Dynamics of epigenetic states during erythroid differentiation

Ross Hardison

July 20, 2010

Barcelona

Investigators on one mouse ENCODE project

• Penn State– Hardison– Stephan Schuster – Frank Pugh– Robert Paulson – Francesca Chiaromonte– Yu Zhang– Webb Miller– Anton Nekrutenko

• Childrens’ Hospital of Philadelphia– Mitch Weiss– Gerd Blobel

• Emory Univ.– James Taylor

• Univ. Massachusetts– Job Dekker

• Duke Univ.– Greg Crawford, consultant– Terry Furey, consultant

• Cal Tech - Barbara Wold

Hematopoiesis

Erythroid differentiation

GATA1, partners, teammates

Gata1–

ES cells

in vitro hematopoietic differentiation

erythropoietinstem cell factor

G1E

Somatic cell model to study GATA1 function

immortalize

Erythroid progenitorsBFU-e, CFU-e

G1E-ER4+estradiol

estradiol

Differentiated erythroblasts

add back GATA-1, hybrid protein with ER

G1E-ER4

Changes in transcription factors and cell morphology during eythroid differentiation

Changes in gene expression during

erythroid differentiation

Features interrogated by ChIP-seq and RNA-seq assays

CTCF

DNase hypersensitive sites

ChIP-seq finds previously known distal CRMs

Known CRMs

DHSs

GATA1

TAL1

H3K4me1

H3K4me3

PcG: H3K27me3

CTCF

+-+-

+-

+-+-

+

+

+ = GATA1-ER activated- = no GATA1

GATA1 activates Zfpm1 by displacing GATA2 and retaining TAL1

Known CRMs

DHSs

GATA1

TAL1

H3K4me1

H3K4me3

PcG: H3K27me3

GATA2 +-

+

+

+-

+-

+-

+-

GATA1 represses Kit by displacing GATA2 and TAL1

Known CRMs

DHSs

GATA1

TAL1

H3K4me1

H3K4me3

PcG: H3K27me3

GATA2 +-

++

+-

+-

+-+-

GATA1 action via TAL1 is common but not universal

Chromatin states from integrative analysis: limited change during differentiation

chromHMM segmentations: Jason Ernst and Manolis Kellis, MIT Broad

GATA1-occupied segments of DNA: Vast majority are in active chromatin, few change states

H3K4me3 H3K4me1 H3K27me3 None

Dominant mark

H3K4me3

H3K4me1

H3K27me3

None

Changes in chromatin state at Btg2

Quantum change in chromatin state distinguishes expressed from non-expressed loci

Enr

ichm

ent

of c

hro

mat

in s

tate

Chromatin state precedes GATA1-induced TF changes

Chromatin stateestablished (mostly):ActiveRepressedDead zones

Induction and repression:Dynamics of transcription factor bindingwithin the already-established chromatin context.

Chromatin condensesNucleus removed

Weisheng Wu, Yong Cheng, Demesew Abebe, Cheryl Keller Capone,Ying Zhang, Ross, Swathi Ashok Kumar, Christine Dorman, David King ….Tejaswini Mishra, Nergiz Dogan

Collaborating labs: Mitch Weiss and Gerd Blobel (Childrens’ Hospital of Philadelphia), James Taylor (Emory)Webb Miller, Francesca Chiaromonte, Yu Zhang, Stephan Schuster, Frank Pugh, Bob Paulson (PSU), Greg Crawford (Duke), Jason Ernst, Manolis Kellis (MIT)

Thanks

Funding: NIH NIDDK, NHGRI (ARRA), Huck Institutes of Life Sciences and Institute for Cyberscience, PSU

Big changes at Hbb-b1

Known CRMs

DHSs

GATA1

TAL1

H3K4me1

H3K4me3

PcG: H3K27me3

Pol2

+-+-

+-

+-+-

+

+

GATA1 causes TAL1 to move at some CRMs, but retains it at others

TAL1 occupancy, G1E

TA

L1 o

ccup

ancy

, ER

4T

AL1

occ

upan

cy, E

R4

GATA1 occupancy, ER4

All GATA1-occupied segments active as enhancers are also occupied by SCL and LDB1

Tripic et al. (2009) Blood

Distinguishing genes induced vs repressed by GATA1

Feature Induction Repression

GATA1 occupancy and transcription start site

Close Distal

Multiple GATA1 occupied segments per gene

More (58% of genes)

Fewer (24% of genes)

GATA1 and TAL1 co-occupancy Almost always Subset has GATA1+ TAL1-

Binding site (WGATAR) is under constraint

More frequently Less frequently

Polycomb mark around TSS Never Subset has Pc

GATA1-induced (>2-fold)1048 genes

• known targets• new gene discovery

GATA1-repressed (>2-fold)1568 genes

• stem cell/progenitor markers• proto-oncogenes (Kit/Myc/Myb)• function unknown

Affy 430 2.0

Kinetics of GATA1-regulated Gene Expression

Globin RNA dominates the transcription profile

RNA-seq

Change in expression

Swathi A. Kumar, Tejaswini Mishra, et al.

Induction and repression occurs within active chromatin

Histone modification profiles across TSS+/ 4kb

Level of expression

High

Low

H3K4me3 at TSS correlates with expression level