Äîêóìåíò âçÿò èç êýøà ïîèñêîâîé ìàøèíû. Àäðåñ îðèãèíàëüíîãî äîêóìåíòà : http://www.mso.anu.edu.au/~charley/papers/ChopraLineweaverASSC09v3.pdf Äàòà èçìåíåíèÿ: Thu May 13 10:03:35 2010 Äàòà èíäåêñèðîâàíèÿ: Tue Oct 2 06:13:50 2012 Êîäèðîâêà: Ïîèñêîâûå ñëîâà: m 17

Palaeoecophylostoichiometrics
Searching for the Elemental Composition of the Last Universal Common Ancestor
Aditya Chopra , Charles H. Lineweaver , Jochen J. Brocks and Trevor R. Ireland Planetary Science Institute Research School of Earth Sciences, Research School of Astronomy and Astrophysics The Australian National University, Canberra, Australia Email: adi@mso.anu.edu.au

Summary: Extant life on Earth shares a common ancestor that probably existed 4 billion years ago. We describe a novel approach to study the root of the phylogenetic tree of life by measuring the elemental abundances of living organisms and their environment. We discuss the conservation of possible to use stoichiometric and changes in the distribution of elem the evolution of life over the past event 2.4 billion years ago. different elemental ratios across different taxa. It may be phylogenetic relationships between taxa to investigate how ents in environments such as the oceans, may have affected 4 billion years ­ particularly during the great oxidation

We outline some strategies to identify modern analogues of geological sites where the common ancestor may have existed and where life first evolved from its pre-biotic origins on Earth and possibly elsewhere in the universe.

Introduction
Darwin's book [1] contained a single figure with hypothetical branches of descent for organisms. Today, Darwin's branches have grown into the universal tree of life, where all extant and many extinct organisms, have found a place [4]­[6]. At the root of the tree, is the Last Universal Common Ancestor (LUCA) (Figure 1) [7], [8]. Biogeochemists are beginning to connect the major patterns of evolutionary diversification seen in the phylogenetic tree with environmental events in Earth's history [9], [10]. The rise in O2 2.4 billion years ago [11], was coupled to changes in the biogeochemical cycles of elements like iron, which was oxidised from soluble Fe2+ to form insoluble Fe3+ precipitates, reducing the amount of free soluble iron available to life in the oceans. Changes in oxygen levels and metal concentrations in the oceans between 2.4 - 1.8 billion years ago would have led to changes in the distribution of metal ions inside cells (the metallome). Metals play a role in almost all metabolic pathways (providing structure and catalytic activity to enzymes) and life would have adapted to the changing conditions leading to new metabolic pathways [12]­[16].


544 Ma
M icrospor idia Ciliates

Animals Plants Fu n g i Fl a g e l l a t e s

E u ka r ya

Slime moulds

1000 Ma
Diplomonads

Bac teria
Pu r p l e b a c te r i a

Green non-sulfur bac teria

1500 Ma
S u l fo l o b u s

Gram positive bac teria

1870 Ma

Archaea

Thermotoga

2450 Ma

P yrodic tium

N itrospira

Ther moproteus P yrobaculum

Cy a n o b a c t e r i a Green sulfur bac teria

P yrococcus M ethanobac ter ium Thermoplasma Archaeoglobus Halobac terium Methanococcus

3500 Ma
Aq u i fex

3500 Ma ?
M ethanosarcina

Last Universal Common A ncestor

Fig. compari selected biogenic

1: Phylogenetic relationships of extant organisms, as inferred from sequence sons of 16S ribosomal RNA genes. The boxed dates indicate the minimum age of branches, based on deductions from molecular biomarkers, isotopic signatures of materials, body fossils and molecular clocks. Adapted from [9] and based on [8].

In comparison to techniques available just a few decades ago, it is possible to measure the elemental composition (the stoichiometry) of life forms and their environments much more easily and to much higher precision. Most of the stable elements in nature can be measured to approximately parts per billion concentrations, using less than a few grams of material. Our approach, termed Palaeoecophylostoichiometrics1, is based on quantifying the stoichiometry of extant bacteria and archaea, known from phylogenetic studies to be most closely related to LUCA. It could reveal new insights into the links between changing paleo-ecologies and early evolution of life and may help to identify the geochemistry of the site for the origin of life.

Stoichiometry of life
Although life is made of elements from its environment and exhibits similar elemental abundances to that of its environment, it is at disequilibrium with the environment. Some elements like carbon, nitrogen and phosphorus are more abundant in life than in the environment, while elements like sodium and potassium are less abundant in life compared to the environment. In [17], we present the view that life may use the disequilibrium in the elemental abundances relative to the environment for its metabolic reactions. Every organism will have an elemental composition that is dependent on its metabolic activity. If the organism's oxygen carrying protein in the blood is hemoglobin that uses iron (as in humans), instead of hemocyanin that uses copper (as in horseshoe crabs [18]), the organism will probably have a higher abundance of iron than an organism using hemocyanin.
Palaeoecophylostoichiometrics is a modification of `Paleoecostoichiophylogenomics' introduced by Elser in [21] to describe an emerging sub-discipline that links biogeochemical and genomic studies.
1


The elemental abundances in an organism will also be affected to some extent by the abundances in the environment. Individuals living in an environment with high concentrations of mercury will probably exhibit increased abundances of mercury than individuals living in environments with low mercury concentrations. Due to slight non-specificity in the mechanism of selectivity between metal ions, elements like lead and strontium can be accumulated in the body as a result of calcium intake, even though they may not be utilised by life. Such environmental effects will introduce noise to any signal that is due to the metabolism of the organism. However, we hypothesise that despite such environment-dependent variations, if the environmental factors are quantified and controlled, there will be signals in the metallome that can be used to distinguish between the physiology and evolutionary history of different taxa. These signals could be based on the relative abundances of elements within the organism and differences between these abundances compared to the abundances found in the environment (such as the growth medium). Redfield found in the Western Atlantic that stoichiometric ratios of both phytoplankton and the surrounding water column were in ratios of C106 :N16 :P1 , by atoms [19]. The similarity in the ratios of phytoplankton and the water column indicated a balanced flow of C, N and P in and out of the environment.
1000
C :P N :P C :N

Canonical Red eld ratios

100

Ratio
10

1

Phytoplankton Insects

Bacteria

Human

Fig. 2: The C:N:P ratios are an example of a stoichiometric signal. Variations from the canonical Redf eld ratios [19] for phytoplankton ref ect differences in the metabolic pathways, environments and evolutionary histories of each taxa. Ratios given are by number of atoms and the uncertainties correspond to standard deviations. "Phytoplankton" represents marine phytoplankton from surface waters at coastal and off-shore sampling sites around North America [20]. Insects, represent whole-body measurements of different species of land insects [21]. Bacteria abundances are representative of 7-40 species depending on the element [22]. Human abundances are from a study involving about 150 individuals [23]. The uncertainty in measurements for bacteria and humans have not been quantif ed by the authors. We have used data for phytoplankton and insects from [20] and [21] as they were two of the most comprehensive data sets which have uncertainties associated with them. Elemental abundances for other taxa, especially in Bacteria and Archaea have not been well quantif ed.


Results in [24] suggest that bulk elemental compositions reflect phylogenetic differences in two phytoplankton plastid superfamilies, those that use chlorophyll-b and those that use chlorophyll-c as the major photosynthetic pigment [25]. While both plastid families share common metabolic functions, the C:N:P stoichiometries of different phytoplankton reflect different ancestral pre-symbiotic host cell phenotypes, while the abundances of trace elements like iron and copper probably reflect differences in the evolution of the acquired plastids in different ocean redox states since the Proterozoic era. We hypothesise that some stoichiometric signals will be conserved between different taxa. Stoichiometric signals are likely to be more similar within low taxonomic orders such as species and genera and less similar between higher taxonomic orders such as phyla, kingdom and domain. In order for a stoichiometric signal to be useful, the level of conservation of the stoichiometric signal must be such that it is sufficiently distinct for particular taxa. Indeed, if the signal remained exactly the same in every taxa, it would not be possible to distinguish between two taxa. In contrast, if the abundances were randomly distributed then it would be impossible to relate different taxa by stoichiometric signals.

Looking back in time
There are a number of features in life on Earth that have been recognised as being conserved over timescales of millions to billions of years. One widely used feature of life on Earth that is used to represent evolutionary descent, in the form of a phylogenetic tree (Figure 1), is the 16S ribosomal RNA gene (16S rRNA). The 16S rRNA's highly conserved sequence is attributed to its essential signalling and structural role in protein synthesis from a genetic code. Phylogenetic trees that are based on other conserved molecules such as transfer RNAs and proteins, are topologically similar to 16S rRNA trees [26]­[30]. Genetic and proteomic conservation translates to conservation of metabolic pathways [31]. Therefore, the stoichiometry of elements that make up proteins in metabolic pathways will be conserved to varying degrees across taxa. We expect that stoichiometric relationships between taxa will, to some extent, reflect phylogenetic relationships established by genomic and proteomic studies [32]­[37]. Figure 3 shows that the relative proportions of the chemical elements in organisms like humans and bacteria, that represent two different domains of life (Eukarya and Bacteria), are similar [17], [38], [39]. In some cases, elemental composition may be more conserved than biomarkers such as lipids or genetic markers such as the 16S rRNA sequence. For example, until recently viruses were not considered part of most phylogenetic trees as they do not contain 16S rRNA [6], [27], [40]­[43]. However, their DNA or RNA based genes, proteins and lipids are made of the same chemical elements that are used by other life forms. Rooted phylogenetic trees can have their branch lengths (distance between two nodes) calibrated for inferring the extent of divergence of species from their common ancestor. The shorter the branch length of an extant species from the root of the tree, represented by LUCA, the more it is likely to resemble LUCA. By studying elemental abundances in bacteria and archaea that have short branch lengths from LUCA, it may be possible to infer the elemental compositon of LUCA and its environment [44], [45].


C

H O

104
N K (number of atoms - normalised to Fe) P

Abundance in Bacteria

10

2

Mg

Na S Cl

Ca

Mn

100
Zr Co Mo Cr

Al Cu B

Fe Zn

Si

10-2

As

Cd

10-4 10-4

Se

10-2

100

102

104

Abundance in Humans
(number of atoms - normalised to Fe)

Fig. 3: A positive correlation between elemental abundances (by number of atoms) in the representatives of two domains of life, Bacteria (bacteria [22]) and Eukarya (humans [23]). Uncertainties were not quantif ed by the authors. The abundances are normalised to iron.

It is possible to use branch lengths as a proxy for divergence times if the rate of evolutionary change is known [46]. The rate is often measured by the number of substitutions in a conserved genetic sequence like 16S rRNA but amino acid sequences for proteins can also be used. However, this is not always possible as molecular clocks are not constant across taxa and over long timescales [47], [48]. The issue can be resolved to some extent by calibration against fossil and biogeochemical records [9], [49]­[51]. By studying bacteria and archaea near the root of phylogenetic trees, it may be possible to relate evolutionary changes in the metallome of life to changes in the environment. Analyses of the elemental composition of taxa with respect to branching order from the root of the phylogenetic tree can be used to determine if there is any identifiable trend of life using a larger suite of elements as a function of time. Those species that are deeply rooted in the tree may utilise fewer elements in their metabolic pathways compared to species that have branched out further from the root. Figure 4 shows a suggestive model that represents the total number of elements in use across all life forms on Earth, as a function of time. Life may have begun with a basic set of elements and then progressively increased its elemental `toolkit'. Initially, life may have been based on abundant elements like carbon, hydrogen and oxygen and ions like calcium or boron to stabilise its structures [53], and catalytic metals like iron [54], [55]. This increase may be in response to changes in the bioavailability of elements. Alternatively, just as a novice chef eggs and sugar, life may have started carbon, hydrogen, oxygen, nitrogen, s with more elements like bromine and use vanilla, cinnamon and the rest of starts baking a cake with simple ingredients like flour, with the most abundant (or bioavailable) elements like ulphur and phosphorus. Later, life began to accessorise selenium ­ like an experienced chef who has learnt to the spice rack.


Estimate of the num ber of elem ents in use by life

30

20

10

Sequence of addition of elements into inventory of life

Cd Sn Se As Sr Ba I Br F Al Si Zn Cu Mo V Cr Mn Co W Ni Fe Na K Mg B Ca Cl P S N O H C

4

3.8

2

1

0

Billion years ago

Fig. 4: A possible sequence of increasing diversity of elements used by life based on estimates of abundances in oceans and discussion in [36]. The order of the elements on the right-hand-side y-axis is speculative but plausible.

Mediated by processes such as endo-symbiosis [56] and horizontal gene-transfer [57], recipes would have been exchanged between different life forms at different times over the past 4 billion years, resulting in usage of new elements and new metabolic pathways. Not shown in Figure 4, there may have been instances of a transient decrease in the number of elements in use by life, due to factors such as extinction of particular taxa or large decrease in the bioavailability of an element, even if for short timescales. Whether our research or others will be able to resolve such transient decrease in the number of elements used by life remains to be seen.

Tracing the evolution of life
Williams and da Silva [58] have suggested that the major changes from the first life form(s) to present life forms were linked to the changing chemistry of the external environment. For example, the development of enclosing membranes provided a stable physical and chemical internal environment for sustaining metabolic reactions of the earliest prokaryotic life form(s). Most origin of life hypotheses require a redox coupled energy source as the earliest metabolisms. An abundant source of a redox potential accessible to life on the Hadean and early Archean Earth, 4.5 to 3.5 billion years ago, are crustal minerals or the redox gradients at hydrothermal vents [52]. These sites most likely had a host of organic molecules such as amino-acids and nucleotides that formed in situ or were brought in by chondritic and cometary material and could have formed auto-catalytic polypeptides and nucleaic acids that developed into the information and metabolic system of the first of the single-celled organisms [55]. On depletion of the initially easily available free energy sources, other metabolisms evolved and some life forms developed the ability to perform oxygenic photosynthesis [59].


Oxygenation of the atmosphere 2.4 billion years ago [11] by efficient oxygenic photosynthetic organisms, led to the oxidation of metals in the environment. The resulting oxidised metals would be incompatible with a reducing cytoplasm, that would have reflected the reducing chemistry of the primitive oceans and atmosphere from which it formed. Isolation of the incompatible pathways within internal compartments through the gradual introduction of vesicles in cells from localised invagination or endosymbiotic events may have resulted in eukaryotic life forms with organelles and a nucleus [59]­[61]. Changes in the environmental abundances would affect the elemental composition of and probably the usage of elements in different taxa that over time would lead to the evolution of new metabolic pathways. If the concentration of a metal used by life decreases in the environment such that an alternative metal is more competitive in terms of bioavailability, reaction rate or reaction efficiency, then life will probably co-opt the alternative metal in a modified metabolic pathway. For example, the rise in atmospheric oxygen concentration in the period 2.4 to 0.8 billion years ago has been linked to changes in the chemical components of the oceans (see Figure 5). This period also corresponds to an increase in number of enzymes that use metals such as copper and molybdenum [32], [36]. As soluble Fe2+ was oxidised to Fe3+ (which forms insoluble precipitates of Fe(OH)3 ), the availability of free or complexed ions of biologically active iron would have decreased substantially in the oceans in the period 2.4 to 0.8 billion years ago. The oxygenation also oxidised another biologically active metal, copper from its Cu+ to Cu2+ state.
0 O
2

­2 ­4 ­6

ATMOSPHERE
O
2

OCEAN
S Fe

S

­2 ­3 ­4

­6 Mo Co Mn Ni Mo
Zn

­7 ­8 ­9 ­10

Ni

Cu

Fe

Mn Co

Zn Cu

­16 ­19 ­22 ­25

4

3

2

1

0

Billion years ago

Fig. 5: The increase in O2 concentration of the atmosphere (top) affected the concentration of metals and other ions in seawater (bottom). Modif ed from Anbar et al. [33] and based on geochemical models and inferences from ancient sediments. Dashed lines are for elements with falling concentrations. Color gradations indicate a transition from anoxic oceans before 2.4 billion years ago (green) to H2 S-rich oceans between 1.8 billion and 800 million years ago (purple), subsequently giving way to complete ocean oxygenation (light blue). [M: moles per litre; atm: atmosphere]

log [Concentration (M)]

­5

log [Pressure (atm)]


This change in the environment may have triggered the development of new metabolic pathways which harnessed the electrochemical potential of the higher oxidation state of copper for roles previously performed by iron [58]. As the availability of metals such as manganese, zinc and cobalt increased in the environment, new metabolic pathways using these metals arose in both prokaryotes and eukaryotes [35]­[37], [61], [64]. An example is the metalloenzyme superoxide dismutase, which is a detoxifying agent to remove the O2- anion, a by-product of metabolic reactions involving one electron reduction of O2 . Most eukaryotes use a form of the enzyme with copper and zinc metal centres at the active site. However, most prokaryotic organisms and mitochondria contained in eukaryotes2 contain a form with manganese and or iron. The concentration of iron and manganese decreased after the oxygenation 2.4 billion years ago and it may explain the emergence of copper- and zinc- based superoxide dismutases in eukaryotes [63]. Since, the elemental abundances in an organism reflects to some extent the elemental abundances in its environment [17], it may be possible to suggest geological sites where LUCA could have existed and help identify modern analogues of geological sites where the origins of life on Earth could have taken place (Figure 6). For example, if LUCA is inferred to have contained and utilised boron, it could add support to the borate-minerals-stabilisingribose model [53], necessary for efficient synthesis of RNA in the transition from prebiotic to biotic life.

100

O

C H

10

N Ca P

1

Abudance (% by m as s )

S

Na

K

Cl Mg Fe

10

-1

10

-2

10

-3

10

-4

Ni

10

-5

Inferred LUCA Hydrothermal vent Radioactive beach

Co

Mo

O

C

H

N

S

P

Na

K

Cl

Mg

Ca

Fe

Co

Ni

Mo

Element

Fig. 6: An environment for LUCA and the origin of life, may be determined by comparing abundances in LUCA and two candidates sites. Abundances and uncertainties are hypothetical. The abundances for a hydrothermal vent show stronger correlation to abundances in LUCA than the correlation between the radioactive beach and LUCA. The result would suggest that hydrothermal vents are more likely than radioactive beaches to represent the geochemistry of the site for the origin of life [54], [55], [69].
2 Mitochondria contained in eukaryotes are believed to have been derived from endosymbiosis between a eukaryote and a prokaryote (-proteobacterium) approximately 1.7 to 2 billion years ago [56], [62].


The Moon-forming life at that time [67]. any inferences about years after the Moon

impact 4.5 billion years ago would have sterilised Earth of any extant LUCA probably existed 4.2 ± 0.4 billion years ago [68]. Therefore, the environment of LUCA will at best relate to a few hundred million forming impact.

We are not attempting to examine past extinct life but rather inferring the evolutionary history of extant taxa. Metabolic pathways in extant taxa are often conserved in the sense that they reflect the pathways in use by the taxa when the taxa first appeared on Earth. However, particular lineages may have gone extinct or metabolic pathways may not be sufficiently conserved within taxa due to responses to environmental changes over billions of years. In future analysis, we will establish the extent to which extant basal organisms preserve a record of the metabolisms in use up to 4 billion years ago. Although the geochemistry of past environments over the past 4 billion years is not well established, there are some constraints on elemental abundances in the oceans, especially in the proterozoic era (2500 - 550 million years ago) based on the geological record and biogeochemical evidence [33], [34], [36]. It is also possible to correlate metal availability in the environment to the evolution of metabolic pathways as in [32]. We have not established the extent to which environmental abundances affect the elemental composition of and usage of elements in different taxa. In future analyses we will investigate if there are signals that are large enough to be detected over the noise introduced by fluctuations in the abundances in the environment.

Sun Earth Crust Life
All atoms that are or have been part of living matter on Earth have either been produced during big bang nucleosynthesis [72] or in different processes of the stellar nucleosynthetic pathways that take place in stars [73]. Our Sun was formed out of a collapsing molecular cloud that was polluted by earlier stellar processes and the proto-planetary disk that was made of the remaining dust eventually gave rise to terrestrial planets like the Earth. The composition of solar system objects including other planets and asteroids reflects to a large extent the composition of the Sun, except that relative to the Sun, all are depleted in the most volatile elements hydrogen, helium and the noble gases. When the Sun formed from the solar nebula, 4.5672 (± 0.0006) billion years ago [74], the solar wind caused the depletion of volatile elements such as hydrogen and helium from the region of the solar nebula where terrestrial planets formed. Rocky planets like Earth, accreted from the fractionated nebular condensate whose composition (particularly C1 chondrites) closely resembles the solar composition, except for the abundances of volatile elements [75]. Input from chondritic material and a late-veneer of volatile comets and other objects from beyond the snow-line led to a abundances more like solar abundances, depleted in volatile of the bulk Earth. Life on Earth is based on elements such as nitrogen which were added to the crust in form of water, carb the chondritic and cometary material. elements due to the impacts of crust which exhibits elemental elements, than the abundances hydrogen, oxygen, carbon and on dioxide and ammonia ice in


Today, life on Earth is primarily composed of the chemical elements oxygen, carbon, hydrogen and nitrogen, which in humans and bacteria make up 96.8 ± 0.1% of the mass. Apart from these bulk elements, phosphorus and sulphur together make up 1.0 ± 0.3% of the mass. The remaining 2.2 ± 0.2% of the mass is dominated by potassium, sodium, calcium, magnesium and chlorine, while 0.03 ± 0.01% is attributed to trace elements such as iron, copper and zinc. Life on Earth is based on the most abundant elements in its environment. Life does not reside in the mantle or the core of the Earth and so its elemental abundances are more reflective of abundances in the upper crust (specifically the biosphere) than abundances in the bulk Earth. The process of formation of a star like the Sun, out of a collapsing molecular cloud polluted by heavy elements from earlier generation of stars, is observed wherever there are molecular clouds. The associated process of terrestrial planet formation is probably common in the universe and it is likely that the elemental abundances of the surfaces of extrasolar habitable planets will also be similar to cosmic abundances as represented by the Sun. Since, the abundance of most elements in life forms and their environments on Earth follow cosmic abundances (Figure 7), perhaps extraterrestrial life will also exhibit elemental abundances similar to those found in life on Earth [38]. Alternatively, if extraterrestrial life is to be found on planets and moons with environments where elemental abundances are different to those found on Earth, it may be possible to predict the elemental abundances and metabolic processes of the extraterrestrial life based on the stoichiometric principles of life on Earth.
105
H O C

N

Abundance in Humans (number of atoms - Normalised to Si)

P K

Ca Na S Mg Si

10

0
F

Cl

Fe Zn Rb Br Sr B

Al Ti

10-5
Ta U Th

Cu Li Cd Pb Ce Se Sn Ba Ge I Mo Cs As Hg Ag Sb Nb Y Zr Ga La Sc In Te V Bi Tl Be Au Sm W

Cr Mn

Ni

Co

10-10 10-10

10

-5

100

105

Abundance in Sun (number of atoms - Normalised to Si)

Fig. 7: A positive correlation between elemental abundances (by number of atoms) in life, as represented by humans [23], and the Sun [75]. The abundances are normalised to silicon.


Our approach to studying the origins and evolution of life by measuring the elemental abundances of extant taxa and their environment will requires a sufficiently large data set looking at a number of taxa with relatively small uncertainties associated with the measurements. A major shortcoming of existing data in the literature was that measurements were made many decades ago, before the availability of high precision analyses offered by analytical techniques today. It is possible that the errors associated with previous measurements of abundances of some elements could be in the range of a few orders of magnitude. Results from authors such as Sedykh et al. [70], who used Inductively Coupled Plasma Atomic Emission Spectroscopy (ICP-AES) techniques and Heldal et al. [71], who used Xray microanalysis to measure elemental abundances of about 10 elements in cyanobacterial species, suggest that once culturing and analytical methods have been standardised, it will be possible to quantify elemental abundances to a precision of 5-10%, in a high-throughput method. If the effect of environmental variations on elemental composition of taxa is sufficiently small then we may be able identify elemental ratios that are conversed across different taxa and correlate changes in the bioavailability of elements in environments with evolution of metabolic pathways. Palaeoecophylostoichiometrics could help us understand LUCA and its environment and the principles that we learn about life on Earth could be applicable elsewhere in the universe.

References
[1] C. Darwin, On the Origin of Species by Means of Natural Selection, or the Preservation of Favoured Races in the Struggle for Life. John Murray, 1859. [2] J. Pereto, J. Bada, and A. Lazcano, "Charles Darwin and the Origin of Life," Origins of Life and Evolution of ´ Biospheres, vol. 39, no. 5, pp. 395­406, 2009. [3] W. B. Whitman, D. C. Coleman, and W. J. Wiebe, "Prokaryotes: The unseen majority," Proceedings of the National Academy of Sciences of the United States of America, vol. 95, no. 12, pp. 6578­6583, 1998. [4] P. Puigbo, Y. Wolf, and E. Koonin, "Search for a 'Tree of Life' in the thicket of the phylogenetic forest," Journal of Biology, vol. 8, no. 6, pp. 59, 2009. [5] K. Swithers, J. P. Gogarten, and G. Fournier, "Trees in the Web of Life," Journal of Biology, vol. 8, no. 6, pp. 54, 2009. [6] H. Brussow, "The not so universal tree of life or the place of viruses in the living world," Philosophical Transactions ¨ of the Royal Society B: Biological Sciences, vol. 364, no. 1527, pp. 2263­2274, 2009. [7] C. Woese, "The universal ancestor," Proceedings of the National Academy of Sciences of the United States of America, vol. 95, no. 12, pp. 6854­6859, 1998. [8] N. R. Pace, "A Molecular View of Microbial Diversity and the Biosphere," Science, vol. 276, no. 5313, pp. 734­740, 1997. [9] J. J. Brocks and J. Banfield, "Unravelling ancient microbial history with community proteogenomics and lipid geochemistry," Nature Reviews Microbiology, vol. 7, no. 8, pp. 601­609, 2009. 10] A. D. Anbar and A. H. Knoll, "Proterozoic Ocean Chemistry and Evolution: A Bioinorganic Bridge?" Science, vol. 297, no. 5584, pp. 1137­1142, 2002. 11] A. Bekker, H. D. Holland, P.-L. Wang, D. Rumble, H. J. Stein, J. L. Hannah, L. L. Coetzee, and N. J. Beukes, "Dating the rise of atmospheric oxygen." Nature, vol. 427, no. 6970, pp. 117­120, 2004. 12] R. J. P. Williams, "Chemical selection of elements by cells," Coordination Chemistry Reviews, vol. 216­217, pp. 583­595, 2001. 13] F. M. M. Morel and N. M. Price, "The Biogeochemical Cycles of Trace Metals in the Oceans," Science, vol. 300, no. 5621, pp. 944­947, 2003. 14] F. M. M. Morel, "The co-evolution of phytoplankton and trace element cycles in the oceans," Geobiology, vol. 6, no. 3, pp. 318­324, 2008. 15] P. G. Falkowski, T. Fenchel, and E. F. Delong, "The Microbial Engines That Drive Earth's Biogeochemical Cycles," Science, vol. 320, no. 5879, pp. 1034­1039, 2008.

[ [ [ [ [ [


[16] P. G. Falkowski and L. V. Godfrey, "Electrons, life and the evolution of Earth's oxygen cycle," Philosophical Transactions of the Royal Society B: Biological Sciences, vol. 363, no. 1504, pp. 2705­2716, 2008. [17] A. Chopra and C. H. Lineweaver, "The major elemental abundance differences between life, the oceans and the Sun," in Proceedings of the 8th Australian Space Science Conference, Canberra, W. Short and I. Cairns, Eds., vol. Australian Space Science Conference Series. National Space Society of Australia Ltd, 2009, pp. 49­55. [18] M. Brenowitz, C. Bonaventura, J. . Bonaventura, and E. Gianazza, "Subunit composition of high molecular weight oligomer: Limulus polyphemus hemocyanin." Archives of biochemistry and biophysics, vol. 210, no. 2, pp. 748­61, 1981. [19] A. Redfield, "The biological control of chemical factors in the environment." American Scientist, vol. 64, pp. 205­221, 1958. [20] T.-Y. Ho, A. Quigg, Z. V. Finkel, A. J. Milligan, K. Wyman, P. G. Falkowski, and F. M. M. Morel, "The elemental composition of some marine phytoplankton," Journal of Phycology, vol. 39, no. 6, pp. 1145­1159, 2003. [21] J. J. Elser, "Biological stoichiometry: a theoretical framework connecting ecosystem ecology, evolution, and biochemistry for application in astrobiology," International Journal of Astrobiology, vol. 2, pp. 185­193, 2003. [22] J. R. Porter, Bacterial Chemistry and Physiology. London: John Wiley and Sons, 1946. [23] W. S. Snyder, M. J. Cook, I. H. Tipton, E. S. Nasset, L. R. Karhausen, and G. P. Howells, Reference Man: Anatomical, Physiological and Metabolic Characteristics, 1st ed., ser. Report of Task Group on Reference Man - International Commission on Radiological Protection. New York: Pergamon Press Ltd., 1975, vol. 23. [24] A. Quigg, Z. V. Finkel, A. J. Irwin, Y. Rosenthal, T.-Y. Ho, J. R. Reinfelder, O. Schofield, F. M. M. Morel, and P. G. Falkowski, "The evolutionary inheritance of elemental stoichiometry in marine phytoplankton," Nature, vol. 425, no. 6955, pp. 291­294, 2003. [25] P. G. Falkowski, M. E. Katz, A. H. Knoll, A. Quigg, J. A. Raven, O. Schofield, and F. J. R. Taylor, "The Evolution of Modern Eukaryotic Phytoplankton," Science, vol. 305, no. 5682, pp. 354­360, 2004. [26] H. Xue, L. T. Ka C. Marck, H. Grosjean, and F. W. J. Tze "Transfer RNA paralogs: evidence for genetic code-amino acid biosynthesis coevolution and an archaeal root of life," Gene, vol. 310, pp. 59­66, 2003. [27] F.-J. Sun and G. Caetano-Anolles, "Evolutionary patterns in the sequence and structure of transfer RNA: early origins ´ of archaea and viruses." PLoS Computational Biology, vol. 4, no. 3, p. e1000018, 2008. [28] S. Gribaldo and C. Brochier-Armanet, "The origin and evolution of Archaea: a state of the art," Philosophical Transactions of the Royal Society B: Biological Sciences, vol. 361, no. 1470, pp. 1007­1022, 2006. [29] C. E. Blank, "Phylogenomic Dating - A Method of Constraining the Age of Microbial Taxa That Lack a Conventional Fossil Record," Astrobiology, vol. 9, pp. 173­191, 2009. [30] J. T.-F. Wong, J. Chen, W.-K. Mat, S.-K. Ng, and H. Xue, "Polyphasic evidence delineating the root of life and roots of biological domains," Gene, vol. 403, no. 1-2, pp. 39 ­ 52, 2007. [31] C. Chothia, J. Gough, C. Vogel, and S. A. Teichmann, "Evolution of the Protein Repertoire," Science, vol. 300, no. 5626, pp. 1701­1703, 2003. [32] J. B. Glass, F. Wolfe-Simon, and A. D. Anbar, "Coevolution of metal availability and nitrogen assimilation in cyanobacteria and algae," Geobiology, vol. 7, no. 2, pp. 100­123, 2009. [33] A. D. Anbar, "OCEANS: Elements and Evolution," Science, vol. 322, no. 5907, pp. 1481­1483, 2008. [34] K. O. Konhauser, E. Pecoits, S. V. Lalonde, D. Papineau, E. G. Nisbet, M. E. Barley, N. T. Arndt, K. Zahnle, and B. S. Kamber, "Oceanic nickel depletion and a methanogen famine before the Great Oxidation Event," Nature, vol. 458, no. 7239, pp. 750­753, 2009. [35] C. L. Dupont, S. Yang, B. Palenik, and P. E. Bourne, "Modern proteomes contain putative imprints of ancient shifts in trace metal geochemistry," Proceedings of the National Academy of Sciences, vol. 103, no. 47, pp. 17 822­17 827, 2006. [36] A. L. Zerkle, C. H. House, and S. L. Brantley, "Biogeochemical signatures through time as inferred from whole microbial genomes," American Journal of Science, vol. 305, no. 6-8, pp. 467­502, 2005. [37] M. A. Fedonkin, "Ancient biosphere: The origin, trends and events," Russian Journal of Earth Sciences, vol. 10, pp. 1­9, 2008. [38] R. E. Davies and R. H. Koch, "All the Observed Universe has Contributed to Life," Philosophical Transactions: Biological Sciences, vol. 334, no. 1271, pp. 391­403, 1991. [39] H. J. M. Bowen, Environmental Chemistry of the Elements. London: Academic Press Inc., 1979. [40] P. Forterre and D. Prangishvili, "The origin of viruses," Research in Microbiology, vol. 160, no. 7, pp. 466­472, 2009. [41] D. Mindell, J. S. Rest, and L. P. Villarreal, Viruses and the Tree of Life, ser. Assembling the Tree of Life. Oxford University Press, New York: Oxford University Press, 2004, ch. 8, pp. 107­118. [42] M. C. Smith, R. N. Burns, S. E. Wilson, and M. A. Gregory, "The complete genome sequence of the Streptomyces temperate phage straight phiC31: evolutionary relationships to other viruses." Nucleic Acids Research, vol. 27, no. 10, pp. 2145­2155, 1999. [43] R. W. Hendrix, M. C. Smith, R. N. Burns, M. E. Ford, and G. F. Hatfull, "Evolutionary relationships among diverse bacteriophages and prophages: all the world's a phage." Proceedings of the National Academy of Sciences of the United States of America, vol. 96, no. 5, pp. 2192­2197, 1999. [44] N. Glansdorff, Y. Xu, and B. Labedan, "The Last Universal Common Ancestor: emergence, constitution and genetic legacy of an elusive forerunner," Biology Direct, vol. 3, no. 1, p. 29, 2008. [45] B. Boussau, S. Blanquart, A. Necsulea, N. Lartillot, and M. Gouy, "Parallel adaptations to high temperatures in the Archaean eon," Nature, vol. 456, no. 7224, pp. 942­945, 2008.


[46] S. B. Hedges and S. Kumar, The Timetree of Life. Oxford University Press, 2009, Discovering the timetree of life., p. 318. [47] L. Bromham and D. Penny, "The modern molecular clock," Nat Rev Genet, vol. 4, no. 3, pp. 216­224, 2003. [48] D. Graur and W. Martin, "Reading the entrails of chickens: molecular timescales of evolution and the illusion of precision," Trends in Genetics, vol. 20, no. 2, pp. 80­86, 2004. [49] C. Marshall, "A Simple Method for Bracketing Absolute Divergence Times on Molecular Phylogenies Using Multiple Fossil Calibration Points." The American Naturalist, vol. 171, no. 6, pp. 726­742, 2008. [50] F. U. Battistuzzi and S. B. Hedges, The Timetree of Life - Archaebacteria. Oxford University Press, 2009, pp. 101­105. [51] F. U. Battistuzzi and S. B. Hedges, The Timetree of Life - Eubacteria. Oxford University Press, 2009, pp. 106­115. [52] D. W. Deamer, The origin of celluar life ­ Planets and Life: The Emerging Science of Astrobiology. Cambridge University Press, 2007, pp. 187­209. [53] A. Ricardo, M. A. Carrigan, A. N. Olcott, and S. A. Benner, "Borate Minerals Stabilize Ribose," Science, vol. 303, no. 5655, p. 196, 2004. [54] G. Wachtershauser, "Origin of life: Life as We Don't Know It," Science, vol. 289, no. 5483, pp. 1307­1308, 2000. ¨ ¨ [55] W. Martin, J. Baross, D. Kelley, and M. J. Russell, "Hydrothermal vents and the origin of life," Nature Reviews Microbiology, vol. 6, no. 11, pp. 805­814, 2008. [56] L. Sagan, "On the origin of mitosing cells," Journal of Theoretical Biology, vol. 14, no. 3, pp. 225­274, 1967. [57] C. R. Woese, "Interpreting the universal phylogenetic tree," Proceedings of the National Academy of Sciences of the United States of America, vol. 97, no. 15, pp. 8392­8396, 2000. [58] R. J. P. Williams and J. J. R. Frausto da Silva, The Chemistry of Evolution. Oxford, UK: Elsevier, 2006. ´ [59] W. Martin and M. J. Russell, "On the origins of cells: a hypothesis for the evolutionary transitions from abiotic geochemistry to chemoautotrophic prokaryotes, and from prokaryotes to nucleated cells," Philosophical Transactions of the Royal Society B: Biological Sciences, vol. 358, no. 1429, pp. 59­85, 2003. [60] R. J. P. Williams and J. J. R. Frausto da Silva, "Evolution was Chemically Constrained," Journal of Theoretical ´ Biology, vol. 220, no. 3, pp. 323­343, 2003. [61] R. J. P. Williams, "The natural selection of the chemical elements," Cellular and Molecular Life Sciences (CMLS), vol. 53, no. 10, pp. 816­829, 1997. [62] V. Emelyanov, "Rickettsiaceae, Rickettsia - Like Endosymbionts, and the Origin of Mitochondria," Biosience Reports, vol. 21, no. 1, pp. 1­17, 2001. [63] F. Wolfe-Simon, D. Grzebyk, O. Schofield, and P. G. Falkowski, "The role and evolution of superoxide dismutases in algae," Journal of Phycology, vol. 41, no. 3, pp. 453­465, 2005. [64] M. K. Chan, S. Mukund, A. Kletzin, M. W. Adams, and D. C. Rees, "Structure of a hyperthermophilic tungstopterin enzyme, aldehyde ferredoxin oxidoreductase." Science, vol. 267, no. 5203, pp. 1463­1469, 1995. [65] R. J. P. Williams and J. J. R. Frausto da Silva, The natural selection of the chemical elements: the environment and ´ life's chemistry. New York: Oxford University Press Inc., 1996. [66] CRC, Handbook of chemistry and physics, 85th ed. Cleveland, Ohio: Chemical Rubber Company, 2004. [67] A. N. Halliday, "A young Moon-forming giant impact at 70-110 million years accompanied by late-stage mixing, core formation and degassing of the Earth," Philosophical Transactions of the Royal Society A: Mathematical, Physical and Engineering Sciences, vol. 366, no. 1883, pp. 4163­4181, 2008. [68] S. B. Hedges, The Timetree of Life - Life. Oxford University Press, 2009, pp. 89­98. [69] Z. Adam, "Actinides and Life's Origins," Astrobiology, vol. 7, no. 6, pp. 852­872, 2007. [70] E. M. Sedykh, O. A. Lyabusheva, A. K. Tambiev, and L. N. Bannykh, "Determination of the elemental composition of cyanobacteria cells and cell fractions by atomic emission and atomic absorption spectrometry," Journal of Analytical Chemistry, vol. 60, no. 1, pp. 29­33, 2005. [71] M. Heldal, D. J. Scanlan, S. Norland, F. Thingstad, and N. H. Mann, "Elemental Composition of Single Cells of Various Strains of Marine Prochlorococcus and Synechococcus Using X - Ray Microanalysis," Limnology and Oceanography, vol. 48, no. 5, pp. 1732­1743, 2003. [72] G. Steigman, "Big Bang Nucleosynthesis: Probing the First 20 Minutes," in Measuring and Modeling the Universe, ser. Carnegie Observatories Astrophysics Series, W. L. Freedman, Ed., vol. 2, 2004, p. 169. [73] E. M. Burbidge, G. R. Burbidge, W. A. Fowler, and F. Hoyle, "Synthesis of the Elements in Stars," Reviews of Modern Physics, vol. 29, pp. 547­650, 1957. [74] Y. Amelin, A. N. Krot, I. D. Hutcheon, and A. A. Ulyanov, "Lead Isotopic Ages of Chondrules and Calcium-Aluminum-Rich Inclusions," Science, vol. 297, no. 5587, pp. 1678­1683, 2002. [75] K. Lodders, H. Palme, and H. -P. Gail, "Abundances of the elements in the solar system," ArXiv e-prints, 2009.