AsianScientist (May 27, 2016) – If you’re a fan of the Douglas Adam’s Hitchhiker’s Guide to the Galaxy series, you’ll know this one: What is the answer to the ultimate question of life, the Universe, and everything?
Far from the profound, life-changing insight expected, the answer generated by the supercomputer Deep Thought was unsatisfying; a seemingly random, discrete number that didn’t have an obvious meaning: (spoiler alert!) 42.
Realizing that their quest for the meaning of life had been thwarted, Deep Thought’s developers (or rather, their descendants 7.5 million years down the line) then asked Deep Thought to calculate what the question was in the first place. Unable to do so, Deep Thought nevertheless offered to help create something that could: a massive simulation called Sol 3, mistaken by its human occupants for a planet called Earth.
Ask synthetic biologist Craig Venter, though, and he will probably tell you that Deep Thought’s answer is one log scale off—the real answer is 473. But unlike Deep Thought, Venter has a clear question in mind: What is the minimum number of genes required for life?
Size isn’t everything
Human beings have about three billion base pairs worth of DNA, organized into 23 chromosomes. Assuming that all genetic material is protein coding (it’s not), and that the average size of a protein is between 141 and 146 amino acids (a gross underestimation), a geneticist named Friedrich Vogel estimated in 1964 that the human genome contains 6.7 million genes.
Almost 30 years later, that estimate was revised to 100,000, based on back-of-the-envelope calculations made by the National Institutes of Health. When the human genome was first sequenced in 2001, Venter’s own estimates put the number of genes at between 26,000 and 40,000. Using mass spectrometry data as a filter, one group estimated in 2014 that the true number of protein-coding genes is actually closer to 19,000—even less than the nematode worm Caenorhabditis elegans’ 20,000.
But scientists have long known that the number of genes doesn’t always correspond with biological complexity. Take for example the organism alleged to hold the title for ‘Largest Genome in the World’—the awesomely-named Polychaos dubium—a humble, algae-eating amoeba that somehow has a staggering 670 billion base pair genome, more than 200 times larger than the human one.
How low can you go?
For the past two decades, Venter and his team have been obsessed with life at the other end of the spectrum, organisms that have as few genes as possible. It is hoped that these organisms with bare-bones genomes can tell us about the minimum requirements for life, a stepping stone towards creating fully synthetic ‘artificial’ life forms.
The work began in 1995, when Venter and his team sequenced the genome of Mycoplasma genitalium, which at 580,000 base pairs and just 525 genes had the smallest known genome of any free-living organism. Switching over to the more robust M. mycoides, which had a genome twice the size but could replicate much faster, the group announced another breakthrough in 2010: The first self-replicating, synthetic bacterial cell.
To achieve this feat, Venter had to push existing technologies to their limit. Firstly, he had to figure out how to accurately synthesize the million base pair M. mycoides genome in the first place, piecing together smaller chemically synthesized fragments and stitching them together in yeast cells. Next, he faced the challenge of carefully extracting the artificial genome from yeast and transferring it to a hollowed out M. capricolum recipient.
US$40 million and 200 man-years later, Venter and his team succeeded. The resulting organism—named JCVI-syn1.0—had the cytoplasm of M. capricolum but expressed the synthetic M. mycoides genome, and was hailed as the first artificial life form.
Trial and error
With all the tools in place, Venter and his team were finally prepared to answer the question of the minimum number of genes required for life. But the process turned out to be much more complicated than he had envisioned.
Like any good experimentalist, he first took the rational approach, using information from scientific literature to sieve out 440 genes that were likely to be non-essential. When those 440 genes were removed from syn1.0, however, the cells did not survive.
Venter then switched over to an empirical approach, devising an iterative design-build-test scheme. Starting with syn1.0’s full set of over 900 genes, the researchers eliminated over 90 percent of the genes that were determined to be non-essential by mutagenesis experiments. The resulting half-sized genome—called RGD1.0—was then chopped up into eight more or less equal fragments. Each of the RGD1.0 fragments were then reinserted into a partial syn1.0 genome containing the remaining seven-eighths, and then transplanted back into M. capricolum to see if live colonies could be grown.
All the M. capricolum cells that received the one-eighth RGD1.0 plus seven-eighths syn1.0 synthetic genomes grew, encouraging the team to try different combinations of RGD1.0 and syn1.0 genomes. In the end, they settled on a winning combination of fragments 2, 6, 7 and 8 from RGD1.0 matched with fragments 1, 3, 4 and 5 from syn1.0.
Syn2.0, as the resulting organism was named, had a total of 516 genes, less than the previous record holder M. genitalium’s 525. But Venter and the team didn’t stop there. Performing another design-build-test cycle, they were able to eliminate a further 43 genes, bringing down the number of genes in syn3.0 to 473.
Down the rabbit hole
Just like Deep Thought’s answer, however, Venter’s findings raise even more questions. First of all, more than 30 percent of the genes essential for syn3.0’s survival have no known biological function. Although the study authors speculate that many of them are likely to fall into one of the four major cell processes of gene expression, membrane structure and function, cytosolic metabolism or genome preservation, they also anticipate a number of genes to perform essential functions that are, as of now, a complete mystery.
More than anything, this study raises the question of what is meant by ‘life’ in the first place. Sure, 473 genes may be sufficient for parasitic bacteria that live in a nutrient rich environment, but which are the genes necessary to make more useful lifeforms, like an oil-slick eating or nitrogen-fixing bacteria? The definition of life, then, seems to be linked to what you want the life form to do.
This article is from a monthly column called From The Editor’s Desk(top). Click here to see the other articles in this series.
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Copyright: Asian Scientist Magazine; Photo: Tom Deerinck & Mark Ellisman, University of California.
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