12732-12742 (2007). Wittmann, M., et al. Synaptic activity induces dramatic changes in the geometry of the cell nucleus: interplay between nuclear structure, histone H3 phosphorylation, and nuclear calcium signaling. J Neurosci 29, 14687-14700 (2009). Soriano, F.X., Papadia, S., Bell, K.F. & Hardingham, G.E. Role of histone acetylation in the activity-dependent regulation of sulfiredoxin and sestrin 2. Epigenetics 4, 152158 (2009). Tea, J.S. & Luo, L. The chromatin remodeling factor Bap55 functions through the TIP60 complex to regulate olfactory projection neuron dendrite targeting. Neural Dev 6, (2011). Choudhary, C., et al. Lysine acetylation targets protein complexes and co-regulates major cellular functions. Science 325, 834-840 (2009). Peixoto, L. & Abel, T. The role of histone acetylation in memory formation and cognitive impairments. Neuropsychopharmacology 38, 62-76 (2013). Alarcon, J.M., et al. Chromatin acetylation, memory, and LTP are impaired in CBP+/mice: a model for the cognitive deficit in Rubinstein-Taybi syndrome and its amelioration. Neuron 42, 947-959 (2004). Bousiges, O., et al. Detection of histone acetylation levels in the dorsal hippocampus reveals early tagging on specific residues of H2B and H4 histones in response to learning. PLoS One 8, e57816 (2013). Roth, T.L. & Sweatt, J.D. Regulation of chromatin structure in memory formation. Current opinion in neurobiology 19, 336-342 (2009). Vogel-Ciernia, A. & Wood, M.A. Neuron-specific chromatin remodeling: A missing link in epigenetic mechanisms underlying synaptic plasticity, memory, and intellectual disability disorders. Neuropharmacology (2013). Crosio, C., Heitz, E., Allis, C.D., Borrelli, E. & Sassone-Corsi, P. Chromatin remodeling and neuronal response: multiple signaling pathways induce specific histone H3 modifications and early gene expression in hippocampal neurons. J Cell Sci 116, 4905-4914 (2003). Jarome, T.J. & Lubin, F.D. Histone lysine methylation: critical regulator of memory and behavior. Rev Neurosci 24, 375-387 (2013). 131 141. 142. 143. 144. 145. 146. 147. 148. 149. 150. 151. 152. 153. 154. 155. 156. 157. 158. 159. Allfrey, V.G., Faulkner, R. & Mirsky, A.E. Acetylation and Methylation of Histones and Their Possible Role in the Regulation of Rna Synthesis. Proc Natl Acad Sci U S A 51, 786-794 (1964). Latham, J.A. & Dent, S.Y. Cross-regulation of histone modifications. Nature structural & molecular biology 14, 1017-1024 (2007). Nightingale, K.P., et al. Cross-talk between histone modifications in response to histone deacetylase inhibitors - MLL4 links histone H3 acetylation and histone H3K4 methylation. Journal of Biological Chemistry 282, 4408-4416 (2007). Zhang, K., Siino, J.S., Jones, P.R., Yau, P.M. & Bradbury, E.M. A mass spectrometric "Western blot" to evaluate the correlations between histone methylation and histone acetylation. Proteomics 4, 3765-3775 (2004). Tsankova, N.M., Kumar, A. & Nestler, E.J. Histone modifications at gene promoter regions in rat hippocampus after acute and chronic electroconvulsive seizures. J Neurosci 24, 5603-5610 (2004). Qi, H.H., et al. Histone H4K20/H3K9 demethylase PHF8 regulates zebrafish brain and craniofacial development. Nature 466, 503-507 (2010). Sapountzi, V., Logan, I.R. & Robson, C.N. Cellular functions of TIP60. The international journal of biochemistry & cell biology 38, 1496-1509 (2006). Cao, X. & Sudhof, T.C. A transcriptionally [correction of transcriptively] active complex of APP with Fe65 and histone acetyltransferase Tip60. Science 293, 115-120 (2001). Kennedy, P.J., et al. Class I HDAC inhibition blocks cocaine-induced plasticity by targeted changes in histone methylation. Nat Neurosci 16, 434-440 (2013). Margaritis, T. & Holstege, F.C. Poised RNA polymerase II gives pause for thought. Cell 133, 581-584 (2008). Otmakhov, N., et al. Forskolin-induced LTP in the CA1 hippocampal region is NMDA receptor dependent. J Neurophysiol 91, 1955-1962 (2004). Chwang, W.B., O'Riordan, K.J., Levenson, J.M. & Sweatt, J.D. ERK/MAPK regulates hippocampal histone phosphorylation following contextual fear conditioning. Learn Mem 13, 322-328 (2006). Levenson, J.M., et al. Regulation of histone acetylation during memory formation in the hippocampus. J Biol Chem 279, 40545-40559 (2004). Duan, Q., Chen, H., Costa, M. & Dai, W. Phosphorylation of H3S10 blocks the access of H3K9 by specific antibodies and histone methyltransferase. Implication in regulating chromatin dynamics and epigenetic inheritance during mitosis. J Biol Chem 283, 33585-33590 (2008). Pirooznia, S.K., et al. Tip60 HAT activity mediates APP induced lethality and apoptotic cell death in the CNS of a Drosophila Alzheimer's disease model. PLoS One 7, e41776 (2012). Johnson, A.A., Sarthi, J., Pirooznia, S.K., Reube, W. & Elefant, F. Increasing Tip60 HAT levels rescues axonal transport defects and associated behavioral phenotypes in a Drosophila Alzheimer's disease model. J Neurosci 33, 7535-7547 (2013). Xu, S., et al. Epigenetic Control of Learning and Memory in Drosophila by Tip60 HAT Action. Genetics (2014). Halkidou, K., Logan, I.R., Cook, S., Neal, D.E. & Robson, C.N. Putative involvement of the histone acetyltransferase Tip60 in ribosomal gene transcription. Nucleic acids research 32, 1654-1665 (2004). Mahajan, M.A. & Stanley, F.M. Insulin-activated Elk-1 recruits the TIP60/NuA4 complex to increase prolactin gene transcription. Molecular and cellular endocrinology 382, 159-169 (2014). 132 160. 161. 162. 163. 164. 165. 166. 167. 168. 169. 170. 171. 172. 173. 174. 175. 176. 177. 178. 179. 180. 181. 182. Wang, Z., et al. Genome-wide mapping of HATs and HDACs reveals distinct functions in active and inactive genes. Cell 138, 1019-1031 (2009). Zippo, A., et al. Histone crosstalk between H3S10ph and H4K16ac generates a histone code that mediates transcription elongation. Cell 138, 1122-1136 (2009). Li, G., et al. Combinatorial H3K9acS10ph histone modification in IgH locus S regions targets 14-3-3 adaptors and AID to specify antibody class-switch DNA recombination. Cell reports 5, 702-714 (2013). Macdonald, N., et al. Molecular basis for the recognition of phosphorylated and phosphoacetylated histone h3 by 14-3-3. Mol Cell 20, 199-211 (2005). Winter, S., et al. 14-3-3 proteins recognize a histone code at histone H3 and are required for transcriptional activation. The EMBO journal 27, 88-99 (2008). Karam, C.S., Kellner, W.A., Takenaka, N., Clemmons, A.W. & Corces, V.G. 14-3-3 mediates histone cross-talk during transcription elongation in Drosophila. PLoS genetics 6, e1000975 (2010). Peleg, S., et al. Altered histone acetylation is associated with age-dependent memory impairment in mice. Science 328, 753-756 (2010). Levenson, J.M. & Sweatt, J.D. Epigenetic mechanisms in memory formation. Nature reviews. Neuroscience 6, 108-118 (2005). Ong, C.T. & Corces, V.G. Enhancer function: new insights into the regulation of tissue-specific gene expression. Nature reviews. Genetics 12, 283-293 (2011). Rosonina, E., et al. Role for PSF in mediating transcriptional activator-dependent stimulation of pre-mRNA processing in vivo. Mol Cell Biol 25, 6734-6746 (2005). Emili, A., et al. Splicing and transcription-associated proteins PSF and p54nrb/nonO bind to the RNA polymerase II CTD. RNA 8, 1102-1111 (2002). Lowery, L.A., Rubin, J. & Sive, H. Whitesnake/sfpq is required for cell survival and neuronal development in the zebrafish. Developmental dynamics : an official publication of the American Association of Anatomists 236, 1347-1357 (2007). Ke, Y.D., et al. Tau-mediated nuclear depletion and cytoplasmic accumulation of SFPQ in Alzheimer's and Pick's disease. PLoS One 7, e35678 (2012). Bentley, D.L. Coupling mRNA processing with transcription in time and space. Nature reviews. Genetics 15, 163-175 (2014). Lee, K.M. & Tarn, W.Y. Coupling pre-mRNA processing to transcription on the RNA factory assembly line. RNA biology 10, 380-390 (2013). Ameur, A., et al. Total RNA sequencing reveals nascent transcription and widespread co-transcriptional splicing in the human brain. Nature structural & molecular biology 18, 1435-1440 (2011). Huang, B., Jones, S.A., Brandenburg, B. & Zhuang, X. Whole-cell 3D STORM reveals interactions between cellular structures with nanometer-scale resolution. Nature methods 5, 1047-1052 (2008). Rust, M.J., Bates, M. & Zhuang, X. Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM). Nature methods 3, 793-795 (2006). Smolle, M. & Venkatesh, S. Transcription Through Chromatin. in Fundamentals of Chromatin (eds. Workman, J.L. & Abmayr, S.M.) 427-489 (Springer New York, 2014). Vermeulen, M., et al. Quantitative interaction proteomics and genome-wide profiling of epigenetic histone marks and their readers. Cell 142, 967-980 (2010). Perner, J. & Chung, H.-R. Chromatin signaling and transcription initiation. Frontiers in Life Science 7, 22-30 (2013). Davis, H.P. & Squire, L.R. Protein synthesis and memory: a review. Psychol Bull 96, 518-559 (1984). Alberini, C.M. Transcription factors in long-term memory and synaptic plasticity. Physiol Rev 89, 121-145 (2009). 133 183. 184. 185. 186. 187. 188. 189. 190. 191. 192. 193. 194. 195. 196. 197. 198. 199. 200. 201. 202. 203. Link, W., et al. Somatodendritic expression of an immediate early gene is regulated by synaptic activity. Proc Natl Acad Sci U S A 92, 5734-5738 (1995). Lyford, G.L., et al. Arc, a growth factor and activity-regulated gene, encodes a novel cytoskeleton-associated protein that is enriched in neuronal dendrites. Neuron 14, 433-445 (1995). Guzowski, J.F., McNaughton, B.L., Barnes, C.A. & Worley, P.F. Environment-specific expression of the immediate-early gene Arc in hippocampal neuronal ensembles. Nat Neurosci 2, 1120-1124 (1999). Chawla, M.K., et al. Sparse, environmentally selective expression of Arc RNA in the upper blade of the rodent fascia dentata by brief spatial experience. Hippocampus 15, 579-586 (2005). Bloomer, W.A., VanDongen, H.M. & VanDongen, A.M. Arc/Arg3. translation is controlled by convergent N-methyl-D-aspartate and Gs-coupled receptor signaling pathways. Journal of Biological Chemistry 283, 582-592 (2008). Chowdhury, S., et al. Arc/Arg3.1 interacts with the endocytic machinery to regulate AMPA receptor trafficking. Neuron 52, 445-459 (2006). Shepherd, J.D., et al. Arc/Arg3.1 mediates homeostatic synaptic scaling of AMPA receptors. Neuron 52, 475-484 (2006). Okuno, H., et al. Inverse synaptic tagging of inactive synapses via dynamic interaction of Arc/Arg3.1 with CaMKIIbeta. Cell 149, 886-898 (2012). Mikuni, T., et al. Arc/Arg3.1 is a postsynaptic mediator of activity-dependent synapse elimination in the developing cerebellum. Neuron 78, 1024-1035 (2013). Torok, D., Ching, R.W. & Bazett-Jones, D.P. PML nuclear bodies as sites of epigenetic regulation. Front Biosci 14, 1325-1336 (2009). Zovkic, I.B., Guzman-Karlsson, M.C. & Sweatt, J.D. Epigenetic regulation of memory formation and maintenance. Learn Mem 20, 61-74 (2013). Abel, T. & Zukin, R.S. Epigenetic targets of HDAC inhibition in neurodegenerative and psychiatric disorders. Curr Opin Pharmacol 8, 57-64 (2008). Eskiw, C.H. & Bazett-Jones, D.P. The promyelocytic leukemia nuclear body: sites of activity? Biochem Cell Biol 80, 301-310 (2002). von Mikecz, A., Zhang, S., Montminy, M., Tan, E.M. & Hemmerich, P. CREB-binding protein (CBP)/p300 and RNA polymerase II colocalize in transcriptionally active domains in the nucleus. J Cell Biol 150, 265-273 (2000). Korzus, E., Rosenfeld, M.G. & Mayford, M. CBP histone acetyltransferase activity is a critical component of memory consolidation. Neuron 42, 961-972 (2004). Barrett, R.M., et al. Hippocampal focal knockout of CBP affects specific histone modifications, long-term potentiation, and long-term memory. Neuropsychopharmacology 36, 1545-1556 (2011). Schermelleh, L., Spada, F. & Leonhardt, H. Visualization and measurement of DNA methyltransferase activity in living cells. Curr Protoc Cell Biol Chapter 22, Unit 22 12 (2008). Gustafsson, M.G. Super-resolution light microscopy goes live. Nat Methods 5, 385387 (2008). Schindelin, J., et al. Fiji: an open-source platform for biological-image analysis. Nat Methods 9, 676-682 (2012). Hardingham, G.E., Fukunaga, Y. & Bading, H. Extrasynaptic NMDARs oppose synaptic NMDARs by triggering CREB shut-off and cell death pathways. Nat Neurosci 5, 405414 (2002). Ogura, T. & Wilkinson, A.J. AAA+ superfamily ATPases: common structure--diverse function. Genes Cells 6, 575-597 (2001). 134 204. 205. 206. 207. 208. 209. 210. 211. 212. 213. 214. 215. 216. 217. 218. 219. 220. 221. 222. Qiu, Z. & Ghosh, A. A calcium-dependent switch in a CREST-BRG1 complex regulates activity-dependent gene expression. Neuron 60, 775-787 (2008). Cai, Y., et al. The mammalian YL1 protein is a shared subunit of the TRRAP/TIP60 histone acetyltransferase and SRCAP complexes. J Biol Chem 280, 13665-13670 (2005). Tapias, A., et al. Trrap-dependent histone acetylation specifically regulates cell-cycle gene transcription to control neural progenitor fate decisions. Cell Stem Cell 14, 632643 (2014). Doucas, V. The promyelocytic (PML) nuclear compartment and transcription control. Biochem Pharmacol 60, 1197-1201 (2000). Cheng, Z., et al. Functional characterization of TIP60 sumoylation in UV-irradiated DNA damage response. Oncogene 27, 931-941 (2008). Legube, G., et al. Role of the histone acetyl transferase Tip60 in the p53 pathway. J Biol Chem 279, 44825-44833 (2004). Blalock, E.M., et al. Gene microarrays in hippocampal aging: statistical profiling identifies novel processes correlated with cognitive impairment. J Neurosci 23, 38073819 (2003). Bousiges, O., et al. Spatial memory consolidation is associated with induction of several lysine-acetyltransferase (histone acetyltransferase) expression levels and H2B/H4 acetylation-dependent transcriptional events in the rat hippocampus. Neuropsychopharmacology 35, 2521-2537 (2010). Ikura, T., et al. Involvement of the TIP60 histone acetylase complex in DNA repair and apoptosis. Cell 102, 463-473 (2000). Squatrito, M., Gorrini, C. & Amati, B. Tip60 in DNA damage response and growth control: many tricks in one HAT. Trends in cell biology 16, 433-442 (2006). Sun, Y., Jiang, X., Chen, S., Fernandes, N. & Price, B.D. A role for the Tip60 histone acetyltransferase in the acetylation and activation of ATM. Proceedings of the National Academy of Sciences of the United States of America 102, 13182-13187 (2005). Kamine, J., Elangovan, B., Subramanian, T., Coleman, D. & Chinnadurai, G. Identification of a cellular protein that specifically interacts with the essential cysteine region of the HIV-1 Tat transactivator. Virology 216, 357-366 (1996). Yamamoto, T. & Horikoshi, M. Novel substrate specificity of the histone acetyltransferase activity of HIV-1-Tat interactive protein Tip60. Journal of Biological Chemistry 272, 30595-30598 (1997). Kimura, A. & Horikoshi, M. Tip60 acetylates six lysines of a specific class in core histones in vitro. Genes to Cells 3, 789-800 (1998). Doyon, Y., et al. Structural and functional conservation of the NuA4 histone acetyltransferase complex from yeast to humans. Molecular & Cellular Biology 24, 1884-1896 (2004). Cai, Y., et al. Identification of new subunits of the multiprotein mammalian TRRAP/TIP60-containing histone acetyltransferase complex. J Biol Chem 278, 4273342736 (2003). Pirooznia, S.K., Chiu, K., Chan, M.T., Zimmerman, J.E. & Elefant, F. Epigenetic regulation of axonal growth of Drosophila pacemaker cells by histone acetyltransferase tip60 controls sleep. Genetics 192, 1327-1345 (2012). Muller, T., et al. A ternary complex consisting of AICD, FE65, and TIP60 downregulates Stathmin1. Biochimica et biophysica acta 1834, 387-394 (2013). Cao, X. & Sudhof, T.C. A transcriptionally active complex of APP with Fe65 and histone acetyltransferase Tip60. Science 293, 115-120 (2001). 135 223. 224. 225. 226. 227. 228. 229. 230. 231. 232. 233. 234. 235. 236. 237. 238. 239. 240. 241. Wu, J., et al. Arc/Arg3.1 regulates an endosomal pathway essential for activitydependent beta-amyloid generation. Cell 147, 615-628 (2011). Rudinskiy, N., et al. Orchestrated experience-driven Arc responses are disrupted in a mouse model of Alzheimer's disease. Nat Neurosci 15, 1422-1429 (2012). Stilling, R.M. & Fischer, A. The role of histone acetylation in age-associated memory impairment and Alzheimer's disease. Neurobiology of learning and memory 96, 1926 (2011). Habets, A.M., Van Dongen, A.M., Van Huizen, F. & Corner, M.A. Spontaneous neuronal firing patterns in fetal rat cortical networks during development in vitro: a quantitative analysis. Exp Brain Res 69, 43-52 (1987). Irie, Y., et al. Molecular cloning and characterization of Amida, a novel protein which interacts with a neuron-specific immediate early gene product arc, contains novel nuclear localization signals, and causes cell death in cultured cells. J Biol Chem 275, 2647-2653 (2000). Fischer, A., Sananbenesi, F., Wang, X., Dobbin, M. & Tsai, L.H. Recovery of learning and memory is associated with chromatin remodelling. Nature 447, 178-182 (2007). Levenson, J.M. & Sweatt, J.D. Epigenetic mechanisms: a common theme in vertebrate and invertebrate memory formation. Cell Mol Life Sci 63, 1009-1016 (2006). Vecsey, C.G., et al. Histone deacetylase inhibitors enhance memory and synaptic plasticity via CREB:CBP-dependent transcriptional activation. J Neurosci 27, 61286140 (2007). Park, C.S., Rehrauer, H. & Mansuy, I.M. Genome-wide analysis of H4K5 acetylation associated with fear memory in mice. BMC genomics 14, 539 (2013). Liu, W., et al. PHF8 mediates histone H4 lysine 20 demethylation events involved in cell cycle progression. Nature 466, 508-512 (2010). Giorgi, C. & Moore, M.J. The nuclear nurture and cytoplasmic nature of localized mRNPs. Semin Cell Dev Biol 18, 186-193 (2007). Gerstein, H., O'Riordan, K., Osting, S., Schwarz, M. & Burger, C. Rescue of synaptic plasticity and spatial learning deficits in the hippocampus of Homer1 knockout mice by recombinant Adeno-associated viral gene delivery of Homer1c. Neurobiol Learn Mem 97, 17-29 (2012). Heessen, S. & Fornerod, M. The inner nuclear envelope as a transcription factor resting place. EMBO Rep 8, 914-919 (2007). Crepaldi, L., et al. Binding of TFIIIC to sine elements controls the relocation of activity-dependent neuronal genes to transcription factories. PLoS Genet 9, e1003699 (2013). Ying, S.W., et al. Brain-derived neurotrophic factor induces long-term potentiation in intact adult hippocampus: requirement for ERK activation coupled to CREB and upregulation of Arc synthesis. J Neurosci 22, 1532-1540 (2002). Katoh-Semba, R., et al. Activation of p38 mitogen-activated protein kinase is required for in vivo brain-derived neurotrophic factor production in the rat hippocampus. Neuroscience 163, 352-361 (2009). Coffey, E.T. Nuclear and cytosolic JNK signalling in neurons. Nat Rev Neurosci 15, 285-299 (2014). Hagenston, A.M. & Bading, H. Calcium signaling in synapse-to-nucleus communication. Cold Spring Harb Perspect Biol 3, a004564 (2011). Dardenne, E., et al. RNA helicases DDX5 and DDX17 dynamically orchestrate transcription, miRNA, and splicing programs in cell differentiation. Cell Rep 7, 19001913 (2014). 136 242. 243. 244. 245. 246. 247. 248. 249. 250. 251. Wang, W.Y., et al. Interaction of FUS and HDAC1 regulates DNA damage response and repair in neurons. Nat Neurosci 16, 1383-1391 (2013). Hashimoto, S., et al. MED23 mutation links intellectual disability to dysregulation of immediate early gene expression. Science 333, 1161-1163 (2011). Glozak, M.A., Sengupta, N., Zhang, X. & Seto, E. Acetylation and deacetylation of non-histone proteins. Gene 363, 15-23 (2005). Patel, J.H., et al. The c-MYC oncoprotein is a substrate of the acetyltransferases hGCN5/PCAF and TIP60. Mol Cell Biol 24, 10826-10834 (2004). Sykes, S.M., et al. Acetylation of the p53 DNA-binding domain regulates apoptosis induction. Mol Cell 24, 841-851 (2006). Gaughan, L., Logan, I.R., Cook, S., Neal, D.E. & Robson, C.N. Tip60 and histone deacetylase regulate androgen receptor activity through changes to the acetylation status of the receptor. J Biol Chem 277, 25904-25913 (2002). Tischmeyer, W. & Grimm, R. Activation of immediate early genes and memory formation. Cell Mol Life Sci 55, 564-574 (1999). Ren, M., Cao, V., Ye, Y., Manji, H.K. & Wang, K.H. Arc regulates experiencedependent persistent firing patterns in frontal cortex. J Neurosci 34, 6583-6595 (2014). von Hertzen, L.S. & Giese, K.P. Memory reconsolidation engages only a subset of immediate-early genes induced during consolidation. J Neurosci 25, 1935-1942 (2005). Rogerson, T., et al. Synaptic tagging during memory allocation. Nat Rev Neurosci 15, 157-169 (2014). 137 Appendix A. Publications accepted or under review / in preparation 1. White, C.A., Oey, N.E., and Emili, A. (2009). Global quantitative proteomic profiling through 18O-labeling in combination with MS/MS spectra analysis. J Proteome Res 8, 3653-3665. 2. Deng, Z.D., McClintock, S.M., Oey, N.E., Luber, B., and Lisanby, S.H. (2014). Neuromodulation for mood and memory: from the engineering bench to the patient bedside. Curr Opin Neurobiol 30C, 38-43. 3. Wee, C.L.*, Teo, S.*, Oey, N.E.*, Wright, G.D., VanDongen, H.M., and VanDongen, A.M. (2014). Nuclear Arc Interacts with the Histone Acetyltransferase Tip60 to Modify H4K12 Acetylation. eneuro, ENEURO. 0019-0014.2014. (*=co-first authors) 4. Oey, N.E., Leung, H.W., Rajaram, E., Beuerman, R.W., Zhou, L., VanDongen, H.M., and VanDongen, A.M. (2014). A Dual-Function Chromatin Modifying Complex Regulates Early Transcription of Arc. eneuro, ENEURO. 0020-14.2015 5. Oey, N.E., Lo, Y.L. (2014). Migraine with Multiple Sensory Auras. J Headache and Pain. Under Revision. 6. Oey, N.E., Leung, H.W., Beuerman, R.W., Zhou, L., VanDongen, H.M., and VanDongen, A.M. (2014). PHF8 is an Early Epigenetic Gateway of Memory Consolidation. Nat Neurosci. In Preparation. 7. Samuel, G.S., Choo, M., Oey, N.E., Ju, H., Chan, W.Y., Kok, S., VanDongen, A.M., Ge, Y., Ng, Y.S. (2014). The Effectiveness of Combining Levodopa Neuromodulation and Virtual Reality Based Therapy in Acute Stroke Rehabilitation: Preliminary Data from a Pilot Randomized Controlled Trial. Experimental Brain Research. In Preparation. 138 [...]... store information through modification of the strength of the synapses that link them Although this may serve as a plausible explanation for working memory that lasts several seconds at maximum, the fact that long- term memory as we know it may persist for months and even years point toward the existence of another, more long- lasting mechanism 4 1.II Molecular Formation 1.II .A Mechanisms of Long- Term Memory. .. cellular correlate of memory, Long Term Potentiation, has an early and a late phase, which parallels the timeline of short -term memory acquisition and long- term memory consolidation, epigenetic events can also be divided into early events that occur within minutes of neuronal activation, and late events that take a longer time to develop In this model, early epigenetic events may underlie earlyphase... demethylase PHF8 and the acetyltransferase TIP60 as a key regulator of the activity- induced expression of Arc, an important mediator of synaptic plasticity Clinically, mutations in PHF8 cause X-linked mental retardation while TIP60 has been implicated in the pathogenesis of Alzheimer’s disease Within minutes of increased synaptic activity, this dual function complex is rapidly recruited to specific neuronal. .. to a neuronal spectrin betaSpIVSigma5 and associates with Promyelocytic Leukemia (PML) bodies, which are major sites of transcriptional regulation68 Subsequent to this, a recent study has shown that ARC protein has both an export signal that allows it to exit into the cytoplasm as well as a retention domain and localization signal that targets it to the nucleus where it may play a role in the PML -dependent. .. known, as in the role of p300/CBP in Rubinstein-Taybi syndromic mental retardation, the exact histone lysines affected and the roles they play in mediating memory formation defects are not yet known75 One particular epigenetic regulator, the HAT enzyme TIP60 (HIV Tat interactive Protein, 60 kDa, also known as KAT5), has emerged as an important effector of neuronal plasticity and memory formation7 6 Initially... circumjacent to a training period does not affect short -term memory but abolishes the ability of animals to form long- term memory, indicating that there is a window of time during which gene transcription and protein synthesis is crucial for the formation of long- term memory1 2-14 It has therefore been known for several decades that there is a fundamental mechanistic difference in the way short- and long- term. .. situating this chromatin modifying complex at the crossroads of transcriptional activation These findings point toward a mechanism by which an epigenetic pathway can regulate neuronal activity- dependent gene transcription, which has implications in the development of novel therapeutics for disorders of learning and memory 2.II Introduction Activity- dependent gene transcription, a pre-requisite for memory. .. LTP and short -term plasticity while late events may be responsible for late-phase LTP and memory consolidation 10 1.II.C Arc – a regulator of synaptic memory The leading mechanisms to explain memory at the level of neuronal synapses are Long Term Potentiation (LTP) and Long Term Depression (LTD), which are defined by the measurable, persistent increase (for LTP) or decrease (for LTD) in the strength of. .. nature of these epigenetic regulators is still obscure In this chapter, I report that PHF8 cooperates with TIP60 in an activitydependent manner to enable the rapid induction of the immediate-early gene Arc by specifically regulating H3K9acS10P, a dual-chromatin mark that is required for transcriptional activation As no direct interaction between a demethylase and an acetyltransferase has yet been reported,... crucial for memory as well, apparently by producing a diminished response to a stimulus37 As such, both early and late LTP and LTD are directly paralleled by many behavioral correlates of learning and memory And just like in short vs long- term memory, only the late phase of LTP and LTD requires novel induction of gene and protein synthesis38,39 Although many proteins have so far been implicated in synaptic . Learning and Memory A. A short history 1 B. Classifications of memory 3 II. Molecular Mechanisms of Long- Term Memory Formation 5 A. Overview: plasticity and activity 5 B. Epigenetics as a. role of the Alzheimer’s Disease-associated epigenetic enzyme TIP60 and an X-linked Mental Retardation (XLMR)-associated protein PHF8 in the rapid neuronal activity- dependent transcription of ARC,. reveals a major uniting mechanism of mRNA metabolism, transcriptional regulation, and mRNA splicing. 115 Figure 32: A graphical abstract of activity- dependent DNA, histone, RNA, and protein changes