Reversing 1.5 Billion Years of Evolution

I’m building a free-living chloroplast to dissect the origins of multicellular life.

This is Part 3 in our New Science research series, where our Fellows explain the visions behind their work. Two additional Fellow essays, not sent via email, are available to read on our website. This, our last post in the series, was written by Kian Faizi.

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“We are not made up, as we had always supposed, of successively enriched packets of our own parts. We are shared, rented, occupied… the colonial posterity of migrant prokaryocytes, probably primitive bacteria that swam into ancestral precursors of eukaryotic cells and stayed there…

I like to think that they work in my interest, that each breath they draw for me, but perhaps it is they who walk through the local park in the early morning, sensing my senses, listening to my music, thinking my thoughts.”
— Lewis Thomas, The Lives of a Cell

Plants weren’t the first to figure out photosynthesis, that careful dance of chemistry that harnesses sunlight to make our food and air. That honor belongs to microbes that first appeared 3.5 billion years ago, in the earliest known days of life on Earth. Some of them, called cyanobacteria, learned to split water into hydrogen and oxygen, and thus created our oxygen-rich atmosphere.

And then, about 1.5 billion years ago, something peculiar happened. A free-living cyanobacterium was engulfed by a larger eukaryotic cell in a phenomenon known as endosymbiosis. That bacterium escaped destruction and became a permanent resident — a cell living within another cell.

Soon, both parties realized they had stumbled upon a mutually beneficial arrangement. The larger, eukaryotic host siphoned food and energy from its photosynthetic guest; in exchange, it offered it shelter from the outside world. The cyanobacterium eventually lost its ability to live independently, handing off regulatory control to the eukaryote in a series of gene transfer events. It became a chloroplast: a specialized compartment that endowed the host with the ability to photosynthesize. All plants, in turn, descended from this single cell.

This transition from free-living cyanobacterium to integrated organelle was a landmark event in evolutionary history. An earlier event led to the creation of mitochondria.

Yet as momentous as endosymbioses are, the precise details of how they arise are still murky. There is much that we stand to learn by reverse engineering the process — but it won’t be easy. Scientific tools are limited, and deciphering this story, obscured by billions of years of complex coevolution, is akin to untying a Gordian knot.

I’m conducting a series of experiments aimed at reversing and reconstructing the chloroplast’s strange and fascinating history. The results could yield new insights into the evolution of multicellular life.

Modern endosymbiotic theory gained widespread acceptance in the 1980s, thanks to the work of Lynn Margulis, its principal champion and “vindicated heretic”. Her 1967 paper, On the origin of mitosing cells, weathered dozens of rejections before it was finally published. It’s now considered a watershed classic. 

Before Margulis, studies into the origins of chloroplasts and mitochondria were driven principally by the work of a few intrepid scientists who made careful observations — and persisted in the face of ridicule. The earliest known mention of endosymbiosis comes from an 1883 article by Andreas Schimper, a German botanist. In On the development of chlorophyll granules and colored bodies, he acknowledged the similarities between cyanobacteria and chloroplasts, before quietly adding a footnote:

“If it should definitively be confirmed that the plastids in egg cells [of plants] are not formed anew, then their relationship to the organism containing them would be somewhat reminiscent of a symbiosis.” (p. 112, footnote 2, emphasis added)

By 1905, the concept had been formalized in essays by the Russian botanist Konstantin Mereschkowski. In On the nature and origin of [plastids] in the plant kingdom, he outlines an endosymbiotic theory that comes astonishingly close to our modern insights. To his more influential contemporaries, though, his ideas were “too fantastic for present mention in polite biological society.” Ultimately, his work did not reach the mainstream — in part because it was overshadowed by his legacy as a serial rapist of children, who often evaded capture by fleeing to new countries.

Mereschkowski’s proposed tree of life, detailing two endosymbiotic events for the origins of nuclei and chloroplasts; the latter hypothesis is widely accepted today. (The theory of two plasma lineages as the foundation of symbiogenesis, a new principle for the origin of organisms, 1910).

American biologist Ivan Wallin argued for an endosymbiotic origin to mitochondria in the 1920s. To support his claim, he carried out a series of audacious experiments aimed at isolating and culturing mitochondria ex vivo. He claimed to have succeeded, but his results later proved irreproducible. Perhaps unsurprisingly, he too found little support among the scientific establishment of his day.

Things began to change as biologists developed new tools in the 1960s. Early genomic analyses crucially showed that chloroplasts contain their own DNA. Electron microscopy yielded the first detailed structural comparisons between chloroplasts and cyanobacteria. Unlike her predecessors, Margulis arguably prevailed thanks to this new experimental evidence.

Today, scientists have a greater palette of technologies to draw from than ever before. We can probe, modify, and dissect living things in ways that were previously impossible. Modern DNA editing, for example, allows us to engineer both the chloroplast and nuclear genomes as we desire; some labs are using these tools in attempts to recreate synthetic endosymbioses. 

All this gives rise to a daring question: Can we restore chloroplasts to a free-living state by re-endowing them with the genes that they’ve lost over 1.5 billion years of evolution? Even incremental progress towards this goal would yield important biological knowledge. As we work to wrestle back control over the organelle, we’ll uncover how the cell cycle, metabolism, and gene regulation are shaped by the interaction of host and endosymbiont. In liberating a chloroplast, we’ll be forced to unravel its complex evolutionary history using active experimentation and tinkering, rather than theoretical musings. To paraphrase Feynman, we should create a free-living chloroplast so that we may understand it.

During my New Science fellowship, I isolated chloroplasts from cells of the green alga Chlamydomonas reinhardtii using established methods. I’m carefully tweaking the chloroplasts’ environment to see which chemicals can support their metabolism in vitro. My first goal is to better understand how to make viable chloroplasts without any genetic engineering.

In the future, I intend to rewind evolutionary history by moving genes that were lost to the host over the last 1.5 billion years back into the chloroplast. In doing so, I aim to provide the first demonstration that chloroplast-derived, nuclear-encoded genes can be successfully repatriated without disrupting their function. I’m currently testing an existing method for inserting DNA into the chloroplast genome. Soon, I’ll compile a list of “high confidence” genes to be relocated. This may require the development of new computational approaches that can screen candidates by mining a combination of sequence data and the published literature.

These efforts will bolster our understanding of chloroplast function and phylogeny. In the future, they may even make it possible to engineer a new, free-living cell using an organelle as a starting point. What will this liberated chloroplast teach us — about its evolutionary history and the meaning of life itself? It’s time we find out.

Edited by Niko McCarty

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Thanks to Lev Tsypin, Adam Strandberg, John McCutcheon, Sasha Targ, and Alexey Guzey for feedback on this essay.

Cite this essay:

Faizi, K. “Reversing 1.5 Billion Years of Evolution.” newscience.org. 2022 September. https://doi.org/10.56416/720qud