Supplemental Information Defining Estrogenic Mechanisms of

Chemistry & Biology, Volume 21
Supplemental Information
Defining Estrogenic Mechanisms of Bisphenol A
Analogs through High Throughput
Microscopy-Based Contextual Assays
Fabio Stossi, Michael J. Bolt, Felicity J. Ashcroft, Jane E. Lamerdin, Jonathan S. Melnick,
Reid T. Powell, Radhika D. Dandekar, Maureen G. Mancini, Cheryl L. Walker, John K.
Westwick, and Michael A. Mancini
SUPPLEMENTAL DATA
Figure S1 (related to Figure 1).
A) Representative images of ER and ER recruitment to the PRL array (top) and correspondent increase
in dsRED2 FISH output (bottom); B-C) co-culture between ER and ER containing cells shows
differential recruitment of SRC-3 (B) or similar recruitment of SRC-1 (C). To discriminate ER from
ER in the co-culture system an ER specific antibody is used.
A.
ERα
ERβ
GFP-ER
dsRED2
FISH
Veh
10nM E2
Veh
10nM E2
GFP-ERα/β
Anti-ERα
Anti-SRC-3
GFP-ERα/β
Anti-ERα
Anti-SRC-1
B.
C.
Figure S2 (related to Figure 2).
BPXs dose response curves in GFP-ER:PRL-HeLa cells
BPC
MH-BPF
50
-6
-5
0
-10
-4
-9
-8
MM2
-8
-7
-6
-5
-9
-8
TMBPA
-9
-8
-7
-6
-5
-5
0
-10
-4
-9
-8
TCBPA
-6
-5
-6
-5
0
-10
-4
-9
-8
MonoMxy
-7
-6
-5
-8
-7
-6
-5
Concentration (M)
-4
-6
-5
-9
-8
-7
-6
-5
Concentration (M)
-4
-5
-4
50
-9
-8
-7
-6
-5
-4
Concentration (M)
BPZ
150
-5
EC50 : 6.36x10 -6
-9
-8
-7
-6
-5
-4
50
-9
-8
-7
-6
-5
Concentration (M)
100
50
0
-10
-9
-8
-7
-6
-5
Concentration (M)
100
0
-10
-6
100
0
-10
-4
BPS
50
0
-10
-7
EC50 : NA
100
-7
TBBPA
150
%E2 10nM
%E2 10nM
-9
-8
BPB
50
-8
EC50 : NA
-6
Concentration (M)
EC50 : ~0.078
100
-9
Concentration (M)
50
0
-10
-4
150
-6
0
-10
-4
100
Concentration (M)
150
EC50 : 4.17x10
-9
EC50 : 1.08x10
%E2 10nM
%E2 10nM
-7
-5
50
BPA
50
-4
100
Concentration (M)
100
Concentration (M)
-6
150
EC50 : 1.08x10
50
-7
50
-5
-5
150
DMBPA
100
-8
-8
100
0
-10
-4
150
EC50 : NA
-9
-9
EC50 : 2.13x10
-7
-6
EC50 : 2.68x10 -7
DMBBPA
Concentration (M)
-7
DMDMBBPA
150
50
-8
Concentration (M)
Concentration (M)
100
0
-10
-4
150
%E2 10nM
-6
%E2 10nM
%E2 10nM
%E2 10nM
50
-9
150
BPF
100
0
-10
-4
50
EC50 : NA
Concentration (M)
%E2 10nM
-7
150
EC50 : ~0.078
-5
100
Concentration (M)
150
-6
EC50 : ~0.078
50
0
-10
-4
-7
50
MHMM1
100
Concentration (M)
-8
150
%E2 10nM
%E2 10nM
%E2 10nM
-9
-9
100
Concentration (M)
EC50 : ~0.078
50
0
-10
0
-10
-4
NSC
-6
100
0
-10
-5
150
EC50 : 3.75x10
0
-10
-6
Concentration (M)
150
0
-10
-7
50
%E2 10nM
-7
Concentration (M)
100
%E2 10nM
-8
50
EC50 : 0.026
-6
%E2 10nM
-9
150
EC50 : 1.62x10
100
%E2 10nM
%E2 10nM
%E2 10nM
EC50 : 4.26x10
BPAP
150
EC50 : NA
100
0
-10
BPAF
150
-7
%E2 10nM
150
-4
-4
Figure S3 (related to Figure 2).
BPXs dose response curves in GFP-ER:PRL-HeLa cells
BPC
MH-BPF
200
150
100
50
150
100
50
-7
-6
-5
-4
-9
-8
-9
-8
-7
-6
-5
-4
-8
-8
-7
-6
200
100
-5
-4
-8
-7
-6
-5
-4
150
100
50
200
100
50
-6
-5
-4
Concentration (M)
-8
MonoMxy BPA
-7
-6
-5
-4
100
50
200
100
50
-9
-8
-7
-6
-5
Concentration (M)
-4
-8
-7
-6
-5
Concentration (M)
-4
-9
-8
-7
-6
-5
-4
150
100
0
-10
-4
-9
-8
-7
-6
-5
-4
Concentration (M)
BPZ
250
EC50 : 7.32x10 -7
200
EC50 : 1.34x10 -7
150
100
50
-9
-8
-7
-6
-5
-4
EC50 : 2.5x10 -6
100
-9
-8
-7
-6
-5
Concentration (M)
0
-10
-9
-8
-7
-6
-5
Concentration (M)
150
0
-10
-5
50
50
-9
-6
EC50 : NA
BPS
EC50 : 1.61x10 -7
-7
200
250
150
0
-10
-8
TB BPA
Concentration (M)
%E2 10nM
%E2 10nM
150
-9
Concentration (M)
EC50 : 4.5x10 -8
BPB
200
0
-10
-4
100
0
-10
250
EC50 : 7.08x10 -7
-5
150
Concentration (M)
250
-6
50
-9
100
BPA
150
0
-10
-7
250
EC50 : 9.31x10 -7
-4
150
Concentration (M)
%E2 10nM
%E2 10nM
200
-5
250
DM BPA
EC50 : NA
-7
-8
100
Concentration (M)
200
-8
-9
50
-9
-6
50
150
0
-10
-7
EC50 : 9.85x10 -8
DMB BPA
250
-9
200
Concentration (M)
%E2 10nM
%E2 10nM
%E2 10nM
-4
-8
DMDMB BPA
250
TC BPA
%E2 10nM
-5
50
250
%E2 10nM
-6
150
Concentration (M)
0
-10
-7
-9
Concentration (M)
100
EC50 : NA
0
-10
0
-10
-4
150
0
-10
200
50
-5
250
BPF
100
-6
EC50 : NA
250
150
-7
200
Concentration (M)
EC50 : 6.38x10 -7
-8
50
-9
100
MH-MM1
EC50 : 3.23x10 -6
TM BPA
-9
-9
250
100
0
-10
150
Concentration (M)
50
250
200
-4
150
Concentration (M)
0
-10
-5
%E2 10nM
200
%E2 10nM
%E2 10nM
EC50 : 4.99x10 -7
50
0
-10
-6
EC50 : 8.09x10 -8
50
NSC
100
200
-7
0
-10
250
150
0
-10
100
50
MM2
200
150
Concentration (M)
250
200
%E2 10nM
-8
Concentration (M)
EC50 : 2.91x10
%E2 10nM
-9
0
-10
250
-8
%E2 10nM
0
-10
200
%E2 10nM
EC50 : 7.09x10
BPAP
250
EC50 : NA
%E2 10nM
%E2 10nM
200
BPAF
250
-8
%E2 10nM
250
-4
-4
Figure S4 (related Figure 5).
Representative images of MCF-7 cells treated for 24hrs with either vehicle or 10nM E2 and then
hybridized with GREB1 intron RNA FISH probes. DAPI stain is in red, RNA FISH in green.
Veh
E2
Figure S5 (related to Figure 6).
Analysis of BPXs chemical structure similarity using Tanimoto distance. Upper panel:
Multidimensional Scaling plot; lower panel: hierarchical clustering
BPA
DM
TM
TB
TC
BPB
Mxy DMB
NSC
BPF
BPS
DMDMB
BPAF
BPC
MM2
MH-MM1
MH-BPF
BPAP
E2
4OHT
BPZ
Table S1 (related to all Figures).
List of BPXs used in the study
Name
BPC
CAS #
14868-03-2
CID #
84677
6807-17-6
81259
50628-55-2
252259
1478-61-1
73864
77-40-7
66166
BPZ
843-55-0
232446
1,1-bis(4hydroxyphenyl)cyclohexane
NSC17960
1844-00-4
226969
79-97-0
6620
dihydroxymethoxychlor olefin
DMB BPA
(4,4'-(1,3Dimethylbutylidene)diphenol)
DM DMB BPA
4,4'-(4-methylpentane-2,2diyl)bis(2-methylphenol)
BPAF
4,4'(hexafluoroisopropylidene)diphenol
BPB
bisphenol B
4,4'-(2Methylpropylidene)bisphenol
DM BPA
2,2-Bis(4-hydroxy-3methylphenyl)propane
Structure
MM2
1576-13-2
4201081
1571-75-1
623849
5613-46-7
79717
80-05-7
6623
Mono Mxy BPA
NA
NA
MH-MM1
1988-89-2
94763
620-92-8
12111
101-53-1
7563
79-95-8
6619
4-[1-(4hydroxyphenyl)propyl]phenol
BPAP
Bisphenol AP
TM BPA
2,2-Bis(4-hydroxy-3,5dimethylphenyl)propane
BPA
bisphenol A
Phenol, 4-(1-phenylethyl)-
BPF
4,4'-methylenebis(phenol)
MH-BPF
4-benzylphenol
TC BPA
Tetrachlorodian
TB BPA
79-94-7
6618
80-09-1
6626
tetrabromobisphenol A
BPS
bis(4-hydroxyphenyl)sulfone

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