Features
In this artist’srendition, DNAcontaining lipid
vesicles form during
the hydration phaseat a hydrothermal
pool. The inset
shows vesicles 10 to20 µm in diameterbudding from a dried
mixture of lipid. Thevesicles containDNA that has
been stained withacridine orange.

ORIGINS OF LIFE

Researchers design cell-like compartments to figureout how Earth’s first cells might have developed
CELIA HENRY ARNAUD, C&EN WASHINGTON

U

nfortunately for scientists, there is no videotape of theuniverse’s history that they can rewind to watch how life
got started on Earth. Instead, they must recapitulate life in
the lab to find plausible models of what happened billionsof years ago. They start with chemical mixtures that might have formed
under conditions on early Earth and see how those scenarios play out.
They especially want to figure out how cells might have formed.
For example, scientists have made significant progress in showing that potentialbuilding blocks of life can spontaneously
form from simple mixtures. But building
blocks alone do not equal life. Scientists are
now trying to figure out how those building
blocks could assemble into something wemight recognize as an early cell.
“There might be many conditions thatgive you suitable building blocks,” says Wilhelm T. S. Huck, a researcher at Radboud
University who is trying to construct living
cells from simple compounds. “The key
problem is how you get from those building
blocks—whatever they were exactly—tosomething that would really be considered aliving cell.”

20 C&EN | CEN.ACS.ORG | FEBRUARY 29, 2016

To try to solve that problem, scientists
study so-called protocells. These cell-likecompartments can take many forms: Most
protocell researchers work with nucleic
acids or other biomolecules enclosed inlipid vesicles, but some researchers thinkthat protocells might not have needed membranes. Some teams have included modern
polymerase enzymes in their compartments,
but others want to find nucleic acids capableof catalyzing their own replication.
But none of these protocells have yet integrated the three capabilities that scientists
equate to life: storage of information (including the instructions for self-replication),
metabolism, and Darwinian evolution.
Instead, much of the protocell work has

focused on the first capability, information
storage. Multiple teams have recently shownways that protocells can grow and divide,both of which are steps on a possible path toself-replication.
In one study, Tadashi Sugawara ofKanagawa University and coworkers described protocells that went through four
steps to divide, demonstrating a primitivemodel of a conventional cell cycle. These
protocells, which consist of bacteria-sized
lipid vesicles, enclose DNA and polymerase
enzymes to replicate DNA (Nat. Commun.
2015, DOI: 10.1038/ncomms9352). In addition to the phospholipids POPC and POPG,
the vesicle membranes contain anothercationic lipid and an amphipathic catalyst,
which has an imidazolium head group anda lipid tail. The catalyst can form a complex with DNA and the cationic lipid thatgenerates more of the lipid from precursor
molecules.
During the first step of the cycle, theingestion phase, an increase in the solution’s pH causes the protocells to fuse with
so-called conveyor vesicles that deliver
nucleotides. In the replication phase, theprotocell’s DNA polymerase uses those nucleotides to make more DNA. In the maturation phase, replicated DNA protrudes into
the membrane to form the catalyst complex.
The researchers add lipid precursors, andthis complex starts generating lipids. As a re-RYAN NORKUS/ LIFE 2015, DOI: 10.3390/LIFE 5010872

With protocells, scientists probe
the chemistry that started biology

COURTESY OF TADASHI SUGAWARA (CE LL CYCLE); NAT. CO MMUN. (MICROGRAPHS)

sult, the protocell membrane starts to grow.
When the membrane becomes unstable, theprotocell divides into two daughter vesicles
during the final phase of the cycle.
The researchers demonstrated two complete cycles but claim more cycles should bepossible. “Our model protocells continue toreplicate DNA as long as nucleotides and thepolymerase are occasionally supplied fromoutside,” Sugawara says.
Vincent Noireaux, a biophysicist at theUniversity of Minnesota, Twin Cities, whoalso develops protocells, notes that eachstep in Sugawara’s model cell cycle is triggered by an external stimulus. “It’s greatwork, but there is still a lot to accomplish
and understand to get a fully autonomouscompartment capable of sustaining itself
without any sort of activation from outside.” Still, he acknowledges that “devisinga minimal metabolism capable of biosynthesis for self-reproduction is incrediblychallenging.”
Another team, led by Tetsuya Yomo ofOsaka University, has also reported vesicles
that can reproduce over multiple generations (Proc. Natl. Acad. Sci. USA 2016, DOI:
10.1073/pnas.1516893113). They mix two
liposome populations, one that contains
RNA and another that contains nucleotides
and the enzyme RNA replicase, which copies RNA. The researchers use freeze-thawcycles to fuse the liposomes and stimulate
RNA replication. When the liposomes
reach a certain size, they divide into smaller
liposomes. Yomo and coworkers were able
to sustain the system for 10 cycles.
Both Yomo and Sugawara use membrane components found in modern cells
instead of simpler compounds that may
have been floating around on early Earth.
That’s because they’re less interested inThis time seriesof micrographs
shows Sugawara’s
protocell dividing
into two daughter
protocells.

Primitivemodel cellcycle
Sugawara’s model
cell cycle has fourstages. In theingestion phase,vesicles deliver
nucleotides toRNA-containingprotocells. In thereplication phase,the protocells use
those nucleotides
to copy the RNA.
In the maturation phase, lipids
from the vesicles
are incorporated
into the protocell
membrane. Finally,
the protocell grows
large enough thatit divides into two
daughter protocells.

how life originally began on Earth than how life arises fromchemistry.
“I am interested in constructing a molecular system thatexhibits dynamic events, such as self-reproduction, recursive proliferation, and evolution, which must be an essential
feature of life,” Sugawara says. “We cannot address questions about the origin of each material, such as protein, RNA,and POPC, but we can address the question of how thosematerials mix to start targeted cellular functions,” Yomosays. Because he uses modern biological molecules, Yomo
avoids calling his structures protocells. “It’s better to call it
an artificial cell model,” he says.
While Yomo, Sugawara, and most other protocell researchers study their systems inside laboratories, David Deamer and Bruce Damer of the University of California, SantaCruz, have gone out on field trips, searching for conditions
that approximate those on prebiotic Earth. Volcanic hydrothermal pools seem to fit the bill.
Deamer recently visited the Bumpass Hell section of
Mount Lassen, a volcano in California. Here, sulfurous
volcanic gases waft up from holes in the hydrothermal field
called fumaroles. Such areas go through wet/dry cycles withrain or hot springs filling puddles that then later evaporate.
In his experiments, Deamer exposes a mixture of lipids
and nucleotides for a couple of hours to the acidic gases
coming out of the fumaroles. After the mixture dries out, hetakes it back to the lab and extracts polymers from the lipidmatrix. “We get something that looks like RNA,” he says.
In previous work, Deamer and colleagues at McMaster
University showed that the nucleotides in a lab version ofthis experiment are trapped between lipid layers (PLOS One
2013, DOI: 10.1371/journal.pone.0062810). It’s only a small
step from there to a protocell in which the polymerized nucleotides are encapsulated by a lipid membrane.
“I encourage my colleagues to take their experiments to aplace like Bumpass Hell,” Deamer says. “Most of them rebel
against that idea because it seems so uncontrolled. But they’remissing so much that can happen in these conditions.”
Yet not all researchers venture to volcanic fields to try to
mimic prebiotic conditions. For some of these lab-boundscientists, the goal is to replicate nucleic acids inside protocells without enzymes, because enzymes are not likely toFEBRUARY 29, 2016 | CEN.ACS.ORG | C&EN21

have been part of primitive cells.
“You can’t have evolution unless thegenetic material is able to replicate,” says
Nobel Laureate in Physiology or Medicine
Jack W. Szostak of Harvard Medical School,
who studies protocells. But there’s not yet a
way of doing complete cycles of RNA replication and cell division without enzymes.
Szostak wants to figure out whether
membranes might promote RNA reactions without modern enzymes. Such a
process would require a way for RNA to
get transported to the interior surface of

22 C&EN | CEN.ACS.ORG | FEBRUARY 29, 2016a membrane. Last year, Szostak’s group
showed that amphipathic peptides can direct RNA molecules to vesicle membranes(Angew. Chem. Int. Ed. 2015, DOI: 10.1002/
anie.201505742).
They developed a system involving tiny,
100-nm-sized vesicles encapsulating undecylimidazole—a model for cationic, amphipathic peptides—and larger, 5- to 25-µmsized vesicles containing RNA. When thetwo vesicles were mixed, the tiny vesicles
brought the undecylimidazole to the surface of the larger vesicles. Within a shortperiod of time, the RNA started to associatewith the membrane.
Researchers have focused on protocell
replication because it’s necessary for evolution, which most people consider to be adefining characteristic of biology. “Only aprotocell that gained evolvability could have
led to present cells,” Yomo says.
But Kepa Ruiz-Mirazo, who studies origins of life at the University of the BasqueCountry, thinks metabolism is the hardest
part of the protocell problem. Modern
metabolism depends on enzymes. “Butyou can’t start with enzymes. They are theresult of the whole process,” he says. If you
look at the possible sources of prebiotic
molecules, such as meteorites, there are noenzymes. “Enzymes are the product of lifeand probably came last in the origins of life,”
Ruiz-Mirazo says.
Ricard Solé, who heads the Complex
Systems Lab at Pompeu Fabra University,
thinks that modeling the thermodynamics
underpinning protocells could point the way
to a nonenzymatic metabolism. His grouphas computationally and theoretically modeled the physical chemistry of a system inwhich the lipid decanoic acid aggregates intoa membranelike structure around decanoic
anhydrides, which are precursors of the lipid
molecules.
“It was an extremely simple metabolism,
which was essentially a single reaction thattransforms precursors in the soup intolipids,” Solé says. “When the precursor
molecules approach the membrane, theyquite easily become new lipids, so the protocell keeps growing. But eventually thisdestabilizes and splits.” With a steady supplyof precursors, the model predicts that thesystem can undergo many cycles (2015,arXiv: 1503.04683v1). Solé describes it as theminimal self-replicating model that doesn’tinvolve RNA or other information storage.
Still, a protocell with all three capabilities is a long way away. “I’m optimistic thatsomebody will get to a point where we have afunctioning protocell model where the vesicle compartment replicates and its nucleic
acid contents can both replicate and carryout simple functions,” Szostak says. “Thatopens up a lot of interesting new questions
we can ask.” For example, researchers could
then study how different chemical and physical conditions affect the evolution of thesystem.
“What we can do as scientists is showwhat is possible, that there are conditions
that allow molecules to organize in a certainway,” says Pasquale Stano, an origins-of-liferesearcher at Roma Tre University. “We’renot going to prove that life originated in acertain way but that, given some conditions,
we don’t need a miracle.” ◾