Dynamics of epigenetic states during
erythroid differentiation
Ross Hardison
July 20, 2010
Barcelona
Investigators on one mouse ENCODE project
•
• Cal Tech
- Barbara Wold
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
Hematopoiesis
Erythroid differentiation
GATA1, partners, teammates
Somatic cell model to study GATA1 function
in vitro hematopoietic differentiation
Gata1–
ES cells
erythropoietin
stem cell factor
immortalize
G1E
Erythroid progenitors
BFU-e, CFU-e
add back GATA-1,
hybrid protein with ER
G1E-ER4
estradiol
G1E-ER4+estradiol
Differentiated
erythroblasts
Changes in transcription factors and cell morphology
during eythroid differentiation
Changes in gene
expression during
erythroid
differentiation
Features interrogated by ChIP-seq and RNAseq assays
DNase hypersensitive sites
CTCF
ChIP-seq finds previously known distal CRMs
Known CRMs
DHSs
GATA1
TAL1
CTCF
H3K4me1
H3K4me3
PcG:
H3K27me3
+
+
+
+
+
+
+
- = no GATA1
+ = GATA1-ER activated
GATA1 activates Zfpm1 by displacing GATA2 and retaining TAL1
Known CRMs
DHSs
+
GATA1
+
TAL1
+
-
GATA2
H3K4me1
+
+
-
H3K4me3
+
-
PcG:
H3K27me3
+
-
GATA1 represses Kit by displacing GATA2 and TAL1
Known CRMs
DHSs
GATA1
TAL1
GATA2
H3K4me1
H3K4me3
PcG:
H3K27me3
+
+
+
+
+
+
+
-
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
Dominant mark
H3K4me3
H3K4me3
H3K4me1
H3K27me3
None
H3K4me1 H3K27me3
None
Changes in chromatin state at Btg2
Quantum change in chromatin state distinguishes
expressed from non-expressed loci
Chromatin state precedes GATA1-induced TF changes
Chromatin state
established (mostly):
Active
Repressed
Dead zones
Induction and repression:
Dynamics of transcription
factor binding
within the alreadyestablished chromatin
context.
Chromatin condenses
Nucleus removed
Thanks
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)
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
Pol2
H3K4me1
H3K4me3
PcG:
H3K27me3
+
+
+
+
+
+
+
-
TAL1 occupancy, ER4
GATA1 causes TAL1 to move at some CRMs,
but retains it at others
TAL1 occupancy, ER4
TAL1 occupancy, G1E
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
Kinetics of GATA1-regulated Gene Expression
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
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
High
Level of expression
Low
H3K4me3 at TSS correlates with expression level