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Electron micrograph of Escherichia coli (Image via Wikipedia)A recent paper describes how the mutation of a single gene is sufficient to turn a harmless bacterium found in our gut into an invasive pathogen. Taken alone, this isn’t terribly surprising; many genes regulate the expression of other genes and some (“master” genes) act as switches that control a whole host of other genes.  The gene mutated in this study isn’t one of these “master” genes, though; it’s a structural gene and there’s a much more  intriguing reason for its broad impact on the bacterium.

Escherichia coli is a rod-shaped bacterium often found in the intestines of warm-blooded animals, including humans. They are often harmless (or even useful — our gut bacteria are important!), but some types (“strains”) may cause illness or food-poisoning. For example, the strain E. coli O104:H4 was responsible for the food-poisonings (traced back to German sprouts) in May of last year.

E. coli K-12 is a strain isolated from the human gut in 1922. In the 1970s, it became the workhorse for molecular biology because of its noninvasive, extracellular, benign commensal nature — in other words, because it’s a harmless bug that lives in our gut but doesn’t get into our cells. We know that under a wide range of conditions and in various animals, K-12 doesn’t invade its host’s cells or behave as a pathogen; in fact, the lab strain has even lost the ability to thrive in the human gut. These characteristics are the reason that the pioneers of the genetic age selected K-12 for their work. It became widely used throughout molecular genetics research and is still used in many labs today (along with derivative strains). In 1997, it was one of the first organisms to have its genome sequenced. As a result of being at the center of molecular biology for nearly 40 years, it may well be the best characterized life form on the planet. Although K-12 isn’t normally virulent, there are virulence genes in its genome, a fact which has puzzled scientists.  We generally expect unused genes to disappear or acquire many mutations over time, but neither of those seems to be the case here.

In a study published this September in mBio, Koli and her colleagues found that the mutation of a single gene was enough to reactivate these virulence genes, transforming K-12 into a very different kind of bacterium. Not only did it gain the ability to invade cells, but a whole suite of genes was simultaneously activated.  Some of the genes are clearly virulence-related, enabling it to replicate inside host cells and decreasing the production of a protein that would prematurely kill host cells; others have more general functions, leading to a change in the bacteria’s shape (from rod-like to spherical) and its metabolism, as well as giving it a wider temperature tolerance. The fact that the mutation of a single gene can affect such a range of traits isn’t surprising.  The gene could be for a “transcription factor”, which is a molecule that bind to DNA and activate other genes; it’s not inconceivable that a mutant form of a transcription factor could activate a whole slew virulence genes in K-12. That’s not what’s happening, though, which is what makes this story so interesting. Instead, the mutation is in a gene encoding a structural protein that forms a kind of scaffolding for DNA.

A surprising fact about DNA is how very long the molecule actually is. Each of your cells contains about 1.8m (nearly 6 ft) of DNA coiled up inside it. At 1.5mm (a bit less than 1/16 of an inch) long, the DNA inside an E. coli is much shorter, but it’s still much larger than the E. coli cell, which is only about 0.002mm long. DNA has to be intricately packaged in order to fit inside a cell. In our cells (and the cells of other animals, plants and fungi), this is accomplished by winding the DNA around proteins called “histones”; a good analogy is the way a string (DNA) is wound around a spool (the histone). There are actually several different histones which interact to form units called nucleosomes, which then also form a helix, packing the DNA even more tightly; I managed to find a pretty good graphical representation for those who are more visually inclined. Bacteria don’t have histones, but they have “histone-like” proteins which perform similar roles; in E. coli, the histone-like proteins are HUa and HUb. The mutation that transformed the harmless E. coli K-12 line into an invasive form was in the gene encoding its HUa protein.

In addition to being wrapped around histones (or histone-like proteins), DNA is further compacted by something called “supercoiling”. Supercoiling is basically a description of the shape a DNA molecule takes when it is wound more tightly or more loosely. In a relaxed DNA molecule, there are a certain number of base pairs for every turn around the helix; changes in this number result in a physical strain on the molecule, which then changes its shape accordingly. A good analogy is the way a wire (like a headphone cable) or a rubber band twirls itself into complicated shapes if you twist it; a really good example is the way the handset wires on old telephones would twist around themselves. Depending on whether the DNA molecule is over- or under-wound, the supercoil spiral (“superhelix”) will twist one way or another; overwound DNA is positively supercoiled while underwound DNA is negatively supercoiled. The mutant histone protein in the invasive strain of E. coli K-12 causes the DNA to be more tightly wound, changing the superhelix from negative to positive supercoiling. The researchers were able to show that it was this change which resulted in the altered gene expression in the novel strain, dramatically altering the aspect and life-history of K-12.

The mutation of a single structural protein in the normally harmless K-12 strain of E. coli results in a conformational change of its entire DNA, exposing many genes and allowing them to become active (while the activity of other genes is repressed). We know that histones play a role in regulating gene expression in general, but this is a particularly striking example because of the dramatic nature of the change — it’s not just that a few genes were up- or down-regulated, but key aspects of the bacterium’s interaction with other cells were changed, as well as the shape and metabolism of the bacterium itself. It’s pretty intriguing to imagine what kind of role this sort of regulation could play, both in the short-term (e.g., stress response) and over the course of evolution. The fact that the up- and down-regulated genes in this example seem to be functionally co-ordinated suggests that this could be a very useful mechanism for efficient co-regulation of suites of functionally related genes, which could be selected together for activation (or repression) in positively or negatively supercoiled DNA. It’s also a wonderful example of the importance of form at every level of biology and a reminder that physical and spatial factors can have important biological consequences.  I think it’s a beautiful story which teaches us something new while showing of how much we still have to discover.

Ref: Koli P, Sudan S, Fitzgerald D, Adhya S & Kar S. (2011) Koli, P., Sudan, S., Fitzgerald, D., Adhya, S., & Kar, S. (2011). Conversion of Commensal Escherichia coli K-12 to an Invasive Form via Expression of a Mutant Histone-Like Protein mBio, 2 (5) DOI: 10.1128/mBio.00182-11
(The paper is open access, which means you can read it without a subscription.)

[I decided to write about this paper after hearing about it on episode 18 of the podcast This Week in Microbiology, which is an excellent weekly podcast about the world of the very small.  If you’re interested in that sort of thing, give TWiM a try!]