Transcript YEO_RNA_2006

REST regulatory networks
Gene Yeo1,2
Stefan Aigner 2, Eric Van Nostrand1,2, Fred H. Gage2
1Crick-Jacobs
Center of Theoretical and Computational Biology
2Laboratory of Genetics
The Salk Institute for Biological Studies, La Jolla, CA
REST repressor/silencing complex
silences neuronal genes in non-neuronal
cells
Differentiated non-neuronal cells
m
mK9
mCpG
Methyl H3K9
HDAC
MeCP2
Histone K4
Demethylase
HDAC
SCP
mSin3
CoREST
REST
mK9
m
mK9
m
mK9
RE1
m
mK9
m
m
HMT K9
m
mK9
MeCP2
m
m
DNMT1
HP1
mK9
mK9
m
mK9
REST binds to a ~20 bp sequence
REST levels decrease during neuronal differentiation
Slide courtesy of XC
REST sites are highly-conserved across
evolution
Superiorcervical
ganglion neuralspecific 10
Synaptosomalassociated
protein 25
Brain-derived
neurotrophic
factor
Algorithm to identify conserved REST
sites
Converged motif
Several REST sites are proximal to
microRNAs
REST-regulated microRNAs are neuronal
REST binds in vivo to predicted sites
REST occupancy near microRNA genes
decreases during neural differentiation
0.8
0.7
0.6
Undifferentiated
Differentiated
0.5
0.4
0.3
0.2
0.1
iR
21
91
m
iR
18
2/
96
/1
83
m
iR
19
2/
19
42
m
iR
13
5b
m
30
e/
30
c1
iR
15
31
m
m
iR
-9
-3
iR
-2
9a
/2
9b
-1
0
m
% of chromatin immunoprecipitated
Anti-REST chromatin immunoprecipitation/qPCR for candidate
REST sites in P19 cells upon neural induction
Neural-specific microRNAs and REST
co-repressors during mouse stem cell
neuronal differentiation
Neural differentiation of P19 mouse embryonic carcinoma cells
RA induction
Western blots
Northern blots
d0 d5 d7 d9 d11
miR-9
miR-124a
d0 d5 d7 d9 d11
REST
msin3A
SCP-1
CoREST
U6
Neuronal marker TUJ1
Neural-specific microRNAs likely target components of the REST co-repressor complex
mir-9 and mir-124a downregulate mSin3a and SCP-1,
respectively
Plasmid
transfected: GFP
Plasmid
transfected:
empty mir-9
msin3A
SCP-1
Loading
control
-actin
m
mK9
mCpG
Methyl H3K9
HDAC
MeCP2
Histone K4
Demethylase
HDAC
SCP
mSin3
CoREST
REST
mK9
m
mK9
m
mK9
RE1
m
mK9
m
m
HMT K9
m
mK9
MeCP2
m
m
DNMT1
HP1
mK9
mK9
m
mK9
Neural-specific microRNAs likely target components of the REST corepressor complex
A sensor assay confirms predicted target sites for miR-124a in the
3’UTR of SCP-1
SCP-1 3’UTR
SCP-1 sensor
ADAR1 3’UTR
Control sensor
Control sensor
SCP1 sensor + mir124
Enhancing neurogenesis
mir-124 induces neuronal gene expression
in P19 mouse EC cells
6
Fold change
(mir124/control)
5
4
24 hours
48 hours
3
2
1
0
GAP43
NFH
Glur2
Promoter driving luciferase
ND1
A model for REST regulation to enhance
neurogenesis
Neuronal progenitors
REST high
Neurons
REST low
REST-regulated microRNAs ON
REST-regulated microRNAs
downregulate components of
the REST repressor complex
REST-regulated microRNAs OFF
REST-regulated protein coding genes
OFF
Acknowledgements: Eunice Meija, Xinwei Cao
REST-regulated protein coding
genes ON
Supported by Crick-Jacobs Center, NSF