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The three-dimensional folding of the -globin gene domain reveals - - PowerPoint PPT Presentation

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The three-dimensional folding of the -globin gene domain reveals formation of chromatin globules. Davide Ba Structural Genomics Group Thursday, February 16, 12 Highlight 2011 The three-dimensional folding of

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The three-dimensional folding of the α-globin gene domain reveals formation of chromatin globules.

Davide Baù

Structural Genomics Group

Thursday, February 16, 12

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  • The three-dimensional folding of the -globin gene

domain reveals formation of chromatin globules

Davide Baù1,4, Amartya Sanyal2,4, Bryan R Lajoie2,4, Emidio Capriotti1, Meg Byron3, Jeanne B Lawrence3, Job Dekker2 & Marc A Marti-Renom1

“Highlight 2011”

Thursday, February 16, 12

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SLIDE 3

Adapted from: Langowski and Heermann. Semin Cell Dev Biol (2007) vol. 18 (5) pp.

μ 10 10 10 Resolution s Time 10 10 10 10 10 10 10 10 μm Volume 10 10 10 10 10 DNA length nt 10 10 10 10

Knowledge

IDM INM

Resolution

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  • 5C “copies” the 3C library into a 5C library containing only ligation junctions
  • Performed at high levels of multiplexing:
  • 2,000 primers detect 1,000,000 unique interactions in 1 reaction

5C Technology

Detecting up to millions of interactions in parallel

http://my5C.umassmed.edu

Dostie et al. Genome Res (2006) vol. 16 (10) pp. 1299-309

Thursday, February 16, 12

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SLIDE 5

The ENCODE data for ENm008 region was obtained from the UCSC Genome Browser tracks for: RefSeq annotated genes, Affymetrix/ CSHL expression data (Gingeras Group at Cold Spring Harbor), Duke/NHGRI DNaseI Hypersensitivity data (Crawford Group at Duke University), and Histone Modifications by Broad Institute ChIP-seq (Bernstein Group at Broad Institute of Harvard and MIT).

RAB11FIP3 DECR2 LOC1001134368 HS8 HS10 HS40 HS33 HS46 HS48 TMEM8 MRPL28 AXIN1 PDIA2 ARHGDIG RGS11 ITFG3 LUC7L HB HB1 HB2 HB HB C16ORF35 500000| 450000| 400000| 350000| 0| 300000| 250000| 200000| 150000| 100000| 50000| POLR3K SNRNP25 RHBDF1 MPG

p13.3 13.2 12.3 p12.1 16p11.2 11.1 q11.2 q12.1 13 16q21 22.1 q23.1

K562 GM06990 DNaseI K562 GM12878 H3K4me3 CTCF K562 GM12878 K562 GM12878 RNA diff CTCF

K562 GM12878

Human α-globin Domain

ENm008 genomic structure and environment

ENCODE Consortium. Nature (2007) vol. 447 (7146) pp. 799-816

Thursday, February 16, 12

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SLIDE 6

RAB11FIP3 DECR2 LOC1001134368 HS8 HS10 HS40 HS33 HS46 HS48 TMEM8 MRPL28 AXIN1 PDIA2 ARHGDIG RGS11 ITFG3 LUC7L HB HB1 HB2 HB HB C16ORF35 500000| 450000| 400000| 350000| 0| 300000| 250000| 200000| 150000| 100000| 50000| POLR3K SNRNP25 RHBDF1 MPG

p13.3 13.2 12.3 p12.1 16p11.2 11.1 q11.2 q12.1 13 16q21 22.1 q23.1

GM12878

250 500 750 >1,000 GM12878 cells 5C counts

b

250 500 750 >1,000 Forward fragments Reverse fragments K562 cells 5C counts

a

Forward fragments Reverse fragments

  • GM12878

K562

Human α-globin Domain

ENm008 genomic structure and environment

ENCODE Consortium. Nature (2007) vol. 447 (7146) pp. 799-816

Thursday, February 16, 12

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SLIDE 7

Biomolecular structure determination 2D-NOESY data Chromosome structure determination 5C data

Structure Determination

Integrative Modeling Platform

http://www.integrativemodeling.org

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SLIDE 8

Integrative modeling

P1 P2 P1 P2 P1 P2

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SLIDE 9

Representation

i i+2 i+1 i+n

Harmonic Lower Bound Harmonic Upper Bound Harmonic

Hi, j = k di, j − di, j

( )

2

if di, j ≤ di, j

0 ;

lbHi, j = k di, j − di, j

( )

2

if di, j > di, j

0 ;

lbHi, j = 0 $ % & ' & if di, j ≥ di, j

0 ;

ubHi, j = k di, j − di, j

( )

2

if di, j < di, j

0 ;

ubHi, j = 0 $ % & ' &

Thursday, February 16, 12

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SLIDE 10

Scoring

Harmonic Harmonic Lower Bound Harmonic Upper Bound

GM1287 K562

70 fragments 1,520 restraints 70 fragments 1,049 restraints

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Optimization

CREATE PARTICLES ADD RESTRAINTS SIMULATED ANEALING MONTE-CARLO LOCAL CONJUGATE GRADIENT LOWEST OBJECTIVE FUNCTION start end 500 rounds 5 steps

50 100 150 200 250 300 350 400 450 500 Iteration 0.0E+00 1.00E+06 2.00E+06 3.00E+06 4.00E+06 5.00E+06 6.00E+06 7.00E+06 IMP Objective function

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Not just one solution

  • b

a c

50 100 150 200 250 300 350 400 Cluster number 100 200 300 400 500 Number of models in cluster Lowest IMP objective function in cluster 231,000 234,500 237,000 241,500 244,000 1 2 3 4 5 Cluster number 2,200 2,300 2,400 2,500 2,600 2,700 2,800 Number of models in cluster 910,280 911,705 913,130 914,555 915,980 Lowest IMP objective function in cluster

Cluster 1 2,780 models 910,280 OF Cluster 4 2,270 models 915,890 OF Cluster 3 2,282 models 915,890 OF Cluster 2 2,668 models 910,400 OF

910,000 911,500 913,000 914,500 916,000 Lowest IMP objective function in cluster

Cluster 1 483 models 232,698 OF Cluster 2 314 models 232,673 OF Cluster 3 275 models 233,110 OF Cluster 4 265 models 233,051 OF Cluster 5 256 models 233,026 OF Cluster 6 228 models 232,819 OF Cluster 7 226 models 233, 398 OF Cluster 8 209 models 233,748 OF Cluster 9 205 models 233,049 OF Cluster 10 168 models 234,747 OF

232,600 233,250 233,800 234,450 235,000

Lowest IMP objective function in cluster

GM12878 K562

Thursday, February 16, 12

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Long-range interactions

  • e

d

200 400 600 800 1,000 Distance (nm) K562 GM12878

c

K562 GM12878 Fragment

  • 100 nm

b a

100 nm

c

  • Thursday, February 16, 12
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SLIDE 14

Chromatin globules

Frequency contact map differences

  • 100 nm

b a

100 nm

c

  • Increased in K562

Increased in GM12878 =

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Genes location within the globules

  • Increased in K562

K562 Distance to center (nm) <400 <300 <350 <250 <200 <150 <100 <50 Relative abundance Promoters Active genes Inactive genes DNase I sites CTCF sites H3K4me3 sites 0.5 1.0 1.5 2.0 2.5

  • b

Promoters Active genes Inactive genes DNase I sites CTCF sites H3K4me3 sites <400 <300 <350 <250 <200 <150 <100 <50 Distance to center (nm) Relative abundance GM12878 0.5 1.0 1.5 2.0 2.5 2.5

  • 100 nm

b a

100 nm

c

  • Thursday, February 16, 12
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SLIDE 16

Model validation

  • 100 nm

b a

100 nm

c

  • K562

GM12878

100 200 300 400 500 Distance (nm)

GM12878 K562

FISH Models (2D)

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SLIDE 17

Summary

5C data results in consistent 3D models

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α-globin- Enhancer looping

  • nly in K562

Looping interaction In GM12878 cells

K562 GM12878 RNA diff

RAB11FIP3 DECR2 LOC1001134368 HS8 HS10 HS40 HS33 HS46 HS48 TMEM8 MRPL28 AXIN1 PDIA2 ARHGDIG RGS11 ITFG3 LUC7L HB HB1 HB2 HB HB C16ORF35 POLR3K SNRNP25 RHBDF1 MPG

Summary

Conformational changes correlate with gene expression

  • 100 nm

b a

100 nm

c

  • Thursday, February 16, 12
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SLIDE 19

The “chromatin globule” model

Münkel et al. JMB (1999) Lieberman-Aiden et al. Science (2009)

PolII HBB Eraf Factory

a b

Osborne et al. Nat Genet (2004)

  • D. Baù et al. Nat Struct Mol Biol (2011) 18:107-14
  • A. Sanyal et al. Current Opinion in Cell Biology (2011) 23:325–33.

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SLIDE 20

Acknowledgments

Emidio Capriotti Marc A. Marti-Renom Amartya Sanyal Bryan R Lajoie Job Dekker Meg Byron Jeanne Lawrence OPEN POSITIONS IN THE LAB Starting early 2012

Thursday, February 16, 12