Complex Molecular Evolutionary
Models and Information Theoretic
Approaches Provide Genomic
Perspectives on Amphibian Evolution
Paul M. Hime
16 May, 2017
Blue Waters Symposium
[email protected]
The Evolution of Life on Earth
• All life traces its origins back
to a single common ancestor
nearly 4 billion years ago
• But today, there are tens of
millions of species!
• Reconstructing the genealogy
of life is fundamental to nearly
all areas of modern biology.
The Evolution of Life on Earth
“Nothing in biology makes sense, except in light of evolution”
Dobzhansky
“Nothing in evolutionary biology makes sense, except in light of phylogeny”
All Organisms on Earth Trace Their Origins
Back to a Single Common Ancestor
Genomes Are Documents of
Evolutionary History
Organisms’ Genomes Evolve through Time
Phylogenetic Reconstruction
• Phylogenies are hypotheses about
ancestor - descendent relationships.
• These can be estimated from genetic
data (in the context of a model).
• Simple case: enumerate all possible
trees, pick the “best”.
• Tree space explodes factorially with
increasing numbers of taxa.
• Use heuristic search strategies to
explore tree- and parameter-space.
Models in Evolutionary Biology
• Evolutionary biology is an inherently historical discipline.
• In evolutionary biology, one cannot “replay the tape” of life...
• We use statistical approaches to compare competing sets of
models, in the light of data which we collect.
“All models are wrong. Some are useful.”
George Box
Data ≠ Information
(Except in the Context of an Appropriate Model)
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Data ≈ Information
(Except in the Context of an Appropriate Model)
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ACCGAGGGCATCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTACGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCATCGATCGACTACCTTAGGGCTCTAGCCTGTTACGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTACGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTACGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCATCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTCCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCATCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTCCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCATCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCCTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTCCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCCTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTGCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCCTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTGCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
*
*
informative site
informative site
Models of Nucleotide Substitution
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ACCGAGGGCATCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTACGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCATCGATCGACTACCTTAGGGCTCTAGCCTGTTACGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTACGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTACGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCATCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTCCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCATCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTCCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCATCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCCTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTCCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCCTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTGCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCCTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTGCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
ACCGAGGGCTTCGATCGACTACCTTAGGGCTCTAGCCTGTTTCGTGCTAGCTGACTGATCGTAGTGTAGCTGACTGTGTG
*
*
informative site
informative site
Multi-sequence Alignment
General time-reversible
substitution matrix
Gamma-distributed rate
heterogeneity across sites
(discretized in practice)
Codon-Based Models of Molecular Evolution
The Multispecies Coalescent Model
Gene divergences always predate species divergences
Stochastic coalescent processes can lead to gene tree / species tree discordance
Modified from Leliaert et al. 2014
Gene Tree - Species Tree Discordance
Population-Level Processes Affect The
Expected Distributions of Gene Coalescence
Luay Nakhleh
Conflicting phylogenetic signal from different loci is expected, especially
for more recent divergence events and large effective population sizes.
What about at deep scales?
Many loci (regions of the genome) may be needed for difficult questions.
The Genomics Revolution in
Evolutionary Biology
It has never been
easier to collect
genomic data in
non-model
organisms.
The Genomics Revolution in
Evolutionary Biology
It has never been
easier to collect
genomic data in
non-model
organisms.
It has never been
easier to collect
genomic data in
non-model
organisms.
Data ≠ Information
(Except in the Context of an Appropriate Model)
Data ≠ Information
(Except in the Context of an Appropriate Model)
Data ≠ Information
(Except in the Context of an Appropriate Model)
Data ≠ Information
(Except in the Context of an Appropriate Model)
Some of the Many Tradeoffs in Phylogenomics
The genomic revolution is now offering unprecedented
opportunities to tackle thorny questions in evolutionary biology.
But these opportunities bring analytical and computational costs.
Some of the Many Tradeoffs in Phylogenomics
The genomic revolution is now offering unprecedented
opportunities to tackle thorny questions in evolutionary biology.
But these opportunities bring analytical and computational costs.
Genomic perspectives on the
amphibian tree of life
• Amphibians provide a rich system for testing phylogenetic
hypotheses at both deep and shallow scales of divergence.
• What is the topology of the amphibian Tree of Life???
• How confident are we in that topology?
• What are the inter-ordinal relationships?
• What is the nature of support across the genome?
• What are “best” practices with large phylogenomic data sets???
Extant Amphibian Diversity
Amphibians - 7,660 species as of 28 April, 2017 (http://AmphibiaWeb.org)
The three amphibian orders likely diverged 300+ MYA.
Map from BiodiversityMapping.org
Extant Amphibian Diversity
Caecilians (Gymnophiona) - 205 species, 33 genera, 10 families
Map from BiodiversityMapping.org
Extant Amphibian Diversity
Salamanders (Caudata) - 695 species, 68 genera, 10 families
Map from BiodiversityMapping.org
Extant Amphibian Diversity
Frogs and Toads (Anura) - 6,760 species, 448 genera, 55-65 families
Map from BiodiversityMapping.org
Amphibian Relationships
Hundreds of studies have addressed phylogenetic affinities of amphibians...
Amphibian Relationships
Hundreds of studies have addressed phylogenetic affinities of amphibians...
This project aims to sample hundreds
of nuclear genes for hundreds of
amphibian species
Inter-Ordinal Amphibian Relationships
Three main hypotheses for phylogenetic relationships among the
three amphibian orders (assuming that Amphibia is monophyletic...)
Batrachia
Acauda
Procera
The potential resolutions of these deep branches each have very
different implications for our understanding of amphibian evolution.
Inter-Ordinal Amphibian Relationships
•
There are 15 possible topologies for the three amphibian orders and
Inter-Ordinal Amphibian Relationships
amniotes,
assuming that Amphibia may or may NOT be monophyletic..
Three main hypotheses for phylogenetic relationships among the
three amphibian orders (assuming that Amphibia is monophyletic...)
Batrachia
Acauda
Procera
These three orders likely diverged ~300 million years ago.
Taxonomic Sampling across Amphibia
• We targeted 325 amphibian taxa (296 “worked”).
• 276 genera (> 50% of recognized genera)...
• 96% of recognized families (and most subfamilies)...
• Taxa were sampled roughly in proportion to species richness.
Taxonomic Sampling across Amphibia
• We targeted 325 amphibian taxa (296 “worked”).
• 276 genera (> 50% of recognized genera)...
• 96% of recognized families (and most subfamilies)...
• Taxa were sampled roughly in proportion to species richness.
• Multiple outgroups were included to root the phylogeny.
Anolis
Chrysemys
Gallus
Homo
Latimeria
Generating Genomic Data in Amphibians
• We developed an amphibian-specific gene capture system which
targets 366 semi-conserved nuclear exons.
• Probes were designed from genomic/transcriptomic* resources for:
1 Caecilian:
Ichthyophis
4 Salamanders:
Ambystoma*
Desmognathus
Cryptobranchus*
Notophthalmus*
6 Frogs:
Gastrophryne
Ascaphus
Pseudacris
Mixophyes
Rana
(Lithobates)
Xenopus
(Silurana)
Nuclear Gene Tree Estimation
•
Independently estimated ML gene trees for all genes in RAxML.
•
Used separate (best-fit) partitioning schemes and nucleotide
substitution models for each gene (GTR+G).
•
Conducted 500 non-parametric bootstrap replicates (across sites) to
assess “support” for branches in these gene trees.
•
Identified outlier taxa for each gene (and removed those taxa).
•
Conducted analyses with unconstrained and constrained topological
backbones.
Species Tree Estimation
•
Use multiple algorithms to estimate the topology of the species tree.
•
“Shortcut” methods attempt to reconcile collections of gene trees into
an estimate of the species tree (Astral and MulRF).
+
+
+
+
...
®
•
Also implemented a gene-tree-free approach (SVDQuartets).
•
Lastly, estimated a concatenated ML tree in RAxML, using a best-fit
partitioning scheme of 76 distinct partitions (separate GTR+G models).
•
Used the non-parametric bootstrap across sites to assess “support”
(sites and genes for Astral).
Brief Overview of Results
• Nearly all families are recovered as
monophyletic.
• Shallow-scale relationships are largely in line
with previous studies.
• Most higher-order clades are recovered.
• Deep branches receive strong bootstrap
support in species tree analyses...
Astral2 Species Tree
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I4421_CAS_224676_Anura_Rh acophoridae_Rhacophorus_rho
48.3333
98
33
• We’re done,
right??!!
I10935_NCBS_
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• Inter-ordinal
support is for
Batrachia
(BS = 100)
I444
I4 4
• High bootstrap
support across
the tree
_couchii_seq1
"TUSBM4QFDJFT5SFF&TUJNBUFEGSPN"ODIPSFE-PDJ"NQIJCJBBOE&BDI0SEFS$POTUSBJOFEUP#F.POPQIZM
sis_ seq1
Inter-Ordinal Support Varies across Loci
(RAxML “Best” Gene Trees)
Acauda
n = 45
Procera
n = 51
Batrachia
n = 98
Support across 500 Bootstrap Replicates
Acauda
Batrachia
Procera
Unbounded Support Using the AIC
100
90
80
∆ AIC
70
60
50
40
30
20
10
0
Acauda
Batrachia
Procera
Unbounded Support Using the AIC
100
Constrain the basal
topology, then calculate AIC
for each model...
90
80
∆ AIC
70
60
50
40
30
20
10
0
Acauda
Batrachia
Procera
Unbounded Support Using the AIC
100
Think of these as measuring
the strength of support
against alternative models.
90
80
∆ AIC
70
60
50
40
30
20
10
0
Acauda
Batrachia
Procera
Backbone of the Amphibian Tree of Life
These topologies
are largely
concordant with
previous studies...
With a few distinct
exceptions...
Backbone of the Amphibian Tree of Life
These topologies
are largely
concordant with
previous studies...
With a few distinct
exceptions...
Conflict in the Frog Tree: Nasikabatrachus
Conflict in the Frog Tree: Nasikabatrachus
Conflict in the Frog Tree: Nasikabatrachus
Different Methods Yield Different Topologies
ASTRAL Tree
RAxML Tree
*
78
100
100
100
100
100
A Framework for Testing Neobatrachian Relationships
A Framework for Testing Neobatrachian Relationships
Conclusions
• The illusion of “support” for topological hypotheses
depends on how hard one looks.
• The bootstrap can help determine the direction of support,
but may not be informative about its magnitude.
• Substantial discordance across loci exists at the base of
the amphibian tree (and may not all be noise!).
• Genomic data and new statistical models are providing
novel insights into evolutionary relationships of amphibians.
• More data ≠ easy answers (that are credible...).
Why Blue Waters?
• Rigorously testing competing topological models across
large numbers of genes is computationally demanding.
• Even embarrassingly parallel approaches (gene-by-gene
AIC) overwhelm typical HPC clusters’ resources.
• MCMC sampling for rugged likelihood surfaces can be
improved with large numbers of Metropolis coupled chains.
• For Bayes factor tests (with N taxa):
Markov chain Monte Carlo scales as N2
Hamiltonian Monte Carlo scales as N1.2
Acknowledgments
Co-Authors/Collaborators
Alan R. Lemmon
Emily C. Moriarty Lemmon
Elizabeth Scott-Prendini
Jeremy M. Brown
Robert C. Thomson
Brice P. Noonan
R. Alex Pyron
Pedro L. V. Peloso
Michelle Kortyna
Justin D. Kratovil
J. Scott Keogh
Tissue Loans from Institutions:
American Museum of Natural History (Darrel Frost, David Kizirian, Julie Feinstein)
California Academy of Sciences (David Blackburn, Jens Vindum)
Florida Museum of Natural History (Pamela Soltis)
University of Kansas Biodiversity Institute and Natural History Museum (Rafe Brown, Linda Trueb, Andrew Campbell)
Louisiana State University Museum of Natural Science (Robb Brumfield, Donna Dittmann)
Museum of Comparative Zoology (Jim Hanken, José Rosado, Breda Zimkus)
Museum of Vertebrate Zoology (Jim McGuire, Carol Spencer, Ted Papenfuss, Marvalee Wake, Sima Bouzid)
Museum Victoria (Jane Melville, Joanna Sumner)
National Museum of Natural History (Kevin De Queiroz, Addison Wynn)
South African National Biodiversity Institute (Zoe Davids)
Saint Louis Zoological Park (Jeffrey Ettling, Mark Wanner, Randall Junge)
University of Michigan Museum of Natural History (Ronald A. Nussbaum, Gregory Schneider)
Yale Peabody Museum (Gregory Watkins-Colwell)
Tissue Loans from Individuals:
J.J. Apodaca, Alan Channing, Becky Chong, Guarino Colli, Tyler Frye, S. Blair Hedges, Elizabeth Jockusch, Christopher
McNamara, Eric O'Neill, Todd Pierson, Steve Richards, Kelly Zamudio
Stephen C. Donnellan
Rachel L. Mueller
Fellowships: NSF Graduate Research Fellowship (3048109801), Blue Waters Graduate Research Fellowship (NSF/NCSA)
Christopher J. Raxworthy
Funding: SSB Graduate Student Research Award, DEB-0949532 (DWW), DEB-1601586 (DWW, PMH)
Krushnamegh Kunte
Santiago Ron
Sandeep Das
Nikhil Gaitonde
David M. Green
Jim Labisko
David W. Weisrock