3box pse -55 dse -220+1 +350 u2 -800-500 +700 +150 -15-200 +215 % of input ptf 4563 figure s1. high...
TRANSCRIPT
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-800 -500 -200 -15 +150 +350 +700
3’boxPSE-55
DSE-220 +1
+350
U2
-800 -500 +700+150-15-200
+215
% o
f in
pu
t
PTF
4 5 63
Figure S1. High resolution chromatin immunoprecipitation of PTF on U2 snRNA genes.qRT-PCR of ChIP analysis of the U2 gene using anti-PTF antibodies and the primers noted on the diagram of the U2 genes shown above.
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% o
f in
pu
t
Histone H4
Histone H2A
Histone H2B
RNU2 locus -actin gene
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1 2 3 4 5 6 7 8
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1 2 3 4 5 6 7 8
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Histone H4
Histone H2A
Histone H2B
% o
f in
pu
t
% o
f in
pu
t
% o
f in
pu
t
% o
f in
pu
t
% o
f in
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t
Figure S2. All four histones have the same pattern of depletion on the U2 and the -actin genes.qRT-PCR of ChIP analysis of the U2 and -actin genes using antibodies against the histones noted.
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1 2 3 4 5 6 7 8
% o
f in
pu
t
- -amanitin
+ -amanitin
% o
f in
pu
t
Histone H3 RNA pol II
RNU2 locus
Figure S3. Nucleosomal depletion on U2 genes is transcription-independent.qRT-PCR of ChIP analysis of the U2 gene using antibodies against H3 and pol II, with or without treatment of the cells with -amanitin.
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+300 +500 +700 +850 +1500 +2100
3’boxPSE-55
DSE-220 +1 +215
CT-rich+600
U2
+850 +1500 +2100+300 +500 +700
7 8
transcription
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% o
f in
pu
t%
of
inp
ut
% o
f in
pu
t
NELF-E
Histone H3
CTCF
Figure S4. Fine mapping of NELF and CTCF on the RNU2 locus.qRT-PCR of ChIP analysis of the RNU2 locus, using antibodies against H3, NELF-E and CTCF using the primers noted on the diagram of the RNU2 locus above.
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0.9Ser7/pol II
Ser5/pol II
RNU2 locus
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1.2Ser7/pol II
Ser5/pol II
-actin gene
3’boxPSE-55
DSE-220 +1 +215
CT-rich+600
U2
4a 4 5 63
Se
r5 r
ela
tive
to
Po
l II
Se
r5 r
ela
tive
to
Po
l II
Se
r7 r
ela
tive
to
Po
l II
Se
r7 r
ela
tive
to
Po
l II
-800
Figure S5. Distribution of Ser5 and Ser7 phosphorylation on the RNU2 locus and the -actin gene. qRT-PCR of ChIP analysis using antibodies against phospho-Ser5 and phospho-Ser7, relative to the Pol II signal. The additional primer, 4a, is noted on the diagram of the RNU2 locus above. All other primers are as shown in Figure 1.
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1 2 3 4 5 6 7 80
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1
1.5
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2.5
3
Sp
t16
leve
l (%
in
pu
t)
Sp
t16
leve
l (%
in
pu
t)
Po
l II
le
vel
(% i
np
ut)
Po
l II
le
vel
(% i
np
ut)
Spt16Pol II
Spt16Pol II
RNU2 locus -actin gene
Figure S6. FACT is associated with both snRNA and protein-coding genes.qRT-PCR of ChIP analysis of the U2 and -actin genes, using antibodies against the Spt16 subunit of FACT. The pol II level is indicated by a dotted line.
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1 2 3 4 5 6 7 80
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Ac-
H4
(rel
ated
to
H4)
Ac-
H4
(rel
ated
to
H4)
RNU2 locus -actin gene
H4
leve
l (%
in
pu
t)
H4
leve
l (%
in
pu
t)
AcH4/H4H4
Figure S7. H4 acetylation across the RNU2 locus and -actin gene.qRT-PCR of ChIP analysis of the U2 and -actin genes using antibodies against H4 and acetyl H4 (Ac-H4). The H4 level is indicated by a dotted line and the Ac-H4 level is shown relative to total H4.
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0.5
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1.5
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2.5
1 2 3 4 5 6 7 80
0.5
1
1.5
2
2.5
1 2 3 4 5 6 7 8
H3-
K36
M3 (
rela
ted
to
H3)
RNU2 locus -actin gene
+DRBDRB-
+DRBDRB-
H3-
K36
M3 (
rela
ted
to
H3)
Figure S8. H3-K36 trimethylation is DRB-sensitive.qRT-PCR of ChIP analysis of the U2 and -actin genes using antibodies against trimethyl H3-K36 (H3-K36M3) with or without treatment of the cells with DRB.
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3.5
4
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H3-
K9M
3 (
rela
ted
to
H3)
RNU2 locus -actin gene
H3
leve
l (%
in
pu
t)
H3
leve
l (%
in
pu
t)
H3-K9M3/H3H3
H3-
K9M
3 (
rela
ted
to
H3)
Figure S9. H3-K9 trimethylation across the RNU2 locus and -actin gene.qRT-PCR of ChIP analysis of the U2 and -actin genes using antibodies against trimethyl H3-K9 (H3-K9M3). The H3 level is indicated by a dotted line and the H3-K9M3 level is shown relative to total H3.
RNU2 locus -actin gene GAPDH gene -actin gene
1 -2780/-2600 -609/-479 -119/+30 -102/+71
2 -2000/-1819 -169/-10 +258/+407 +246/+415
3 -919/-739 +222/+361 +658/+817 +666/+895
4 -40/+35 +742/+911 +1018/+1167 +989/+1122
5 +270/+460 +1112/+1242 +1328/+1501 +1656/+1789
6 +651/+820 +1382/+1511 +3333/+3507 +2550/+2664
7 +1422/+1560 +2540/+2671
8 +2022/+2202 +3541/+3682
Figure S10. Regions analysed by qRT-PCR after chromatin immunoprecipitation.Regions amplified by each pair of primers on the RNU2 locus, the -actin, GAPDH and -actin genes are indicated relative to the transcription start site.