Modulation of enhancer looping and differential gene targeting by Epstein-Barr virus transcription factors directs cellular reprogramming.
Epstein-Barr virus (EBV) epigenetically reprogrammes B-lymphocytes to drive immortalization and facilitate viral persistence. Host-cell transcription is perturbed principally through the actions of EBV EBNA 2, 3A, 3B and 3C, with cellular genes deregulated by specific combinations of these EBNAs through unknown mechanisms. Comparing human genome binding by these viral transcription factors, we discovered that 25% of binding sites were shared by EBNA 2 and the EBNA 3s and were located predominantly in enhancers. Moreover, 80% of potential EBNA 3A, 3B or 3C target genes were also targeted by EBNA 2, implicating extensive interplay between EBNA 2 and 3 proteins in cellular reprogramming. Investigating shared enhancer sites neighbouring two new targets (WEE1 and CTBP2) we discovered that EBNA 3 proteins repress transcription by modulating enhancer-promoter loop formation to establish repressive chromatin hubs or prevent assembly of active hubs. Re-ChIP analysis revealed that EBNA 2 and 3 proteins do not bind simultaneously at shared sites but compete for binding thereby modulating enhancer-promoter interactions. At an EBNA 3-only intergenic enhancer site between ADAM28 and ADAMDEC1 EBNA 3C was also able to independently direct epigenetic repression of both genes through enhancer-promoter looping. Significantly, studying shared or unique EBNA 3 binding sites at WEE1, CTBP2, ITGAL (LFA-1 alpha chain), BCL2L11 (Bim) and the ADAMs, we also discovered that different sets of EBNA 3 proteins bind regulatory elements in a gene and cell-type specific manner. Binding profiles correlated with the effects of individual EBNA 3 proteins on the expression of these genes, providing a molecular basis for the targeting of different sets of cellular genes by the EBNA 3s. Our results therefore highlight the influence of the genomic and cellular context in determining the specificity of gene deregulation by EBV and provide a paradigm for host-cell reprogramming through modulation of enhancer-promoter interactions by viral transcription factors.
School of Life Sciences, John Maynard-Smith Building, University of Sussex, Falmer, Brighton, United Kingdom.
Michael J
MJ
McClellan
C David
CD
Wood
Opeoluwa
O
Ojeniyi
Tim J
TJ
Cooper
Aditi
A
Kanhere
Aaron
A
Arvey
Helen M
HM
Webb
Richard D
RD
Palermo
Marie L
ML
Harth-Hertle
Bettina
B
Kempkes
Richard G
RG
Jenner
Michelle J
MJ
West
D000429:Alcohol Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D001667:Binding, Competitive; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D065150:Cellular Reprogramming; D056970:Co-Repressor Proteins; D004742:Enhancer Elements, Genetic; D020031:Epstein-Barr Virus Infections / Q000378:metabolism / Q000473:pathology; D019309:Epstein-Barr Virus Nuclear Antigens / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018390:Gene Targeting; D004854:Herpesvirus 4, Human / Q000378:metabolism*; D054884:Host-Pathogen Interactions; D006801:Humans; D008954:Models, Biological; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D018797:Cell Cycle Proteins; D056970:Co-Repressor Proteins; C402641:EBNA-2 protein, Human herpesvirus 4; C508151:EBNA-3A antigen; C422762:EBNA-3B antigen; C498774:EBNA-3C, epstein-barr virus; D019309:Epstein-Barr Virus Nuclear Antigens; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins; D011994:Recombinant Proteins; D012097:Repressor Proteins; D014764:Viral Proteins; D000429:Alcohol Oxidoreductases; C421248:CTBP2 protein, human; D011505:Protein-Tyrosine Kinases; C499349:WEE1 protein, human