Supporting InformationSherratt et al. 10.1073/pnas.1506516112
Fig. S1. Toepad morphology of fore- and hindfeet among recent (representing five ecomorphs) and fossil anoles. Lamella numbers for the modern species aregiven beside digits III and IV. The amber fossils are colored by ecomorph according to DFA results if assigned with a probability greater than 0.90 (Table S3).Modern specimens from Museum of Comparative Zoology Reptile collection, Harvard University (prefix MCZ R).
Sherratt et al. www.pnas.org/cgi/content/short/1506516112 1 of 12
Twig
(A
nolis
inso
litus
)Tr
unk
(A. b
revi
rost
ris)
Gra
ss-b
ush
(A. d
olic
ocep
halu
s)Tr
unk-
crow
n (A
. chl
oroc
yanu
s)Tr
unk-
grou
nd
(A. c
ybot
es)
O
K
J
F
MCZ R-59788T-0845
MCZ R-124846MCZ R-74865
MCZ R-69274MCZ R-155836
MCZ R-186752T-0339
MCZ R-107016 MCZ R-107014
Fig. S2. Cranial shape of five recent anoles representing five ecomorphs compared with four amber fossils. Crania of modern juveniles (Left) and adults(Center), as well as amber fossil skulls (Right) are shown in lateral and dorsal views. The amber fossils are colored by ecomorph if assigned with a probabilitygreater than 0.90 (Table S3). Modern specimens from Museum of Comparative Zoology Reptile collection, Harvard University (prefix MCZ R), or Field Tag forT.J. Sanger (prefix T).
Sherratt et al. www.pnas.org/cgi/content/short/1506516112 2 of 12
Fig. S3. Variation among the fossils and how it corresponds with variation among the ecomorphs for all 10 cranial measurements. Size-corrected residuals areplotted for the amber fossils labeled by letters (A–T) and modern specimens (colored dots by ecomorph). Amber fossils are colored according to the DFA results(detailed in Table S3) when assigned with a probability greater than 0.90. A plot of the first two canonical variables of the DFA using all 10 variables (LowerRight) illustrates how well the cranial measurements discriminate the ecomorphs (modern specimens only).
Sherratt et al. www.pnas.org/cgi/content/short/1506516112 3 of 12
Leng
th (r
esid
uals
)-0.2
-0.15
-0.1
-0.05
0
0.05
0.1
0.15
AMB
J
Pelvis width
-0.1
-0.05
0
0.05
0.1
LA E
RH
M
Sacrum width
-0.25
-0.2
-0.15
-0.1
-0.05
0
0.05
0.1
A
RMJH
Pubis length
-0.25
-0.2
-0.15
-0.1
-0.05
0
0.05
0.1
0.15
0.2
A J
Sternum width
-0.15
-0.1
-0.05
0
0.05
0.1
O
A JC
Humerus
-0.2
-0.15
-0.1
-0.05
0
0.05
0.1
0.15
OA RCF JH IG
Ulna
-0.2
-0.15
-0.1
-0.05
0
0.05
0.1
L
MEC GAJFIDB H
Femur
-0.3
-0.25
-0.2
-0.15
-0.1
-0.05
0
0.05
0.1
LE MC JA F TB IGH
D
Tibia
-0.3
-0.2
-0.1
0
0.1
LE MA JC TH IGB FD
Metatarsal IV
-0.3
-0.25
-0.2
-0.15
-0.1
-0.05
0
0.05
0.1
MEC H IJ TB DA GF
Hindtoe IV
-0.25
-0.2
-0.15
-0.1
-0.05
0
0.05
0.1
0.15
LME H JC ID FA
Hindtoe III
-0.25
-0.2
-0.15
-0.1
-0.05
0
0.05
0.1
0.15
C H JO
Foretoe III
-0.25
-0.2
-0.15
-0.1
-0.05
0
0.05
0.1
CG
H J O
Foretoe IV
Leng
th (r
esid
uals
)Le
ngth
(res
idua
ls)
Leng
th (r
esid
uals
)Le
ngth
(res
idua
ls)
EcomorphGB TR TC TG TW
Amber EcomorphGB TR TC TG TW
Amber EcomorphGB TR TC TG TW
Amber
EcomorphGB TR TC TG TW
Amber EcomorphGB TR TC TG TW
Amber EcomorphGB TR TC TG TW
Amber
EcomorphGB TR TC TG TW
Amber EcomorphGB TR TC TG TW
Amber EcomorphGB TR TC TG TW
Amber
EcomorphGB TR TC TG TW
Amber EcomorphGB TR TC TG TW
Amber EcomorphGB TR TC TG TW
Amber
EcomorphGB TR TC TG TW
Amber-10
-8
-6
-4
-2
0
2
4
6
Can
onic
al 2
-10 -5 0 5 10Canonical 1
DFA, All postcranial variables, modern only
Fig. S4. Variation among the fossils and how it corresponds with variation among the ecomorphs for all 13 postcranial measurements. Size-corrected residualsare plotted for the amber (A–T) and modern specimens. Amber fossils are colored according to the discriminant function analysis (DFA) results (Table S3) whenassigned with a probability greater than 0.90. A plot of the first two canonical variables of the DFA using all 13 variables (Lower Right) illustrates how well thepostcranial measurements discriminate the ecomorphs (modern specimens only).
Sherratt et al. www.pnas.org/cgi/content/short/1506516112 4 of 12
Fig. S5. Summary tree showing the phylogenetic relationships of the amber fossils inferred by parsimony analyses. Names of Anolis subclades are given on theright. The amber fossils are represented by letters corresponding to those in Fig. 2, and their relationships are indicated by yellow-orange branches; brokenbranches indicate the prune and graft results of alternative placements. DFA classifications for the fossils are indicated by pie charts, colored by ecomorph (onlyprobabilities greater than 0.1 are shown).
Sherratt et al. www.pnas.org/cgi/content/short/1506516112 5 of 12
foretoe IIIforetoe IV
hindtoe IV
hindtoe III
frontal w
max l
prf w
skull w
jugal horbit d
mand lmand w
snout l
pmx w
2mm
tibia l
femur l
fused
sacrum w
metatarsal IV
hindtoe IV
humerus l
radius l
Postcranial
A. chlorocyanus (MCZ R-187268)47mm SVL (trunk-crown)
Cranial
Lamella Number
fused
hindtoe III
foretoe IIIforetoe IV
2mmA. baleatus (MCZ R-186663)49mm SVL (crown-giant)
unfused
unfusedb
a
A. chlorocyanus (MCZ R-59788)
A. coelestinus (MCZ R- 187469)
A. coelestinus (MCZ R- 187469)
Fig. S6. Schematic of the morphometric measurements and meristic counts taken from fossil and recent anole specimens shown on an adult trunk-crownanole (A). A juvenile crown-giant anole (B) of approximately the same SVL. Red arrows point to where there are clearly unfused (B) and fused (A) epiphyses anddiaphyses of fore and hindlimb bones, which allow the identification of juveniles vs. adults. Specimens from Museum of Comparative Zoology Reptile col-lection, Harvard University (prefix MCZ R).
Sherratt et al. www.pnas.org/cgi/content/short/1506516112 6 of 12
Table
S1.
The20
fossils
exam
ined
inthis
study
Specim
enSp
ecim
enID
Elem
ents
preserved
Specim
enlocation
Notesonspecim
enLo
calityoforigin
Scan
ned
wherean
dbywhom
kVμA
Voxe
lsize
(mm)
Rotation
step
(o)
Exposure
(ms)
AAMNH
DRSH
1W
hole
body
American
Museum
ofNaturalH
istory,
New
York
CuratorDav
idGrimaldi
Dominican
Rep
ublic
Harva
rdCen
trefor
Nan
oscaleSy
stem
sMaterialSynthesis
Facility(CNS);
EmmaSh
erratt
(ES)
&Jasm
ine
Casart
7012
00.02
670.11
1,00
0
BI
Whole
body
Privateco
llection
Owned
byMarily
nHalonen
and
Michae
lCusanovich
of
Tuscon,Arizo
na
Dominican
Rep
ublic
CNS;
D.Lu
keMah
ler
5518
50.04
520.25
—
CII
Whole
body
Privateco
llection
Owned
byJim
Work
ofCen
tral
Point,
Oregon
Dominican
Rep
ublic
CNS;
ES65
195
0.06
090.11
1,00
0
DIII
Whole
body
Privateco
llection
Jose
Calbetoof
Guay
nab
o,Pu
erto
Rico
Dominican
Rep
ublic
CNS;
ES55
165
0.01
490.11
1,41
5
EM-106
2Pa
rt(hindlim
bsan
dtail)
Privateco
llection
Owned
byEttore
MoroneofTo
rino,
Italy
Dominican
Rep
ublic
Imag
ingan
dAnalysisCen
tre,
NaturalHistory
Museum
London
(IAC-N
HM);ES
&RussellGarwood
(RG)
105
195
0.01
070.06
354
FM-115
3W
hole
body
Privateco
llection
Owned
byEttore
MoroneofTo
rino,
Italy
Dominican
Rep
ublic
IAC-N
HM;ES
&RG
105
195
0.01
350.06
354
GM-341
0W
hole
body
Privateco
llection
Owned
byEttore
MoroneofTo
rino,
Italy
Dominican
Rep
ublic
IAC-N
HM;ES
&RG
105
195
0.03
300.06
354
HM-525
Whole
body
Privateco
llection
Owned
byEttore
MoroneofTo
rino,
Italy
Dominican
Rep
ublic
IAC-N
HM;ES
&RG
105
195
0.02
100.06
500
INMBA
Entom
P52
Whole
body
Naturhistorisches
Museum,Basal
Switze
rlan
d
Obtained
byC.
Baroni-Urban
i,NMBA,Basel
LaTo
camine,
Cordillera
Septentrional,
Dominican
Rep
ublic
Synch
rotron
Rad
iationFa
cility
(ESR
F)in
Grenoble,Fran
ce,
Ren
audBoistel&
AnthonyHerrel
——
0.00
746
——
JOAAAA
Whole
body
Privateco
llection
Supplie
dbyMarco
Greco
(Ambras
Greco
SAS)
of
Mila
n,Italy
Dominican
Rep
ublic
CNS;
ES75
130
0.02
370.11
1,00
0
Sherratt et al. www.pnas.org/cgi/content/short/1506516112 7 of 12
Table
S1.
Cont.
Specim
enSp
ecim
enID
Elem
ents
preserved
Specim
enlocation
Notesonspecim
enLo
calityoforigin
Scan
ned
wherean
dbywhom
kVμA
Voxe
lsize
(mm)
Rotation
step
(o)
Exposure
(ms)
KSM
NSDo-
4871
-MPa
rt(hea
d)
Staa
tliches
Museum
fürNaturkunde
Stuttgart,
German
y
Donated
byGeo
rgDommel
of
Düsseldorf,
German
yin
1985
Dominican
Rep
ublic
CNS;
ES50
140
0.00
770.11
1,41
5
LSM
NSDo-
4886
-BPa
rt(tail,hindlim
b)
Staa
tliches
Museum
fürNaturkunde
Stuttgart,
German
y
Purchased
in19
85from
ErichBey
na,
Santo
Domingo,
Dominican
Rep
ublic
Dominican
Rep
ublic
CNS;
ES50
180
0.01
240.11
1,00
0
MSM
NSDo-
5566
-XPa
rt(trunk,
forelim
b,
hindlim
bs)
Staa
tliches
Museum
fürNaturkunde
Stuttgart,
German
y
Purchased
1991
from
D.M.Sa
lem,
8950
Sunrise
Lake
Blvd.80
/311
,Su
nrise,FL
3332
2,USA
Dominican
Rep
ublic
CNS;
ES50
130
0.02
560.11
1,41
5
NSM
U74
976
Part
(hea
d)
ShulerMuseum
of
Paleontology,
South
Methodist
University,Dallas
TX
Donated
byWilliam
S.Lo
weof
Granbury
Texa
s
Dominican
Rep
ublic
University
ofTe
xas
High-Resolution
X-ray
CTFa
cility,
RichKetch
am
8020
00.06
800.13
—
OUSN
M58
0060
Part
(hea
d,forelim
b)
Smithsonian
Institution
National
Museum
ofNaturalH
istory,
WashintonDC
Purchased
from
AmbericaWest
LaTo
camine,
Cordillera
Septentrional,
Dominican
Rep
ublic
University
ofTe
xas
High-Resolution
X-ray
CTFa
cility,
Matthew
Colbert
180
133
0.01
270.23
—
““
““
““
CNS;
ES60
165
0.00
510.18
1,00
0P
M-209
6Pa
rt(trunkan
dforelim
bs)
Privateco
llection
Owned
byEttore
MoroneofTo
rino,
Italy
Dominican
Rep
ublic
IAC-N
HM;ES
&RG
105
195
0.01
680.06
354
QRAAA
Part
(forelim
bs)
Privateco
llection
Supplie
dbyMarco
Greco
(Ambras
Greco
SAS)
of
Mila
n,Italy
Dominican
Rep
ublic
CNS;
ES75
130
0.00
890.11
1,00
0
RM-301
6Pa
rt(forelim
b,torso)
Privateco
llection
Owned
byEttore
MoroneofTo
rino,
Italy
Dominican
Rep
ublic
IAC-N
HM;ES
&RG
105
195
0.00
920.06
354
SM-127
3Pa
rt(forelim
bs)
Privateco
llection
Owned
byEttore
MoroneofTo
rino,
Italy
Dominican
Rep
ublic
IAC-N
HM;ES
&RG
105
195
0.00
680.06
354
TSM
NSDo-
5742
Part
(body,
nohea
d)
Staa
tliches
Museum
fürNaturkunde
Stuttgart,
German
y
Donated
byGeo
rgDommel
of
Düsseldorf,
German
yin
1980
Dominican
Rep
ublic
CNS;
ES50
165
0.03
080.11
1,41
5
Inform
ationisprovided
onstateofpreservation(elemen
tspreserved
);cu
rren
tlocationan
dlocalityoforigin
(includingthemine,
wherekn
own);an
dX-ray
microco
mputedtomographyscan
settings,with
inform
ationonwherethescan
was
donean
dbywhom.F
ossilO,o
riginally
scan
ned
atUniversity
ofTe
xas,was
rescan
ned
atHarva
rdat
ahigher
resolutionforthisproject
(datafrom
thehigher
resolutionscan
wereusedforthemorphometrican
dphylogen
etic
analyses).Th
reefossils
previouslywerepublished
:A(18),I(15
),an
dN(19).
Sherratt et al. www.pnas.org/cgi/content/short/1506516112 8 of 12
Table
S2.
Mea
suredan
destimated
bodysize
(SVL),morphometricva
riab
les(inmm),an
dmeristicco
unts
take
nfrom
each
amber
specim
enan
dusedin
thediscrim
inan
tfunction
analyses
(DFA
s)presentedin
Table
S3
Specim
enA
BC
DE
FG
HI
JK
LM
NO
PQ
RS
T
Mea
suredSV
L23
.830
.052
.830
.225
.723
.225
.423
.1SV
Lestimatefrom
lumbar
length
25.7
27.1
24.4
24.2
31.0
31.0
26.3
SVLestimatefrom
ilium
length
27.6
53.6
17.7
21.8
23.0
21.8
29.6
25.1
SVLestimatefrom
bodylength
24.8
SVLestimatefrom
man
dible
length
28.2
27.7
27.5
24.2
21.1
21.5
20.6
21.2
19.7
SVLestimatefrom
other
(see
notes)
19.4–21
.019
.9–21
.718
.5–20
.025
.2–44
.719
.9–28
.2Man
dible
length
8.63
8.46
8.40
7.40
6.43
6.97
6.57
6.29
6.57
6.00
Prem
axillalength
1.23
1.06
1.29
0.89
0.97
1.05
0.96
1.02
Max
illalength
4.20
5.40
5.27
4.52
4.13
3.91
3.55
3.79
3.64
Skullwidth
4.13
4.94
4.88
3.80
4.23
3.80
3.64
3.63
3.70
Frontalwidth
betwee
norbits
0.64
0.77
0.54
0.51
0.54
0.39
0.35
0.53
Prefrontalwidth
2.89
2.91
2.80
2.46
2.47
2.19
2.58
Orbitdiameter
3.38
3.47
3.50
3.31
2.84
3.04
2.96
2.90
2.90
Snoutlength
3.65
3.81
3.73
3.01
2.59
2.73
2.15
2.51
2.35
Man
dible
width
3.53
6.80
4.12
4.20
3.24
3.38
3.30
3.13
3.15
Jugal
height
1.92
1.81
1.58
1.52
1.68
1.50
1.36
1.54
1.62
Sternum
width
2.29
1.82
Humeruslength
4.38
9.00
3.17
3.20
3.48
4.64
3.40
Ulnalength
3.62
7.80
3.72
2.95
2.72
2.79
2.60
3.03
2.57
4.18
3.13
2.70
ForetoeIV
length
4.83
1.64
2.47
2.16
1.93
2.18
1.77
2.46
1.88
ForetoeIII
length
5.17
1.90
2.83
2.43
2.40
2.33
2.03
2.93
2.18
Pelviswidth
2.38
2.25
2.12
1.47
2.46
Sacrum
width
2.00
1.51
1.51
2.52
1.92
1.62
Pubislength
2.48
1.78
1.70
2.45
1.96
Femurlength
6.05
5.70
12.45
3.23
4.85
6.26
5.95
4.56
4.88
4.74
8.04
7.15
Tibia
length
4.89
5.24
10.98
2.44
4.44
5.35
4.56
4.02
4.38
4.22
6.75
6.21
4.29
MetatarsalIV
length
3.37
3.16
6.40
1.73
2.72
3.04
2.92
2.74
2.83
2.80
3.88
3.80
2.84
HindtoeIV
length
4.72
5.16
9.80
3.10
4.38
4.89
4.63
4.54
4.61
4.13
6.34
4.42
HindtoeIII
length
2.53
5.54
1.66
2.31
2.82
2.56
2.49
2.41
3.65
3.30
HindtoeIV
lamellae
1416
1816
17HindtoeIII
lamellae
2326
1618
2321
1923
17Fo
retoeIV
lamellae
1617
1518
2121
ForetoeIII
lamellae
1313
1620
20
Reg
ressionmodelsofmodernspecieswereusedto
estimateSV
Lofthefossils
from
fourlog-transform
edva
riab
les:thelength
ofalumbar
verteb
ra;length
oftheilium;bodylength;an
dman
dible
length.
Detailsan
deq
uationsaregiven
inMethods.Allsquared
correlationco
efficien
tsare0.96
,excep
tbodylength,w
hichis0.99
.SVLestimates
inbold
arethose
usedin
theDFA
s.W
hen
more
than
oneestimatewas
available,theva
luech
osenwas
decided
acco
rdingto
thepreservationofthefossil.
Rows1–
6providearangeofSV
Lestimates:specim
enA
broke
n,mea
suredSV
Lan
underestimate;
specim
enCinco
mplete,
mea
suredSV
Lan
underestimate;
specim
enD
inco
mplete,ilium
frag
men
tpreserved
;specim
enG
inco
mplete,mea
suredSV
Lan
underestimate;
specim
enH
broke
n,m
easuredSV
Lan
underestimate;
specim
enK,
other
SVLestimated
from
rangeofecomorph-specificslopes
forman
dible
length;specim
enN,o
ther
SVLestimated
from
rangeofecomorph-specificslopes
forman
dible
length;specim
enO,o
ther
SVLestimated
from
rangeofecomorph-specific
slopes
forman
dible
length;specim
enP,
other
SVLestimated
from
rangeofecomorph-specific
slopes
forulnalength
(r2=0.84
);specim
enS,
other
SVLestimated
from
rangeof
ecomorph-specific
slopes
forulnalength
(r2=
0.84
).
Sherratt et al. www.pnas.org/cgi/content/short/1506516112 9 of 12
Table S3. Results of discriminant function analyses (DFAs) using size-corrected residuals of the morphometric variables (details of thesize correction in Table S2)
Specimen TC TR TG TW GB λ Approximate F (df) Percent misclassified
A 1.00 0.00002 40.50 (84, 227.59) 0B 0.96 0.00099 35.25 (44, 304.19) 0C 1.00 0.00018 50.11 (44, 258.28) 0D 0.99 0.00256 28.33 (36, 260.31) 1.22E 1.00 0.00148 46.78 (28, 257.42) 0F 1.00 0.00004 41.46 (72, 238.29) 0G 0.69 0.31 0.00004 44.46 (68, 241.70) 0H 0.99 0.00002 40.26 (84, 227.59) 0I 0.99 0.00037 45.59 (40, 259.70) 0J 1.00 0.00004 34.31 (88, 243.69) 0K 0.82 0.18 0.01746 18.97 (32, 304.00) 7.4
[0.81] [0.18] 0.02303 16.91 (32, 304.00) 13.8L 0.98 0.00227 49.70 (24, 252.39) 1.22M 1.00 0.00075 36.86 (40, 259.70) 2.4N 0.86 0.12 0.02797 18.21 (28, 300.68) 5.3
[0.52] [0.48] 0.03315 16.88 (28, 300.68) 10.6O 1.00 0.00122 26.99 (48, 275.54) 1.1
[0.99] 0.00183 23.75 (48, 275.54) 1.1P {0.13) {0.49} {0.34} 0.22769 20.81 (8, 152.00) 42.7Q {0.31} {0.13} {0.20} {0.35} 0.22769 20.81 (8, 152.00) 42.7R 0.94 0.00626 43.90 (20, 243.06) 3.7S {0.60} {0.35} 0.22769 20.81 (8, 152.00) 42.7T 0.99 0.04449 43.50 (12, 233.12) 24.2
Probabilities of classification to the five ecomorphs are presented, with a post hoc Wilks’ λ test of the null hypothesis that the means of all of theindependent variables are equal across groups (F statistic and degrees of freedom), and proportion of individuals misclassified in each analysis. Probabilitiesbelow 0.1 are not shown. Each DFA used a different set of variables specific to the fossil (Table S2). For three fossils that comprised an isolated skull (K, N, andO), a second DFA using raw (not size-corrected) variables was conducted, the results of which are given in brackets. For fossils P, Q, and S, only lamella countswere available for the DFA, so these results are given in braces. Bold indicates DFA probabilities above 0.90.
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Table
S4.
Resultsofparsimonyan
dBay
esianphylogen
etic
analyses
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Movie S1. Photographs and animated 3D renderings of the 20 amber fossils, in order from A to T. For each fossil, a photograph of the specimen is shownfollowed by a 3D reconstructed skeleton and/or soft tissue animation from X-ray micro-CT data. In specimens L and M, there are ant inclusions (genus Azteca)that have been false-colored brown and rendered opaque.
Movie S1
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