Phylogenetic Interpretation Dr Laura Emery Laura.Emery@ebi.ac.uk www.ebi.ac.uk

Objectives After this tutorial you should be able to… Discuss the impact of a range of biological phenomena upon phylogenetic inference Appreciate some challenges and limitations of phylogenetic approach Interpret published phylogenies (and your own) We don't have enough time to go into any of these methods in great depth, and if you want to fully understand how they work, then you will need to go away and do some extra reading. But the point of this tutorial is to make you aware of some of the most commonly available methods, their assumptions and limitations. And also to direct you to information that you can read up on if you want further help

Phylogenetic interpretation is essential throughout data analysis Data assessment - known biology - additional data (e.g. geography) Formulate hypotheses Decide upon and implement method Phylogenetic Result(s) Investigate unexpected and unresolved aspects further - consider including more data Answered your question? No No We often get asked: how should I interpret my own data? Data analysis is more complex then many lab biologists expect Every stage in this process requires the ability to interpret phylogenies Caveat: Finished versus unfinished analyses and their interpretation Most of the examples we will look at today have been fully analysed When you come to analyse your own data, initially things will not look so straightforward since data analysis is an iterative process. Nevertheless the concept you will learn in this lecture still apply. The chances are that you will be able to reach fewer conclusions when you first come to analyse your own data. Please be patient and persistents Yes Can you validate this? Final phylogeny and analysis Yes

Phylogenetic interpretation skill set Tree-thinking skills Revise: relatedness, trait evolution, confidence, homology Knowledge of phylogenetic methods and their limitations Knowledge of biological processes affecting sequence evolution gene duplication, recombination, horizontal gene transfer, population genetic processes, and many more! Knowledge of the data you wish to interpret Covered in introduction to phylogenies 1. We will be introducing a few more tree thinking skills but we dealt with these in the first session with this morning 2. It is important to know that phylogenetic methods to the extent that you know what assumptions they are making and their limitations, and you know how to interpret relevant statistical information 3. Knowledge of biology-that's it will cover in this session. Will do this by examining case studies based upon real data, so you can see what kind of trees and patterns of evolution produced from these processes 4. Knowledge of the data you are attempting to interpret

Recap of tree-thinking skills Relatedness Trait evolution Confidence Homology

1. Relatedness: taxa that share a more recent common ancestor are more closely related most recent common ancestor shared with second cousin most recent common ancestor shared with first cousin You are more closely related to your first cousin than you are to your second cousin because the most recent common ancestor that you share with your first cousin i.e. your grandmother was born more recently than the most recent common ancestor that you share with your second cousin (i.e. your great-grandmother)

2. Trait evolution It can be useful to map traits onto phylogenies as a first step in inferring their evolutionary histories Interpreting trait evolution in its phylogenetic context is rarely straightforward! Assumptions must be made regarding the loss and gain of traits It is often useful to construct alternative scenarios Then we have to decide upon the most plausible (character state methods e.g. MP and ML can be applied)

Example: The Evolution of Mitochondria origin of eukaryotes Ginger et al. 2010

Example: The Evolution of Mitochondria G = gain L = loss G G G G G G G G G G G G origin of eukaryotes Ginger et al. 2010 Scenario one: Mitochondria evolved from mitosomes

Example: The Evolution of Mitochondria G = gain L = loss G L G L G L G L origin of eukaryotes Ginger et al. 2010 Scenario two: Mitochondria occurred at the origin of eukaryotes

3. Tree Confidence Question Does this tree support the grouping of pelecaniforms and ciconiiforms as a monophyletic group?

4. Homology is similarity due to shared ancestry Example: limbs and wings Limbs are homologous they share a common ancestor Wings are not homologous they are an analogous as they have evolved similarity independently

Homology Question: Trap-jaws in ants Based on this phylogeny, which scenario do you think is more likely? trap-jaws are homologous trap-jaws are analogous and have evolved independently four times Convergent evolution (analogy rather than homology). Trap-jaws are believed to have evolved independently at least four times Moreau et al. 2006

Homology Question: Trap-jaws in ants Based on this phylogeny, which scenario do you think is more likely? trap-jaws are homologous trap-jaws are analogous and have evolved independently four times L L L L Convergent evolution (analogy rather than homology). Trap-jaws are believed to have evolved independently at least four times G L L L Moreau et al. 2006 Scenario one: Trap-jaws are homologous

Homology Question: Trap-jaws in ants Based on this phylogeny, which scenario do you think is more likely? trap-jaws are homologous trap-jaws are analogous and have evolved independently four times G G Trap-jaws or an evolutionary adaptation that allow ants to shut their jaws at 145mph to attack prey and escape predation, as it propels them into the air! Convergent evolution (analogy rather than homology). Trap-jaws are believed to have evolved independently at least four times G more parsimonious Moreau et al. 2006 Scenario two: Trap-jaws are analogous

Phylogenetic interpretation skill set Tree-thinking skills Revise: relatedness, trait evolution, confidence, homology Knowledge of phylogenetic methods and their limitations Knowledge of biological processes affecting sequence evolution gene duplication, recombination, horizontal gene transfer, population genetic processes, and many more! Knowledge of the data you wish to interpret Covered in introduction to phylogenies 1. We will be introducing a few more tree thinking skills but we dealt with these in the first session with this morning 2. It is important to know that phylogenetic methods to the extent that you know what assumptions they are making and their limitations, and you know how to interpret relevant statistical information 3. Knowledge of biology-that's it will cover in this session. Will do this by examining case studies based upon real data, so you can see what kind of trees and patterns of evolution produced from these processes 4. Knowledge of the data you are attempting to interpret

Processes that affect sequence evolution Gene/genome duplication and divergence Recombination Horizontal gene transfer Coevolution Migration Rate and time of divergence Other

1. Gene duplication Gene duplication and subsequent divergence can result in novel gene functions (it can also result in pseudogenes) Genes that are homologous due to gene duplication are paralogous Genes that are homologous due to speciation are orthologous

Gene duplication question This is a tree of gene family that has undergone one gene duplication event in its evolutionary past. Where on the tree did this occur? Is the event well-supported? SPARC and SPARCL1 Cells Tissues & Organs 2007

2. Recombination Single or small numbers of events: Within genes Between genes Where there is extensive recombination - a phylogenetic approach is inappropriate (not tree-like) Cut and pasting of gene/genome segments

Recombination example: Dengue-2 virus Strain highlighted in red box is a recombinant Others highlighted are two strains from the same mosquito data from E. Holmes, figure from A. Rambaut

Recombination Question Can you spot the recombinant strain? Mauro et al 2003

3. Horizontal Gene Transfer (HGT/LGT) Horizontal gene transfer violates the assumption that sequences have evolved in a tree-like manner Where sparse, can be detected by comparing with species phylogeny Where extensive, phylogenetic approach is inappropriate Gogarten & Townsend 2005

Phylogenetics is not appropriate for highly recombinant taxa Phylogenetics assumes that patterns of relatedness among taxa follow a tree- like structure Recombination and horizontal gene transfer produce networks Avoid phylogenetics for: Intraspecific sexual species (recombination at each meiosis) Asexual species with extensive HGT (e.g. some Bacteria)

Horizontal gene transfer question Can you spot the horizontally transferred gene? Bombyx is a silk moth

4. Coevolution Where parasites or symbionts co-evolve with their hosts, both topologies are expected to be very similar. The parasites are inherited vertically Weiss 2009 from Reed et al 2007

Coevolution Question Do these phylogenies provide evidence that the lice are inherited vertically? Hafner & Nadler 1988

6. Migration Patterns of migration influence phylogenetic topology, especially in structured populations

Phylogeography example: Chimpanzees P. troglodytes and P.schweinfurthii are more dissimilar than you would expect given their proximity > Chimpanzees can't cross rivers! Gao et al 1999

Migration Question What can you infer about patterns of migration of the Taiwanese stag- beetle based upon this phylogeny? Black = Taiwan Assuming even sampling, it looks as the beetle originated in China and has radiated outwards

5. Rate and time of divergence Phylogenies can be used to date divergence times when some temporal information is known e.g. carbon dating from fossil evidence e.g. dates of sample isolation Genetic change = Evolutionary rate x Divergence time (substitutions/site) (substitutions/site/year) (years) If all lineages evolve at the same rate (i.e. there is a molecular clock) then branch lengths should reflect divergences times C D E A B

Is there a molecular clock? Zuckerland and Pauling (1962) No. substitutions in haemoglobin roughly proportional to time based upon fossil datings

Dating divergence with a molecular clock X d = genetic distance (branch length) We know time T since a and c diverged We want to find out time X since a and b diverged Use T to estimate the evolutionary rate r r = d(a-c) / 2T Use r to estimate time X X = 1/2 (d(a-b) / r)

Dating Drosophila Divergence around Hawaii The volcanic activity around Hawaii has produced a chain of islands; the oldest is furthest away from the mainland Several species including Drosophila have diverged with island formation Figure Andrew Rambaut from Fleischer, McIntosh &Tarr 1998

Dating Drosophila Divergence in Hawaii Island formation dates reflecting species’ divergence were plotted against genetic distance (branch length) Genetic distance scaled linearly with divergences date, indicating the presence of a molecular clock Genetic distance gradient = evolutionary rate NB: Not all species exhibit a molecular clock! Time Fleischer, McIntosh &Tarr 1998

7. Other biological processes can complicate molecular analyses Population genetic processes Epidemiological processes Gene conversion Codon bias Hypermutable sites Concerted evolution Reassortment Many more…

Summary: Phylogenetic interpretation skill set Tree-thinking skills Revise: relatedness, trait evolution, confidence, homology Knowledge of phylogenetic methods and their limitations Knowledge of biological processes affecting sequence evolution gene duplication, recombination, horizontal gene transfer, population genetic processes, and many more! Knowledge of the data you wish to interpret Covered in introduction to phylogenies 1. We will be introducing a few more tree thinking skills but we dealt with these in the first session with this morning 2. It is important to know that phylogenetic methods to the extent that you know what assumptions they are making and their limitations, and you know how to interpret relevant statistical information 3. Knowledge of biology-that's it will cover in this session. Will do this by examining case studies based upon real data, so you can see what kind of trees and patterns of evolution produced from these processes 4. Knowledge of the data you are attempting to interpret

Further Reading Molecular Evolution: A Phylogenetic Approach (1998) Roderic D M Page & Edward C Holmes, Blackwell Science, Oxford. The Phylogenetic Handbook (2003), Marco Salemi and Anne-Mieke Vandamme Eds, Cambridge University Press, Cambridge. Inferring Phylogenies (2003) Joseph Felsenstein, Sinauer. Molecular Evolution (1997) Wen-Hsiung Li , Sinauer

Train online www.ebi.ac.uk/training/online Free online courses Learn in your own time, at your own pace Created for life- science researchers No previous knowledge of bioinformatics needed www.ebi.ac.uk/training/online

Acknowledgements People Funding EMBL member states and… Andrew Rambaut (University of Edinburgh) …and the EBI training team Paul Sharp (University of Edinburgh) Nick Goldman (EMBL-EBI) Benjamin Redelings (Duke University) Brian Moore (University of California, Davis) Olivier Gascuel (University of Montpelier) Aiden Budd (EMBL-EBI) Funding EMBL member states and…

Thank you! www.ebi.ac.uk Twitter: @emblebi Facebook: EMBLEBI

Now it's your turn… Open your tutorial manual and begin Tree-thinking quiz 2 (appendix 2) The manual is available to download from: http://www.ebi.ac.uk/training/course/scuola-di-bioinformatica- 2013 When you are finished you can mark your own. Remember to ask for help at any stage!