Monday, 13 August 2007

Countdown to a synthetic lifeform

Countdown to a synthetic lifeform
11 July 2007 news service Peter Aldhous

Synthetic life could be just around the corner - depending on what you mean by "synthetic".
Last week, genomics pioneer Craig Venter announced that his team has passed an important milestone in its efforts to create a bacterial cell whose genome is entirely synthetic - constructed chemically from the building blocks of DNA. Venter claims this goal could be achieved within months.
But while Venter's synthetic genome will be housed within an existing bacterial cell, other scientists are aiming for the even more ambitious target of building an entire living cell from the basic chemical ingredients. Giovanni Murtas of the Enrico Fermi Centre at the University of Rome 3, Italy, reported last week at the Synthetic Biology 3.0 meeting in Zurich, Switzerland, that his team had taken a step toward this goal by successfully synthesising proteins in cell-like compartments.

According to George Church at Harvard Medical School in Boston, who has devised a complete blueprint for a synthetic cell, an investment of around $10 million would be enough to turn the "bottom-up" dream into reality. "Our approach doesn't require any super new technology," he says. Whichever definition of synthetic life you adopt, it seems now to be a question of when rather than if. "We are at the doorstep of being able to create life," says Steen Rasmussen, a physicist trying to create artificial living systems at the Los Alamos National Laboratory in New Mexico.

“Whichever definition of synthetic life you adopt, it seems now to be a question of when rather than if”
Murtas and his team have managed to initiate the process of protein synthesis in cell-like self-assembling spheres bounded by lipid membranes, known as "liposomes". A similar feat was achieved in 2004 by Vincent Noireaux and Albert Libchaber of Rockefeller University in New York, but while they seeded their lipid vesicles with an extract of Escherichia coli bacterial cells, Murtas and his colleagues used a recipe of 37 enzymes and a range of smaller molecules to enable protein synthesis.

After drying lipid molecules onto the walls of a plastic tube, Murtas's team added the mix of enzymes and chemicals, plus the gene for green fluorescent protein (GFP). Some of the resulting liposomes subsequently made GFP for several hours.

This is clearly some way from a living cell, and to obtain something indisputably alive the genetic material needs to copy itself, and the vesicles divide. With this in mind, Murtas and his colleagues are trying to incorporate into their vesicles genes for enzymes that can form new lipids, which they hope will make the liposomes grow to the point that they split apart to form smaller daughter vesicles.

Murtas is interested in synthetic cells as a model of what happened when the earliest forms of life emerged. His team's achievement falls short of a true bottom-up construction because the recipe they used for protein synthesis had to include structures known as ribosomes, composed of RNA and proteins, which they obtained from E. coli. These "biochemical machines" direct protein synthesis, and to be truly synthetic a cell would have to include structures capable of a similar job that were assembled from their basic components.
"That is probably the biggest challenge," says Church. Though biochemists have been able to assemble ribosomes in the lab for some years, it has required high temperatures and harsh chemical conditions - not the sort of environment to be found in living cells.
But progress is being made, and last year Church, working with Tony Forster of Vanderbilt University in Nashville, Tennessee, published a detailed blueprint for assembling a synthetic cell from scratch (Molecular Systems Biology, DOI: 10.1038/msb4100090). It includes 115 genes which would be combined with various biochemicals to make a self-assembling cell able to live under carefully controlled lab conditions. The details have still to be worked out, but Church believes there should be no fundamental barriers. He sees the team's artificial organism becoming a workhorse for biotechnology that could be adapted to do useful tasks such as making complex biochemicals.

Despite the scope of Forster and Church's vision, it is Venter and his team who are grabbing the headlines. They have been working for years to develop a minimal genome containing less than 400 genes but which nevertheless has everything it takes to sustain a free-living cell. They have investigated which genes are essential by a process of elimination: knocking out genes in the bacterium Mycoplasma genitalium, which itself has an exceptionally small genome. Venter ignited controversy last month by trying to patent the resulting minimal genome (New Scientist, 16 June, p 13),

Venter's next step will be to synthesise the minimal genome, and put it into a bacterial cell, and for this he needs a technique for replacing a bacterium's natural genome with a synthetic one. "It's really essential to what they want to do," says David Deamer, a biophysicist at the University of California, Santa Cruz.

Pic1 : Transplanting a genome

The novel "genome transplant" that Venter's team announced last week has proved in principle that they can do just that. The researchers, led by John Glass of the J. Craig Venter Institute in Rockville, Maryland, managed to transfer the genome of Mycoplasma mycoides to a related parasite called Mycoplasma capricolum. Both species infect goats, sheep and cows. Judging from the proteins they produced, the resulting cells seem to have been completely transformed into M. mycoides (Science, DOI: 10.1126/science.1144622).

The team took a strain of M. mycoides resistant to the antibiotic tetracycline, broke open the cells and then used enzymes to digest away their proteins, leaving its circular chromosomes intact. Next, they incubated these chromosomes with M. capricolum cells in a culture medium containing a polymer called polyethylene glycol (PEG). PEG makes cell membranes fuse, and the researchers speculate that some M. capricolum cells fuse together, encapsulating an M. mycoides chromosome as they do so. The cells containing multiple genomes soon divide, putting one genome into each daughter cell.

The researchers then treated their cultures with tetracycline, which wipes out those containing the host M. capricolum genome while cells containing the M. mycoides genome survive (see Diagram). Though the transplantation worked in only about 1 in every 150,000 cells, that was enough to give healthy colonies of transformed bacteria containing no M. capricolum DNA.
Venter says that efforts to synthesise his minimal genome from scratch are still in progress, but once it is ready, the transplant method should allow the first bacterium with a synthetic genome to be created with little delay. "It could be weeks or months," he says. Not everyone accepts that Venter's bacterium will qualify as a "synthetic" organism. "It's a misnomer," says Deamer, who argues that a better name would be a radically engineered organism.

“Not everyone accepts Venter's bacterium as truly 'synthetic', and say it would be better to call it radically engineered”
So when are we likely to see unequivocally synthetic life, with the entire cell built from scratch? "It could be five months or 10 years," says Church. "These things aren't so much a question of timescales as the amount of money available."

From issue 2611 of New Scientist magazine, 11 July 2007, page 6-7
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