‘Essentiality’ scan reveals microbe’s ‘must-have’ list

Researchers have spent years taking apart one of the world’s simplest microbes, Mycoplasma pneumoniae, piece by piece, and created a detailed list of what molecular parts the living cell can and cannot do without, knowledge that could accelerate the development of “living medicines” built from this very microbe. 

Their efforts have revealed how much real estate engineers have to edit and repurpose the bacterium for therapeutic purposes, for example to combat antibiotic resistance or cancer.  

The study, published today in Molecular Systems Biology, is the most comprehensive “essentiality map” for any living organism built to date. Researchers create the maps to see which bits of the genome living organisms truly need to stay alive, and how badly they falter when any given bit goes missing. 

Yet even for Escherichia coli, the most well-studied microbe on Earth, most surveys probe only one in every ten or twenty DNA positions, leaving wide blind spots and classifying genes in crude yes‑or‑no terms. The current study went far beyond by probing every other DNA letter, allowing researchers to rank genomic components not just as essential or non-essential, but along a continuum of importance. 

“We wanted to know which tiny parts of a bacterium’s DNA are truly essential, not just the obvious genes,” says Dr. Samuel MiravetVerde, a microbiologist at ETH Zurich who coled the work while at the Centre for Genomic Regulation (CRG) together with Dr. Raul Burgos and ICREA Research Professor Luis Serrano.  

Mycoplasma pneumoniae’s entire genome spans only about 816,000 DNA letters long, roughly a millionth the size of the human genome. The researchers used a technique called transposon sequencing to disrupt 450,000 of these DNA letters. 

Out of 707 proteincoding genes, 220 proved absolutely critical, 86 were almost critical, another 84 helped but weren’t essential, and 317 were dispensable under lab conditions. That means that roughly half of the microbe’s genetic instruction book can be deleted or damaged without killing it, at least in the lab conditions used. 

The study also examined the microbe’s regulatory elements. These are tiny regions of DNA that sit next to genes and tell the cell when, where or how strongly to use them. Out of the 1,050 regulatory elements studied, only 25 turned out to be truly necessary. That finding fits with what the researchers already suspected: the bacterium has very simple on/off switches and often runs its genes at full blast. Dr. Miravet-Verde likens the effort to checking not only the big gears in a machine, but also the small screws and switches that keep it running. 

Because every DNA letter now has a quantitative fitness score from “essential” to “dispensable”, the researchers can now predict how much a possible tweak to the microbe’s genome can slow growth or stress the cell, and choose the gentlest route to modify the bacterium instead of by costly trialanderror. 

The discovery matters because Mycoplasma pneumoniae is already being refitted as a therapeutic chassis for medical purposes. The research group, alongside biotechnology spin-off Pulmobiotics, have repurposed the bacterium and trained it to deliver drugs that can clear stubborn antibiotic-resistant lung infections in mice. They are also testing other variants to deliver anticancer drugs directly inside lung tumours. 

The new essentiality map gives the researchers more freedom to remove what they don’t need and a better roadmap to insert what they do, making M. pneumoniae one of the most engineerfriendly microbes available for the purposes of synthetic biology. 

“When you want to add a therapeutic payload, you must insert new DNA somewhere. If you land in the wrong spot, you can cripple a vital gene, so having this map confirm there are thousands of safe landing zones mapped at very high resolution gives us a very high degree of confidence,” explains Dr. Serrano. 

“It can also allow us to strip away surplus functions that reduce any chance the microbe might misbehave or survive where it shouldn’t, a key safety feature for ‘living medicines’,” adds Dr. Burgos. 

One of the surprising findings made by the study is that some “essential” genes could be split in two and the cell still lived. The discovery hints that some of the bacterium’s genes may be stitched together from smaller ancestors, offering a window on how life’s machinery evolved. “It’s like discovering a car can still drive even if part of the engine is cut in half,” says Dr. Miravet-Verde. 

The researchers next want to study why these essential genes can be split and the evolutionary implications of these events along the tree of life. “Were these once two separate proteins that fused over evolution?” asks Dr. Miravet Verde. “Answering that could help us design new, modular proteins for synthetic biology or understand how the proteins we see now in nature were originally formed.”