[astro] did life evolve in ice?

[Eugen Leitl]

http://discovermagazine.com/2008/feb/did-life-evolve-in-ice/article_print

Did Life Evolve in Ice?

02.01.2008

Funky properties of frozen water may have made life possible.

by Douglas Fox

scientist Hauke TrinksHauke Trinks spent 13 months in the Far North studying
ice and its potential as an incubator for life.

Image courtesy of Marie Tieche

One morning in late 1997, Stanley Miller lifted a glass vial from a cold,
bubbling vat. For 25 years he had tended the vial as though it were an exotic
orchid, checking it daily, adding a few pellets of dry ice as needed to keep
it at –108 degrees Fahrenheit. He had told hardly a soul about it. Now he set
the frozen time capsule out to thaw, ending the experiment that had lasted
more than one-third of his 68 years.

Miller had filled the vial in 1972 with a mixture of ammonia and cyanide,
chemicals that scientists believe existed on early Earth and may have
contributed to the rise of life. He had then cooled the mix to the
temperature of Jupiter’s icy moon Europa—too cold, most scientists had
assumed, for much of anything to happen. Miller disagreed. Examining the vial
in his laboratory at the University of California at San Diego, he was about
to see who was right.

As Miller and his former student Jeffrey Bada brushed the frost from the vial
that morning, they could see that something had happened. The mixture of
ammonia and cyanide, normally colorless, had deepened to amber, highlighting
a web of cracks in the ice. Miller nodded calmly, but Bada exclaimed in
shock. It was a color that both men knew well—the color of complex polymers
made up of organic molecules. Tests later confirmed Miller's and Bada’s
hunch. Over a quarter-century, the frozen ammonia-cyanide blend had coalesced
into the molecules of life: nucleobases, the building blocks of RNA and DNA,
and amino acids, the building blocks of proteins. The vial’s contents would
support a new account of how life began on Earth and would arouse both
surprise and skepticism around the world.

Although one of Miller’s final experiments, it certainly wasn’t the final
word. The last several years have seen a steady stream of corroborating
evidence, including one experiment—so new it has not yet been published—that
Miller’s colleague, the late Leslie Orgel, called “astonishing.”

For decades, those studying the origin of life have imagined that it emerged
in balmy conditions from primordial soups, tropical ponds, even boiling
volcanic vents. Miller and a few other scientists began to suspect that life
began not in warmth but in ice—at temperatures that few living things can now
survive. The very laws of chemistry may have favored ice, says Bada, now at
the Scripps Institution of Oceanography in La Jolla, California. “We’ve been
arguing for a long time,” he says, “that cold conditions make much more
sense, chemically, than warm conditions.”

polar bearImage courtesy of Marie Tieche

Miller’s frozen experiment is a striking testament to the idea. Although life
requires liquid water, small amounts of liquid can persist even at –60°F.
Microscopic pockets of water within the ice may have gathered simple
molecules like the ones Miller synthesized, assembling them into longer and
longer chains. A single cubic yard of sea ice contains a million or more
liquid compartments, microscopic test tubes that could have created unique
mixtures of RNA that eventually formed the first life.

If life on Earth arose from ice, then our chances of finding life elsewhere
in the solar system—not to mention elsewhere in the galaxy—may be better than
we ever imagined.

The vial of ammonia and cyanide chilling in Miller’s lab was just one of the
chemical cocktails he kept, aging like wine in a cellar. Some of the samples
sat in freezers, others under the sink, and still others in water baths
maintained at various temperatures. They were part of an effort to understand
chemical reactions that must have unfolded over millennia on early Earth. The
location of every sample was stored in Miller’s head; occasionally he would
give one to a student to analyze.  +++

Matthew Levy, once a graduate student of Miller’s and now a molecular
biologist at the Albert Einstein College of Medicine in New York City,
recalls being handed one of the 25-year-old samples to work on. “I was
scared,” he says. “I was thinking, these samples are older than I am.” Levy
burned holes in his shirts over the next few weeks as he dissolved the
samples with hydrochloric acid and ran them through an instrument called a
high-performance liquid chromatograph to identify the chemicals that had
formed. Red and green pens on the device traced out telltale peaks on a
scrolling strip of paper. Those peaks corresponded to seven different amino
acids and 11 types of nucleobases.

“What was remarkable,” Bada says, “is that the yield in these frozen
experiments was better, for some compounds, than it was with room-temperature
experiments.”

There were people who found the results a little too remarkable. When Bada
and Miller submitted their findings to a top-tier science journal, the
article was rejected. A reviewer of the manuscript felt that those molecules
must surely have formed while the samples were thawing, not while frozen at
the ridiculously low temperature of –108°F. So Miller, Bada, and Levy did
more experiments to show that thawing played no role. They published their
results in another journal, Icarus, in 2000.

The skepticism they faced was understandable. Chemical reactions do slow down
as the temperature drops, and according to standard calculations, the
reactions that assemble cyanide molecules into amino acids and nucleobases
should run a hundred thousand times more slowly at –112°F than at room
temperature. By that reckoning, even if Miller had run his experiment for 250
years—let alone 25—he should have seen nothing.

This is the main argument against Miller’s experiment, and against a cold
origin of life in general. But strange things happen when you freeze
chemicals in ice. Some reactions slow down, but others actually speed
up—especially reactions that involve joining small molecules into larger
ones. This seeming paradox is caused by a process called eutectic freezing.
As an ice crystal forms, it stays pure: Only molecules of water join the
growing crystal, while impurities like salt or cyanide are excluded. These
impurities become crowded in microscopic pockets of liquid within the ice,
and this crowding causes the molecules to collide more often. Chemically
speaking, it transforms a tepid seventh-grade school dance into a raging
molecular mosh pit.

“Usually as you cool things, the reaction rates go down,” concluded Leslie
Orgel, who studied the origins of life at the Salk Institute in La Jolla,
California, from the 1960s until his death last October. “But with eutectic
freezing, the concentrations go up so fast that they more than make up” for
the difference.

Cyanide is a good candidate as a precursor molecule in the life-in-a-freezer
model for several reasons. First, planetary scientists suspect that cyanide
was abundant on early Earth, deposited here by comets or created in the
atmosphere by ultraviolet light or by lightning (once the atmosphere became
oxygen rich, 2.5 billion years ago, the process would have stopped). Second,
although cyanide is lethal to modern animals, it has a convenient tendency to
self-assemble into larger molecules. Third, and perhaps most important, no
matter how much cyanide rained down, it could become concentrated only in a
cold environment—not in warm coastal lagoons—because it evaporates more
quickly than water.

“The strong point of freezing,” according to Orgel, “is that you concentrate
things very efficiently without evaporation.” Freezing also helps preserve
fragile molecules like nucleobases, extending their lifetime from days to
centuries and giving them time to accumulate and perhaps organize into
something more interesting—like life.

Orgel and his coworkers proposed these ideas in 1966, when he showed that
frozen cyanide efficiently assembles into larger molecules. Alan Schwartz, a
biochemist at the University of Nijmegen in ut to explore whether a layer of
ice covering early Earth’s oceans might have gathered and assembled organic
molecules.  +++

scientist Hauke TrinksTrinks operates his microscope at an improvised desk. .

Image courtesy of Hauke Trinks

With a few crates of supplies and two sled dogs, Trinks and his partner,
Marie Tieche, hunkered down in a cabin on Nordaustland for 13 months. Each
morning they monitored the temperature of the ice and prepared the day’s
experiments. To study the networks of liquid pockets, Trinks injected dyes
into the ice and watched through a microscope as they spread.

Winter deepened, 24-hour darkness descended, and the mercury plummeted to
–20°F. Trinks continued his experiments, sometimes banging pans together to
chase polar bears away. Once a walrus lunged up through the ice and dragged
several of Trinks’s instruments into the ocean.

He built a makeshift lab table from planks of wood and discarded gasoline
cans. He examined slices of sea ice under the microscope, his hood pulled
tight around his eyes. Turning a knob with a gloved hand, he nudged a metal
electrode nearly as fine as a red blood cell closer to an ice crystal. The
needle on his voltmeter jerked sideways, registering a sharp drop in voltage
on the crystal’s surface—evidence of a microscopic electric field that might
arrange and orient molecules on the ice’s surface.

By the time Trinks returned to Hamburg in 2003, he had formulated a theory
that ice was doing much more than just concentrating chemicals. The ice
surface is a checkerboard of positive and negative charges; he imagined those
charges grabbing individual nucleobases and stacking them like Pringles in a
can, helping them coalesce into a chain of RNA. “The surface layer between
ice and liquid is very complicated,” he says. “There is strong bonding
between the surface of the ice and the liquid. Those bondings are important
for producing long organic chains like RNA.”

At a lecture in Hamburg in 2003, Trinks met up with chemist Christof
Biebricher, who was studying how the first RNA chains could have formed in
ttest it sounded messy—more like a margarita recipe than a serious scientific
investigation. “Chemists,” says Biebricher, “do not like heterogeneous
substances like ice.” But Trinks convinced him to try it in his laboratory at
the Max Planck Institute for Biophysical Chemistry in Göttingen, Germany.

Biebricher sealed small amounts of RNA nucleobases—adenine, cytosine,
guanine—with artificial seawater into thumb-size plastic tubes and froze
them. After a year, he thawed the tubes and analyzed them for chains of RNA.

For decades researchers had tried to coax RNA chains to form under all sorts
of conditions without using enzymes; the longest chain formed, which Orgel
accomplished in 1982, consisted of about 40 nucleobases. So when Biebricher
analyzed his own samples, he was amazed to see RNA molecules up to 400 bases
long. In newer, unpublished experiments he says he has observed RNA molecules
700 bases long. Biebricher’s results are so fantastic that some colleagues
have wondered whether accidental contamination played a role. Orgel defended
the work. “It’s a remarkable result,” he said. “It’s so remarkable that
everyone wants better evidence than they would for an unremarkable result.
But I think it’s right.”

Biebricher had loaded the deck somewhat, because he wasn’t growing RNA chains
from nothing. Before he froze his samples, he added an RNA template—a
single-strand chain of RNA that guides the formation of a new strand of RNA.
As that new RNA strand grows, it adheres to the template like one half of a
zipper to the other. This must be how the first genes, made of RNA, would
have copied themselves. But the first step was the formation of the original
RNA molecule that served as a template, and how that step happened remains a
mystery.

Ice may prove the crucial ingredient here, too. Deamer and his former student
Pierre-Alain Monnard (now at Los Alamos National Laboratory in New Mexico)
have run experiments frozen at 0°F for a month, without the aid of templates.
In those relatively brief experiments they already see RNA molecules How do
you get from tiny snippets of RNA to longer, well-crafted chains that could
have acted as the first enzymes, doing fancy things like copying themselves
The shortest RNA enzyme chains known today are about 50 bases long; most have
more than 100. To work effectively, moreover, an RNA enzyme must fold
correctly, which requires exactly the right sequence of bases.


A young scientist named Alexander Vlassov stumbled upon a possible answer. He
was working at SomaGenics, a biotech company in Santa Cruz, California, to
develop RNA enzymes that latch on to the hepatitis C virus. His RNA enzymes
were behaving strangely: They normally consisted of a single segment of RNA,
but every time he cooled them below freezing to purify them, the chain of RNA
spontaneously joined its ends into a circle, like a snake biting its tail. As
Vlassov worked to fix the technical glitch, he noticed that another RNA
enzyme, called hairpin, also acted strangely. At room temperature, hairpin
acts like scissors, snipping other RNA molecules into pieces. But when
Vlassov froze it, it ran in reverse: It glued other RNA chains together end
to end.

Vlassov and his coworkers, Sergei Kazakov and Brian Johnston, realized that
the ice was driving both enzymes to work in reverse. Normally when an enzyme
cuts an RNA chain in two, a water molecule is consumed in the process, and
when two RNA chains are joined, a water molecule is expelled. By removing
most of the liquid water, the ice creates conditions that allow the RNA
enzyme to work in just one direction, joining RNA chains.

The SomaGenics scientists wondered whether an icy spot on early Earth could
have driven a primitive enzyme to do the same. To investigate this, they
introduced random mutations into the hairpin RNA, shortened it from its
normal length of 58 bases, and even cut it into pieces—all in an effort to
produce RNA enzymes that were as dodgy and imperfect as early Earth’s first
enzymes likely were. These pseudoprimitive RNA enzymes do nothing at room
temperature. But freeze them and they become active, joining other RNA
molecules at a slow bouraged short segments of RNA to stick together and
behave as a single, larger RNA molecule. “Freezing stabilizes the complexes
formed from multiple pieces of RNA,” concludes Kazakov. “So small pieces of
RNA could be enzymes, not just large 50-base molecules.”

Equally telling, the pseudoprimitive RNA enzymes that Vlassov made grabbed
and joined just about any other molecule. Enzymes on early Earth might have
done the same, joining random segments of 5 or 10 RNA bases to form a variety
of sequences.

iceIn the Arctic, Trinks photographed ice under his microscope.

Image courtesy of Marie Tieche

All these processes would occur in microscopic pockets of liquid within the
ice. “You have billions and billions of different possibilities,” Trinks
says, “because you have billions of these small channels,” each like a
microscopic test tube containing a unique RNA experiment. On the young Earth,
pockets of liquid could have expanded into a network of channels that mixed
their contents during freeze-thaw cycles, like day-night temperature changes
in summer. In winter, the liquid pores would have contracted and become
isolated again, returning to their separate experiments. With all the mixing,
something special might eventually have formed: an RNA molecule that made
rough copies of itself. And as Earth warmed, these molecules might have found
a home in newly thawed seas or ponds, where something even more complex might
have emerged—such as a cell-like membrane. “You have something that is
multiplying itself, and you have variation that is inherited,” says Antonio
Lazcano, a biology researcher and professor at the National Autonomous
University of Mexico, in Mexico City?. “There you have the onset of Darwinian
evolution. I’m willing to call that living.”

No one can really know if this is how life began. Other theories posit that
mineral surfaces organized key molecules or volcanic sources synthesized
amino acids. These theories need not be mutually exclusive. Glaciers on early
Earth could have scooped up mineral dust; volcanoes could have rained ash
onto nearby sea ice. Primordial ice “must have been full of
impurities,” Lazcano says, “and those impurities must have had
catalytic effects, enhancing the synthesis or destruction of some compounds.”

Shortly after Miller finished his 25-year experiment, he suffered a stroke
that ended his career. His laboratory, with 40 years of samples, was emptied
in 2002 to make way for a building renovation. Experiments that had run for
years or decades were discarded without ever being analyzed. As Bada rescued
a few items from his mentor’s freezer, safety personnel stood by in hazmat
suits, sent by university officials concerned about rumors of toxic cyanide.
Any sample that couldn’t be identified was incinerated. Miller was present
for a few hours of this ordeal, struggling to find words to identify the
vials that he had known so well.

Miller died on May 20, 2007, but the provocative theory he helped nurture
lives on. In the latest twist, Miller’s ideas are influencing not just
theories about life’s origin on Earth but also investigations about the
potential for life elsewhere in the solar system. In fact, it was a dinner
conversation with Bada regarding Jupiter’s moon Europa that prompted Miller
to open his 25-year-old samples back in 1997. While most scientists were
focusing on the possibility of life in Europa’s ocean, he and Bada had been
talking about what biochemistry might happen in the 10-mile-thick layer of
ice atop the ocean. Those speculations are more relevant than ever, with
recent discoveries of geysers on Saturn’s icy moon Enceladus and elaborate
organic molecules on Titan, another Saturnian moon. Recent studies show that
Mars too has vast quantities of buried ice, especially at its poles.

If life arose in one of these frozen zones, it might still exist there.
Although life as we know it requires liquid water, there are places where
life survives well below freezing. In the microscopic veins that permeate
Arctic ice, for example, the high concentration of salt can maintain traces
of water in a liquid state down to –65°F. Bacteria and diatoms inhabit those
liquid veins, and Hajo Eicken, a glaciologist at the University of Alaska at
Fairbanks, suspects that similar habitats could exist in the lower, warmer
layers of ice on Europa, and perhaps on the other moons as well. “There’s
potentially hundreds of meters of ice, if not maybe a few kilometers, that
may well be quite habitable,” Eicken says.

Liquid water—and life—occurs in other cold places, too. Films of liquid water
persist far below freezing, like coatings of condensation, on the surfaces of
some minerals. Under some conditions, these films may stay liquid down to
–90°F. Bacteria beneath films of liquid water only several molecules thick
have been found clinging to microscopic grains of clay in ice cores from
Greenland. Slowly consuming the iron in a single grain, these bacteria could
get by for a million years before exhausting their food supply; at colder
temperatures, where metabolic demands are lower, they might survive hundreds
of millions of years.

If life arose in ice on Earth, then why not on Mars, Europa, or Enceladus?
“You’ve got to keep an open mind in this business,” Bada says. “If I were
going to make a bet about what we’d find if we discover life elsewhere in the
universe, I would suspect it would be more cold-adapted than hot-adapted.”