Survey
* Your assessment is very important for improving the work of artificial intelligence, which forms the content of this project
Download The NIH Mouse Regulome
Document related concepts
no text concepts found
Transcript
The NIH Mouse Regulome Project ES cells Hemangioblasts Mouse blastocyst E3.5 Substan:al developmental gap (marked epigene:c and transcrip:onal differences) Early streak E10 Hematopoie:c stem cells Ac:vated B lymphocytes C. Diphtheriae Diphtheria toxin A Diphtheria toxin A class switch recombination! DNA breaks! VDJ! IgM! IgG! IgE! IgA! Diphtheria toxin A AID VDJ! IgG! IgE! IgA! 1-‐ Iden:fy all genomic sites occupied by regulatory factors The ES and B cell regulomes DNaseI hypersensi:vity 1-‐ Iden:fy all genomic sites occupied by regulatory factors TF binding DNaseI DNaseI hypersensi:vity 1-‐ Iden:fy all genomic sites occupied by regulatory factors 2-‐ Classify promoters and enhancers DNaseI hypersensi:vity Map TSS (ENSEMBL) Map Nipbl, Med12, p300 DNaseI Not en:rely overlapping 3-‐ Refine promoter/enhancer classifica:on Enhancers 10 1.5 Enhancers 1.510 Enhancers 10 1.5 Map H3K4me1 vs H2AZ 90,015 DNaseI domains 1.0 H3K4me1 H3K4me1 H3K4me1 10 1.0 1.0 10 10 Promoters (19%) 0.5 10 0.5 0.510 10 Other (26%) 0 10 0 0 10 10 -0.5 10 -0.5 -0.5 10 10 -1.0 Promoters Promoters Promoters 10 -1.0 -1.0 10 10 -1.0 -0.5 0 0.5 1.0 1.5 2.0 10 10 100.5 10 -1.0 10 -0.5 100 1.0 10 1.5 2.0 -1.0 -0.5 0 10 0.510 1.010 1.510 2.010 10 10 10 10 10 10 10 10 10 H2AZ H2AZ H2AZ Enhancers (55%) Paris Metro Map Opéra Bourse Pyramides Concorde Louvre Bonne Nouvelle Richelieu Rivoli Les Halles Châtelet Musée d’Orsay St-‐Germain des-‐Prés Solférino Sèvres Babylone St-‐Michel Odéon Froissard Hôtel de Ville Bas:lle Notre-‐Dame La Sorbonne Mutualité Temple Sully 4-‐ Map cognate promoter-‐enhancer interac:ons ChIA-‐PET Paris Metro Map Opéra Bourse Pyramides Concorde Louvre Bonne Nouvelle Richelieu Rivoli Les Halles Châtelet Musée d’Orsay St-‐Germain des-‐Prés Solférino Sèvres Babylone St-‐Michel Odéon Froissard Hôtel de Ville Bas:lle Notre-‐Dame La Sorbonne Mutualité Temple Sully 4-‐ Map cognate promoter-‐enhancer interac:ons (83)! PolII looping PolII chroma:n bound (7)! ChIA-‐PET (27)! (41)! (15)! (38)! 5C or HiC studies Background interac:ons 44,398 Func:onal interac:ons 343 (0.8%) Phillips-‐Cremins et al, Cell, 2013 Advantages of ChIA-‐PET 1) Specificity 5C or HiC studies Background interac:ons 44,398 ChIA-‐PET detects long-‐range interac:ons involving ac:ve promoters and enhancers Func:onal interac:ons 343 (0.8%) Phillips-‐Cremins et al, Cell, 2013 Anchored enhancers are H3K27Ac+ (func:onal) Anchored promoters are transcrip:onally ac:ve >90% of ChIA-‐PET anchors within DNaseI islands 2) ChIA-‐PET maps leapfrog interacDons ENCODE: enhancers are assigned to cognate promoters based on proximity ~250kb DHS Pax5 enhancers 3) ChIA-‐PET uncovers new enhancers known enhancers E4 E5 Igκ locus J Jκ Cκ C Rpia κ light chains first NFκB site E1 E2 known enhancers Mfap5 E6 AID Solid phase high-‐throughput Transcrip:on TALEN nuclease Apobec1 DNA break Cd83 (control) Enhancers control PolII recruitment/ac:vity PolII (WT) PolII (E2 KO) Δ 190kb 4) ChIA-‐PET uncovers two kinds of gene clusters 1,231 single-‐promoter clusters Gpr183 cluster Cd83 1 promoter 12 enhancers 72 interac:ons 1,481 mul:ple-‐promoter clusters Rela cluster 66 promoters 398 interac:ons Gene families Does gene clustering influence transcrip:on? Expression (RNA-‐Seq) PolII-‐ Single promoter clusters Mul:ple promoter clusters Clec2d lincRNA E85930 Cd69 Transcrip:on of lincRNAs and their cis-‐targets is ooen coordinated Bcl11a lincRNA E123592 ES cells Promoters anchored in: Sox2 B cells ES cells 5,099 4,673 2,217 ES cells B cells Smarcd2 Tcam1 B cells Gh Cd79b Cd79b Scn4a different enhancers 94% 6% ExcepDon ES cells 5,099 4,673 2,217 B cells Acbd4 Xehim1 Xehim2 same enhancers 4 enhancers (~150kb) * * * * B cells ES cells * * * * * * * 10 enhancers (~350kb) * * * Validate CpG methylated CpG demethylated CH 3 CH 3 CH 3 CH 3 Silent promoters/enhancers Ac:ve promoters/enhancers 100 90 80 70 60 % CpG methyla:on 50 40 30 20 10 0 B cell B cell B cell B cell promoters enhancers promoters enhancers TALENs The enhancer landscape of broadly expressed genes changes during development Changes are mirrored by the dynamics of CpG methyla:on What’s the impact on transcrip:on? Differen:al enhancer usage does impact transcrip:on levels (transcrip:on is commensurate to the number of linked enhancers) loose > 2 enhancers No change gain > 2 enhancers Loose > 2 No change gain > 2 ES cells (4 enhancers) 4 fold increase in transcrip:on B cells (10 enhancers) How do genes like Myc change their enhancer landscape during development? Create comprehensive maps of TF binding in B and ES cells ES cell-‐specific factors? Digital genomic footprinDng B cell-‐specific factors? Create comprehensive maps of TF binding in B and ES cells Digital genomic footprinDng renders flanking chroma:n DNaseI hypersensi:ve TF protects binging site from DNaseI diges:on DNaseI ChIP-‐Seq valida:on PU.1 CTCF Crystal structure valida:on NFκB-‐cRel Ebf1 ES cell-‐specific factors? B cell-‐specific factors? Tissue-‐specific transcrip:on factors Tissue-‐specific genes Broadly-‐expressed genes “Relay race” transcripDonal regulaDon Cell type A! TFs! TFs! Broadly-‐expressed gene Cell type B! TFs! CH3 CH3 Why do higher organisms modulate the enhancer landscape of broadly expressed genes? 1-‐ To fine-‐tune protein levels Ac:vity 2-‐ To place basic cellular processes (prolifera:on, mo:lity, signal transduc:on) nnunder :ssue-‐specific regula:on Ac:vated cycling G0 quiescent Cell cycle entry Myc Transcriptome amplification ~10 fold (Nie et al. Cell 2012 Kouzine et al. Cell 2013) Ac:vated cycling G0 quiescent B cells must rapidly engage a burst of Myc expression to cope with fast dividing pathogens ES cells ~10 fold amplifica:on No drastic changes in cell proliferation ES cells (4 enhancers) 4 fold increase in transcrip:on B cells (10 enhancers) Kyong-‐Rim Kieffer Kwon Ewy Mathe Jason Qian Wolfgang Resch Nathanael Pruew Laura Vian Steevenson Nelson Arito Yamane Hirotaka Nakahashi NIAMS-‐NIH Funding NIAMS NCI Ethan Taylor Medical Arts NIH Myong-‐Hee Sung Ofir Hakim Songjoon Baek Lars Grøntved Gordon L. Hager NCI-‐NIH Jizhong Zou Stem Cell Ins:tute NIAMS-‐NIH Zhonghui Tang Guoliang Li Chia-‐Lin Wei Xiaoan Ruan Yijun Ruan Jackson Labs Hossein Zare Viworio Sartorelli NIAMS-‐NIH Sasha Kovalchuk NIAID-‐NIH Deepak Reyon J. Keith Joung Harvard
Related documents