Daniel Vaulot and Adriana Lopes dos Santos

Origin of Eukaryotes

2023-06-19

Outline

• Early Life Evolution

• How many domains ?

• The endosymbiotic theory

Early Life Evolution

Early History of Earth

• Earth is ~ 4.5 billion years old. There is not a single rock that is older than this…
• Liquid water required for life: first appeared ~ 4.3 billion years ago (volcanic outgassing and icy comets/asteroids).
• Evidence for life ~ 4.1 billion years ago (carbon isotope ratios, life prefers C₁₂)

Parameters that limit life

• The laws of chemistry and physics have universal principles which enable us to extrapolate to the conditions under which life would have appeared and could survive elsewhere.
• These principles suggest that life requires:
  • a liquid solvent
  • an energy source
  • building blocks (aas, lipids, nucleotides, etc).
• Experimental evidence indicates that organic molecules such as RNA nucleotides, amino acids, and lipids can form spontaneously under conditions that were present on the early Earth, providing the preconditions needed for the first living systems (e.g. Oparin, Stanley Miller, Juan Oro, Jeffrey Bada and others… JPL Astrobiology research USA)
• Temperature, pH, pressure, salinity, and radiation can influence the availability of nutrients and energy sources,
• Microbial diversity is structured by these parameters in any environment.
• Identifying the minima and maxima of each parameter and their combined effects, is imperative to advance our understanding of limits of life on Earth and the potential existence of life elsewhere.

Life can adapt to a wide range of parameters

• Microorganisms (Bacteria and Archaea) have been detected in a variety of extreme environments and virtually in any location where liquid water is available for life to use.

https://www.frontiersin.org/articles/10.3389/fmicb.2019.00780/full

Hyperthermophilic ancestor? On going debate

• The surface of the early Earth is thought to have been much hotter than today. Only superheat-loving microbes (hyperthermophiles) would have been able to thrive and survive in such an ‘early times of life’ scenario.
• Hyperthermophilic archaea and bacteria are found on the deepest, shortest branches of the SSU rRNA phylogenetic tree (thick branches)
• Hyperthermophilic bacteria and archaea often display chemotrophic metabolism where compounds like molecular hydrogen, carbon dioxide, sulphur and ferric/ferrous iron are used as electron donors.

LUCA: Last Universal Common Ancestor
Stetter, K. O. Hyperthermophiles in the history of life. Philos. Trans. R. Soc. B Biol. Sci. 361, 1837–1842 (2006).

Molecular adaptations to life at high temperature

• RNA stability: higher GC content
• DNA stability
  • high intracellular solute levels stabilize DNA
  • reverse DNA gyrase (found only in hyperthermophiles): introduces positive supercoils into DNA
  • DNA-binding proteins (histones) compact DNA into nucleosome-like structures
• Protein folding
  • Chaperones: class of proteins that refold partially denatured proteins
• Lipid stability: possess dibiphytanyl tetraether type lipids; form a lipid monolayer membrane structure

Reverse DNA gyrase

Results suggest a nonhyperthermophilic LUCA and bacterial ancestor, with hyperthermophily emerging early in the evolution of the archaeal and bacterial domains.

Life originated at hydrothermal systems on ocean floor ?

• The extremely hot temperatures and levels of ultraviolet radiation at Earth’s surface were likely to be very hostile to the origin of life as we know it.
• Life may have originated at hydrothermal systems where conditions would have been more stable.
• Steady and abundant supply of energy (e.g., H₂ and H₂S) may have been available at these sites.
• Geochemistry can support abiotic production of molecules required for life (e.g., amino acids, lipids, sugars, and nucleotides), as shown by the experiment before hydrothermal vents were discovered.

How many domains

The three domains of life - Carl Woese

• Distinctive ribosomal RNAs and tRNAs,
• Absence of murein cell walls
• Presence of ether-linked lipids in phytanyl chains.

Molecular features shared only between archaea and eukaryotes appeared consistent with 3 domains hypothesis

• DNA replication machinery
• DNA packed by histones in nucleosome-like structures
• Structurally complex RNA polymerases
• Translation machinery

Two vs Three domains of life

• In 1984, based on analysis of rRNA, the evolutionary biologist James Lake (University of California, Los Angeles) proposed that eukaryotes are sisters to archaea that he called eocytes, which means dawn cells. The idea evolved into the two-domain scenario.

• In this evolutionary scenario, Archaea and Bacteria represent the only primary domains of life, and eukaryotes later emerged from lineages within the Archaea domain.
  

Lake, J. A. Origin of the eukaryotic nucleus determined by rate-invariant analysis of rRNA sequences. Nature 331, 184–186 (1988).

Sampling the deep-sea

In 2010, a 2m long gravity core (GC14) was retrieved from the Arctic Mid-Ocean Ridge during summer, approximately 15 km northwest of the active venting site Loki’s Castle (between Greenland and Norway), at 3283m below sea level.

Why Loki ?

Loki is a god in Norse Mythology. He creates trouble. He is elusive and ambiguous.

Discovery of Lokiarchaeota - 2015

• A team of microbiologists, led by Thijs Ettema of Uppsala University in Sweden, had been studying since 2010 a subset of Archaea lineages, called the Deep-Sea Archaeal Group (DSAG) or TACK (Thaumarchaeota, Aigarchaeota, Crenarchaeota and Korarchaeota).

• They sequenced DNA extracted directly from Loki samples and assembled genomes exclusively from sequences (Genome-resolved metagenomics)

• They discovered a new archaea phylum called Lokiarchaeota which branches with eukaryotes.

Spang, A. et al. Complex archaea that bridge the gap between prokaryotes and eukaryotes. Nature 521, 173–179 (2015).

Metagenome assembled genomes

https://merenlab.org/2020/06/27/seminar-series-on-microbial-omics/

Asgard superphylum

• In 2016 and 2017 Ettema’s team and other researchers announced the discovery of 3 new archaeal phyla related to the Lokis: Thorarchaeota, Odinarchaeota, Heimdallarchaeota.

• The entire grouping of new superphylum was named Asgard after the realm of the Norse gods.

• Phylogenomic analyses strongly indicated that the new archaea lineages representing the Asgard superphylum form a monophyletic group with eukaryotes.

• Members of Asgard were found to carry genes encoding proteins that were assumed to be specific to eukaryotes (e.g. actins).

Zaremba-Niedzwiedzka, K. et al. Asgard archaea illuminate the origin of eukaryotic cellular complexity. Nature 541, 353–358 (2017).

Asgard superphylum

Everybody was not convinced

The final proof - Isolation of a pure co-culture

• Took long time …. “Through subsequent transfers, we were able (…) to obtain a pure co-culture of the target archaeon MK-D1 and Methanogenium after a 12-year study—from bioreactor-based pre-enrichment of deep-sea sediments to a final 7 years of in vitro enrichment.”

• Doubling time was estimated to be approximately 14–25 days

• The genome encodes proteins that are only found in Eukarya (eukaryotic signature proteins): membrane trafficking, vesicle formation and/or transportation, ubiquitin and cytoskeleton formation

• It catabolizes ten amino acids and peptides through syntrophic growth with Halodesulfovibrio (sulfate-reducing bacterium) and Methanogenium (methanogenic Archaea)

Imachi, H. et al. Isolation of an archaeon at the prokaryote–eukaryote interface. Nature 577, 519–525 (2020)

Candidatus Prometheoarchaeum syntrophicum

• Etymology:

  • Prometheus (Greek): a Greek god who shaped humans out of mud and gave them the ability to create fire; archaeum from archaea (Greek): an ancient life.  
  • Syn (Greek): together with; trophy (Greek) nourish; icus (Latin) pertaining to.

• The cells produce long branching (g) and straight (h) membrane protrusions

• Cytoplasmatic membrane contains “biphytane”(j) (probably monolayer)

An ever-evolving story - June 2023

Eme, L. et al. Inference and reconstruction of the heimdallarchaeial ancestry of eukaryotes. Nature 1–8 (2023)

Endosymbiotic Origin of Eukaryotes

Earth’s history

• LUCA emerged at least 3.5 - 4 billion years ago.
• An explosion of metabolisms (metabolic diversification) occurred allowing life (microbes) to exploit the various resources available on Earth.
• Oxygenic photosynthesis in the ancestors of cyanobacteria appeared on ~ 3.2 billion years ago

Earth’s history

• Oxygen accumulated in the atmosphere lead to the Earth’s first mass extinction (GOT).
• The rising atmospheric oxygen triggered cellular compartmentalization and eukaryogenesis
• LECA emerged ~1–1.9 billion years ago
• Eukaryogenesis – the emergence of eukaryotic cells – represents a pivotal evolutionary event.
• The compartmentalization of cellular processes allowed the evolution of complex multicellular organisms.

The origin of the eukaryotic cell

Most early models envisaged that eukaryotic cells evolved from prokaryotic cells (bacteria) by a process of complexification.

The role of symbiosis

• The idea was originally suggested by Schimper in 1883 in a small note.

• Famintsyn (1889) worked extensively on algal symbiosis. He also attempted to culture chloroplasts outside the of the host.

• Merezhkowsky (1905) remarked the similar structural characteristics of cyanobacteria and chloroplasts.

Lynn Margulis

• In 1967, Lynn Margulis championed the idea of an endosymbiotic origin of organelles and further hypothesized that not only mitochondria, chloroplast have symbiotic origin but also cell flagella derived from symbiotic spirochetes (this one she got wrong!).

• She presented a comprehensive symbiotic view of eukaryotic cell evolution based on cytological, biochemical, and paleontological evidence in her seminal work “On the origin of mitosing cells”.

The endosymbiotic theory

Molecular phylogeny

Part of Margulis ideas started to become credible with the advent of molecular phylogeny that allowed demonstrating that conserved genes in chloroplast and mitochondrial genomes clustered with, respectively, cyanobacterial and alphaproteobacterial genes in phylogenetic trees.

Schwartz, R. M. & Dayhoff, M. O. Origins of Prokaryotes, Eukaryotes, Mitochondria, and Chloroplasts. Science 199, 395–403 (1978).

Chloroplast - Primary endosymbiosis

• Photosynthetic eukaryotes emerged from a heterotrophic eukaryote which ate a gram-negative cyanobacterium (2 membranes),
• The cyanobacterium was retained and not digested.
• The ingested cyanobacterium gradually integrated into the cellular machinery as a new organelle: the plastid.

Primary endosymbiosis occured at least twice

• The first occurred ~ 1.6 billion years ago and gave rise to glaucophytes, red algae and green algae (the ancestors of all plants)
• The second occurred ~ 90–140 million years ago, establishing a permanent photosynthetic compartment (the chromatophore) in amoebae in the genus Paulinella. 

Chloroplast - Secondary endosymbiosis

• Secondary symbiosis was the process of a heterotrophic eukaryote engulfing a green or red algal cell (also eukaryotes).
• The retained, non-digested cell, became gradually the chloroplast.

Chloroplast - Secondary endosymbiosis

• Red, green, and glaucophyte algae (and plants) have tremendous diversity but they still only account for a fraction of the diversity of photosynthetic eukaryotes.
• Secondary plastids are characterized by additional membranes surrounding the plastid
  • Two membranes from primary plastid (cyanobacteria)
  • Cytoplasmic envelope of the primary algal endosymbiont.
  • Phagocytotic vacuole in which the algal prey was taken up.

The nucleomorph

Cryptophytes and Chlorarachniophytes retained a nucleomorph which is a remnant of the nucleus of the primitive photosynthetic cell. This is a further evidence of the endosymbiotic theory.

The apicoplast

• Apicomplexa are non-photosynthetic successful group of endoparasites of animals, from corals to humans (e.g. Plasmodium causing malaria).

• Evolved from free living algae.

• Contain an apicoplast which is a plastid that lost its photosynthetic ability.

• Bound by four membrane. Own genome and expresses a small number of genes.

Foth, B. J. & McFadden, G. I. The apicoplast: A plastid in Plasmodium falciparum and other apicomplexan parasites. in International Review of Cytology vol. 224 57–110 (Academic Press, 2003).

Protist diversity

Take home messages

• Origin of eukaryotes is key question

• Discovery of Asgard induce change of paradigm

• Endosymbiosis is key to understand evolution of eukaryotes

Questions ?