Reconstructing the tree of life using molecular sequence data

Reconstructing the tree of life using molecular sequence data

John Dunbar SN: 69789839

Abstract: The Tree of Life (ToL) and the evolutionary histories of species have proved to be an extremely complex field of study. There has been much debate on how the universal ToL stands. Traditional taxonomy methods of identifying shared morphological characteristics have been beneficial to constructing phylogenetic trees and inferring the ToL. However, molecular sequence data in the 1960s revolutionised phylogenetics and has enabled us to resolve many relationships between species. Although horizontal gene transfer (HGT) made interpreting molecular sequence data very difficult, good computational analysis has made inferring the deepest roots of ToL more possible.

Table of Contents

Introduction….................................................................................................. 2 Traditional & Phylogenetic classification…………........................................ 2 Molecular sequencing and the impact of Horizontal Gene Transfer (HGT) & Genome Fusion………………………………………………………………. 4 DNA Barcoding……………………………………………………………… 7 Conclusion…………………………………………………………………… 9 References……………………………………………………………………. 10

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Reconstructing the tree of life using molecular sequence data Introduction In the publication “On the Origin of Species”, Darwin, (1859) depicted a tree like diagram in attempt to illustrate the relatedness and histories of all living organisms and demonstrate that all life descended from a common ancestor. This is known as the Tree of Life (ToL). In doing so, Darwin opened up a new world for exploration, and a century and a half later, with the aid of traditional taxonomy methods and molecular sequence data, many phylogenetic trees have been established. However, the ToL and the evolutionary histories of species have proved extremely complex and there has been much debate on how the universal ToL stands.

Traditional & Phylogenetic classification Traditional taxonomy is founded on the grouping of organisms’ in terms of classification, identification, and naming based on shared morphological characteristics. The system of naming and grouping organisms was established by Carolus Linnaeus in 1735 when he published his book “Systema Naturae” (Futuyma, 2009). The rationale at the time was to develop an understanding of the scale of life in the hope to reveal God’s plan of creation (Understanding Evolution, 2011). This system consists of first assigning a species with a two-part name, known as binomial nomenclature, i.e. a genus name followed by a species name. Secondly, species are grouped into larger groups known as taxonomic levels which are categorised starting with the highest level as: Kingdom, Phylum, Class, Order,

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Family, Genus, and Species. This is known as hierarchical classification (Futuyma, 2009).

However, the Linnaean system does not identify the organism’s position on the ToL or its evolutionary history because it ranks organisms into equivalent taxonomic levels regardless of their evolutionary history. For example, the cats (Felidae) and the orchids (Orchidaceae) are both grouped at the family level, even though Orchidaceae has an evolutionary history expanding over 70 million years longer than Felidae (Understanding Evolution, 2011).

We can demonstrate where organisms are positioned within the ToL by using methods known as phylogenetic classification. Phylogenetics is concerned with the classification of species in a way that reflects their evolutionary relatedness and classifies them into groups called clades (Fig.1) which include a species and all its descendants.

These clades are then depicted on a tree like diagram called a phylogenetic tree based on Figure 1. Demonstrates a clade on a phylogenetic tree.

Taken from (Understanding Evolution, 2011)

phenotypic characters such as internal and

external morphology and, behaviour and molecular sequence data, which will be discussed in the next chapter. A tree can be established by identifying a particular trait in a clade and determining if the trait is ancestral (plesiomorphic) or a newly derived trait (apomorphic) (Fig.2). All possessing synapomorphic traits (traits that are shared by two or more lineages within a clade) are branched together high in the tree while those believed to be more basal (primitive) and missing the trait but possessing other shared Figure 2. Demonstrates apomorphic and plesiomorphic characters .

Taken from (Understanding Evolution, 2011)

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plesiomorphic traits are placed lower on the tree indicating that they all share a common ancestor.

Often, new data will conflict with existing trees. Take, for example, the evolution of snakes, which are known to have arisen during the Cretaceous period approx. 98 million years ago (Wilson et al., 2010). Like the mammals, after the K-T extinction (an extinction event 65 million years ago) they exploded in diversity during the Tertiary (Benton, 2000).

Macrostomata, the wide gape that gives snakes the ability to swallow prey many times larger than their skull, is believed to have evolved later in the clade Alethinophidia (Lee et al., 2007). The most basal macrostomatans are Xenopeltids and the Core Macrostomata are concerned with the more recently evolved booids which include giant constricting snakes and the more advanced snakes, such as those that contain a venom apparatus (Lee et al., 2007).

This hypothesis suggests that a restricted gape in extant basal snakes is a plesiomorphic character. However, recent findings in India of a 3.5 meter long snake found within a sauropod dinosaur nest site, recovered from Upper Cretaceous rocks by Wilson et al., (2010), demonstrates that early snakes did have the ability to attain a large body size and jaw mobility did in fact allow them to subdue large prey. The finding also suggests that the restricted gape in extant basal snakes is a result of miniaturisation of habitat and prey rather than a plesiomorphic character.

Molecular sequencing and the impact of Horizontal Gene Transfer (HGT) & Genome Fusion A new era of inferring the ToL arrived in the 1960s, when molecular biology techniques allowed us to read the genealogical history of an organism from its DNA sequence (Zuckerkandl & Pauling., 1965). In contrast to the traditional taxonomic methods, sequencing 4

of an organism’s DNA revealed its evolutionary history and seemed promising in uncovering the ToL (Woese, 2000). Traditional morphological identification grouped Tardigrada into a monophyletic group with Onychophora and Arthropoda but recent large scale molecular data places Tardigrada as a sister group to Arthropoda (Campbell et al., 2011). Ribosomal RNA (rRNA) for example, is among the ideal molecules for sequencing, due to its universal function (O’Malley & Koonin, 2011). Sequencing rRNA enabled biologists to build up a comprehensive overview of historical events that become recorded in molecules and could therefore, help us infer the universal ToL (McInerney et al., 2007). However, during the 1990s, it became apparent that it wouldn’t be an easy task to construct the ToL using genomics. New data acquired from the sequencing of genomes conflicted with other molecular data and this initially seemed to cast doubt on molecular sequencing as a reliable tool for inferring the ToL. The errors in the molecular data occurred through a process known as horizontal gene transfer (HGT). HGT occurs when organisms’ genes are incorporated into another organism’s genome (Woese, 2000). The errors in interpretation arose because of the fact that HGT was not discovered back then.

HGT might initially appear to be an undesirable event, particularly for rooting trees, but in regards to cellular defence, cells appear more than capable of defending against foreign DNA (Woese, 2000), suggesting that perhaps there are advantages to acquiring new genes.

There are extant multicellular organisms in which HGT is believed to have had a significant impact on their anatomy and physiology. Take for instance, the sea slug Elysia chorotica, an animal which acquires food via photosynthesis for part of its life (Rumpho et al., 2008).

Initially E. chorotica’s nutritional source is a filamentous algae Vaucheria litorea. After eating, E. chorotica employs chloroplasts from V. litorea in the digestive apparatus to carry out photosynthesis in a suitable organ (Rumpho et al., 2008). However, this process is time limited as active photosynthesis can be carried out for approx. 10 months only. While this 5

process sounds ideal, especially during times of food scarcity, short lived proteins such as, psb0 encoded in the nucleus of V. litorea must be regularly imported by the chloroplasts in order to ensure continuous function (Skulachev, 2010). Acquiring the chloroplasts from algae is not enough to carry out photosynthesis alone, the presence of psb0 genes are critical and E. chorotica contains the psb0 genes which is identical, to that of V. litorea, as revealed through sequencing (Rumpho et al., 2011). However, Wagele et al., (2011) argues that HGT is not responsible for the acquisition of psb0 by E. chorotica which is needed to sustain photosynthesis, and instead suggests that this gene is actually already possessed by the plastids and are only aided by the intracellular environment of E. chorotica as it refrains from degrading plastids.

HGT has had a minor impact on multicellular organisms, with some estimates of 2% of an entire genome arising from HGT (McInerney et al., 2007). However, the impact on single celled organisms is far greater and HGT has the potential to erase much of the history recorded in prokaryotes prior to the evolution of eukaryotes (Simonson et al., 2005). The importance of this is that it poses a difficulty in interpreting the true evolutionary history of prokaryotes from molecular sequence data. This means that the original tree-like root on the ToL has become compromised and it is now difficult to determine if the ToL is a tree like structure at the root or if it is now net or web-like as a consequence of HGT. A net or weblike tree illustrates that the rate at which HGT has occurred is too great to determine which is its true ancestor and which is the original source of the transferred DNA (Williams et al., 2011). However, if many molecules are sequenced and good computational analysis is applied, it may still be possible to retrieve the true tree of cellular division (Puigbo et al., 2009, Williams et al., 2011).

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The evolution of eukaryotes (Fig.3) occurred through a phenomenon known as genome fusion. This refers to a process whereby an organism incorporates its genome into another organism’s genome (Cotton & McInerney, 2010).

Eukaryotes arose as a result of genome fusion, between

two

diverse

prokaryotic

genomes,

Figure 3. The three domains of life, eubacteria and archaebacteria giving rise to the eukaryota. Taken from (Understanding Evolution, 2011)

eubacteria and archaebacteria, providing each other with biochemical services (Pisani et al., 2007), resulting in a more complex cell of which the potential is evident by the diversity of complex life on earth (Rivera & Lake, 2004).

However,

the

eubacteria

and

archaebacteria are both distinct separate domains on the base of the ToL and this genome fusion means that both domains merged into one, and this means that the tree Figure 4. A schematic diagram of the ring of life. Taken from (Rivera & Lake, 2004)

of life now resembles a ring of life (Fig.4) (Rivera & Lake, 2004).

DNA Barcoding A molecular-based tool for identifying and categorising specimens has emerged over the last decade (Hebert et al., 2003), and this may also aid in reconstructing the universal ToL (Casiraghi et al., 2010). This simple concept known as DNA barcoding is achieved by applying “barcodes� to specimens by marking a particular region on a DNA sequence called the cytochrome c oxidase 1 (CO1) gene (Kress et al., 2005). CO1 is a 648-bp region on a

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mitochondrial gene and it is a more suitable marker than nuclear DNA. This is, because evolution occurs faster in mitochondrial DNA (mtDNA), resulting in a larger accumulation of changes between species that are closely related (Hebert et al., 2004b). Although several studies using DNA barcoding have been successful (Hebert et al., 2003, Hebert et al., 2004a, Hebert et al., 2004b), DNA barcoding has been highly controversial (Meyer & Paulay, 2005).

Criticisms have been made, for example, in using single genes as universal markers, and this is particularly true for CO1, as while it is successful in animals, it is not appropriate in barcoding plants (Kress et al., 2005), because the gene evolves too slowly in plants and fungi for it to discriminate between species levels (Pires & Marinoni, 2010), in which case other genes are being explored (Kress et al., 2005).

In recent years, DNA barcoding has become more accepted by the scientific community (Moritz & Cicero, 2004), bringing us closer to inferring the universal ToL. Although taxonomists were largely sceptical and had reservations in regarding to DNA barcoding methods, fearing that it might replace traditional taxonomic methods, it has now become apparent that both methods are crucial and complement each other (Moritz & Cicero, 2004). However, DNA barcoding is still in its infancy and many issues need to be addressed.

For instance, Casiraghi et al., (2010) cautions that evolution is continuous and species are not frozen in time. Therefore, we need to consider that once a given species is assigned a barcode, how that will read as that species evolves in the future. Databases have been established which confirm the growing support for DNA barcoding. For example, The consortium for the barcode of life (CBOL) (http://barcoding.si.edu/) and The Barcode of Life Data Systems (BOLD) (http://www.boldsystems.org/views/login.php) are online resource databases that provide a global bank containing DNA barcode records (Edwards, 2005, Ratnasingham & Hebert, 2007). As data accumulates, detailed phylogenies will become evident and DNA barcoding will become a formidable tool for inferring the universal ToL. 8

Conclusion Although binomial nomenclature does little to explain the evolutionary history of organisms, the traditional methods of identifying shared morphological characteristics have been beneficial to constructing phylogenetic trees and inferring the ToL. Molecular sequence data in the 1960s revolutionised phylogenetics and has enabled us to resolve many relationships between species. However, DNA sequencing often conflicts with traditional taxonomy and has revealed the damaging effect that HGT has had on interpreting the evolutionary history that is preserved in DNA. It first seemed that the legitimate root of the ToL was not retrievable but we now know that large scale molecular and morphological work can resolve deep branches and with the advancement of DNA barcoding, long term data compiling is likely to open up new avenues in molecular sequencing and resolve detailed phylogenetic trees. As it stands, the ToL resembles a ring of life as a result of endosymbiosis between eubacteria and archaebacteria giving rise to the eukaryote.

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