+O+---
+
AqNR2 (FAs)
HNF4s (FAs)
PNR
H1
2
seq co ac
u en t i va
ce c t o r
ore
L6H1
1s
al t
b ri
Rig
dg
id
e
to H H11
4 a ; H1
2H
nd
H1
-bo
0
nds
Bu l
i n pk y si d
o ck e-ch
et
ai n
s
s
AqNR1 (FAs)
AncNR:
Lig
a
ope nd po
n o cke
r fil t
led
3 la
yer
A
O (835)
O**
–
–
–
–
–
+
+
O (659)*
+
+
RXRs/USP (FAs, retinoids)
+
O (574)
+
–
–
–
SF-1 (phospholipids)
LRH-1 (phospholipids)
rodent LRH-1
+
O (2285)
+
+
–
–
+
O (1633)
+
–
–
–
+
O (759)
–
–
–
+
F (<50)
O (574)*
+
–
+
–
–
3
–
+
+
+
O (701)*
O (418-450)
+
–
–
–
MR, GR, PR, AR (steroids)
Branchiostoma SR (steroids)
+
O (413-538)
+
–
–
–
+
O (438)*
+
–
–
–
ERRs (phenolics)
+
+
–
–
1
Nurr, NOR1, NGF1B, DHR38
+
F/O (<50-322)
F (<50)
+
–
–
2
PPARs (FAs, phosphoglycerides) +
RORs (cholesterol, retinoids)
+
Rev-Erbs/E75 (heme)
+
O (1150)
+
–
–
–
O (888-1969)
+
–
–
–
O (606)
+
O (674)
–
–
–
–
–
ECR (steroids)
FXR (bile acids)
LXR (oxysterols)
–
+
+
O (1257)
+
–
–
–
+
O (1018)
+
–
–
–
VDR (steroids, bile acid)
CARb (steroids)
+
O (800)
+
–
–
–
+
+
O (646)
O (1107)
+
+
–
–
+
–
–
–
DAF-12 (steroids)
TR (thyroxine)
RARs (retinoids)
+
O (719)
+
–
–
–
+
O (485)
+
–
–
–
+
O (488)
+
–
–
–
AqNR1 (FAs)
AqNR2 (FAs)
+
O (835)
O**
–
–
–
HNF4s (FAs)
PNR
TLL, TLX, DSF, FAX1
+
O (680)
+
?
+
+
–
–
–
–
COUP-TFs (serum components)
TR2, TR4 (FAs)
RXRs/USP (FAs, retinoids)
+
+
+
O (659)*
+
+
+
–
–
–
–
–
–
SF-1 (phospholipids)
LRH-1 (phospholipids)
rodent LRH-1
GCNF, GRF
mollusk ERs
annelid ERs (steroids)
+
O (2285)
O (1633)
O (759)
+
+
–
–
+
+
–
+
–
–
–
–
–
–
–
–
3
+
+
+
+
+
+
+
–
+
+
+
+
+
+
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
+
–
–
–
–
1
2
–
–
–
–
–
–
–
–
–
+
+
+
–
–
–
–
–
–
–
–
–
PXR (steroids, xenobiotics)
Branchiostoma ER
vertebrate ERs (steroids)
MR, GR, PR, AR (steroids)
Branchiostoma SRs (steroids)
+
+
+
O (574)
F (<50)
O (574)*
PXR (steroids, xenobiotics)
+
+
O (701)*
O (418-450)
O (413-538)
O (438)*
F/O (<50-322)
F (<50)
O (1150)
O (888-1969)
O (606)
O (674)
O (1257)
O (1018)
O (800)
O (646)
O (1107)
DAF-12 (steroids)
TR (thyroxine)
RARs (retinoids)
+
+
+
O (719)
O (485)
O (488)
ERRs (phenolics)
Nurr, NOR1, NGF1B, DHR38
PPARs (FAs, phosphoglycerides)
RORs (cholesterol , retinoids)
Rev-Erbs/E75 (heme)
ECR (steroids)
FXR (bile acids)
LXR (oxysterols)
VDR (steroids, bile acid)
CAR (steroids)
Gain of ligand-binding and regulation
–
COUP-TFs (serum components)
TR2, TR4 (FAs)
Branchiostoma ER
vertebrate ERs (steroids)
AncNR
–
+
+
mollusk ERs
annelid ERs (steroids)
B
–
O (680)
GCNF, GRF
Gain of ligand-independent activity
–
+
TLL, TLX, DSF, FAX1
Loss of ligand-binding and regulation
+
?
+
+
+
+
+
+
+
+
+
+
+
+
–
+
+
Fig. S9. Reconstruction of structural and functional characters on an alternate NR phylogeny. This phylogeny represents the next-best
alternative to the ML phylogeny; it differs in that it groups AqNR1 and AqNR2 as sponge-specific duplicates. A. Maximum parsimony
reconstruction of structural and functional characters assuming an ancestral ligand-binding, ligand-dependent transcriptional activator.
B. Maximum parsimony reconstruction assuming an ancestral ligand-independent activator. In both panels, ligand-regulated
transcriptional activators are shown in green, with ligands in parentheses. Red, activators with no known ligand. Underlined, receptors
with transcriptional activity in the absence of ligand or other modifications. Black, repressors that do not activate transcription. The
ancestral NR (AncNR, green circle) is shown, with the most parsimonious character reconstruction at that node. Hash marks on branches
show gains of ligand-independent activity with or without loss of ligand binding (filled and empty red boxes, respectively). States of
protein structural characters are shown in the table. Specific characters are described in Fig. 5 of the main text of the paper.