title: Identification of a novel distal regulatory element of the human Neuroglobin gene by the chromosome conformation capture approach.
abstract: Neuroglobin (NGB) is predominantly expressed in the brain and retina. Studies suggest that NGB exerts protective effects to neuronal cells and is implicated in reducing the severity of stroke and Alzheimer's disease. However, little is known about the mechanisms which regulate the cell type-specific expression of the gene. In this study, we hypothesized that distal regulatory elements (DREs) are involved in optimal expression of the NGB gene. By chromosome conformation capture we identified two novel DREs located -70 kb upstream and +100 kb downstream from the NGB gene. ENCODE database showed the presence of DNaseI hypersensitive and transcription factors binding sites in these regions. Further analyses using luciferase reporters and chromatin immunoprecipitation suggested that the -70 kb region upstream of the NGB gene contained a neuronal-specific enhancer and GATA transcription factor binding sites. Knockdown of GATA-2 caused NGB expression to drop dramatically, indicating GATA-2 as an essential transcription factor for the activation of NGB expression. The crucial role of the DRE in NGB expression activation was further confirmed by the drop in NGB level after CRISPR-mediated deletion of the DRE. Taken together, we show that the NGB gene is regulated by a cell type-specific loop formed between its promoter and the novel DRE.
authors.affiliation: School of Biological Sciences, The University of Hong Kong, Pokfulam Road, Hong Kong S.A.R., China.
authors.forename: Kin Tung
authors.initials: KT
authors.lastname: Tam
authors.affiliation: Gene Control Mechanisms and Disease Group, Department of Medicine, Division of Brain Sciences and MRC Clinical Sciences Centre, Imperial College School of Medicine, London W12 0NN, United Kingdom.
authors.forename: Ping Kei
authors.initials: PK
authors.lastname: Chan
authors.affiliation: School of Biological Sciences, The University of Hong Kong, Pokfulam Road, Hong Kong S.A.R., China.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.affiliation: School of Biological Sciences, The University of Hong Kong, Pokfulam Road, Hong Kong S.A.R., China.|Gene Control Mechanisms and Disease Group, Department of Medicine, Division of Brain Sciences and MRC Clinical Sciences Centre, Imperial College School of Medicine, London W12 0NN, United Kingdom.|Department of Paediatrics and Adolescent Medicine, The University of Hong Kong, Pokfulam Road, Hong Kong S.A.R., China.
authors.forename: Pui Pik
authors.initials: PP
authors.lastname: Law
authors.affiliation: School of Biological Sciences, The University of Hong Kong, Pokfulam Road, Hong Kong S.A.R., China.|School of Professional and Continuing Education (HKU SPACE), The University of Hong Kong, Pokfulam Road, Hong Kong S.A.R., China.
authors.forename: Zhipeng
authors.initials: Z
authors.lastname: Tian
authors.affiliation: Department of Paediatrics and Adolescent Medicine, The University of Hong Kong, Pokfulam Road, Hong Kong S.A.R., China.
authors.forename: Godfrey Chi
authors.initials: GC
authors.lastname: Fung Chan
authors.affiliation: Department of Cell Biology, Erasmus MC, PO Box 2040, 3000 CA Rotterdam, The Netherlands kctanun@hkucc.hku.hk.
authors.forename: Sjaak
authors.initials: S
authors.lastname: Philipsen
authors.affiliation: Gene Control Mechanisms and Disease Group, Department of Medicine, Division of Brain Sciences and MRC Clinical Sciences Centre, Imperial College School of Medicine, London W12 0NN, United Kingdom kctanun@hkucc.hku.hk.
authors.forename: Richard
authors.initials: R
authors.lastname: Festenstein
authors.affiliation: School of Biological Sciences, The University of Hong Kong, Pokfulam Road, Hong Kong S.A.R., China kctanun@hkucc.hku.hk.|School of Professional and Continuing Education (HKU SPACE), The University of Hong Kong, Pokfulam Road, Hong Kong S.A.R., China.
authors.forename: Kian Cheng
authors.initials: KC
authors.lastname: Tan-Un
mesh_terms: D001665:Binding Sites; D064113:CRISPR-Cas Systems; D045744:Cell Line, Tumor; D002883:Chromosomes, Human, Pair 14 / Q000737:chemistry*; D003850:Deoxyribonuclease I / Q000235:genetics / Q000378:metabolism; D050989:GATA2 Transcription Factor / Q000235:genetics* / Q000378:metabolism; D000072669:Gene Editing; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D005914:Globins / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D020014:K562 Cells; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D000078787:Neuroglobin; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D009928:Organ Specificity; D011485:Protein Binding; D000094704:RNA, Guide, CRISPR-Cas Systems / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D050436:Regulatory Elements, Transcriptional; D015398:Signal Transduction
chemical_list: D050989:GATA2 Transcription Factor; C494711:GATA2 protein, human; D009419:Nerve Tissue Proteins; D000078787:Neuroglobin; D000094704:RNA, Guide, CRISPR-Cas Systems; D034741:RNA, Small Interfering; D005914:Globins; D008156:Luciferases; D003850:Deoxyribonuclease I