The contoured landscape of the fingertip, the aged face, the shriveled fruit: These are the living manifestations of wrinkling. By examining bacterial communities known as biofilms, researchers may have gained insight into how these sometimes beautiful, sometimes bothersome structures arise. In a new study, the scientists showed that they can control the self-assembly of biofilms, and the death of cells within them, to form specific designs—work that may yield a useful technique for tissue engineering.
Biofilms form when bacteria grow on a surface and adhere together. The microbes produce a scaffolding made of proteins, starches, and other molecules; this scaffolding holds the cells in place as they divide, until gradually a visible glob appears. The biofilm can develop various features, such as ridges, formed by folds in the scaffolding. These features can be impressive: If the bacteria were human-sized, the ridges would be as tall as New York City's Chrysler Building.
Systems biologist Gürol Süel at the University of California, San Diego, wanted to understand what generates the ridges, which are thought to help the biofilm exchange nutrients more efficiently. Süel and his colleagues found dead cells accumulated under the ridges, but didn't know which came first: the cells' deaths or the folding of the scaffolding. To solve this chicken-or-egg problem, Süel and his colleagues first set out to explore cell death in the biofilms. The team made many mutants of Bacillus subtilis, a soil bacterium. Each mutant was missing a gene thought to be involved in biofilm formation. In each case, the team noted whether the gene enhanced or inhibited cell death. The team also kept track of when and where the cells died. Overall, there was a pattern to the death of the cells, they discovered: Only certain patches of cells would disintegrate.
And after those cell patches disintegrated, wrinkles appeared at those locations, Süel and his colleagues report this week in the early online edition of the Proceedings of the National Academy of Sciences. And when mutant cells did not die as often as normal, they found, the biofilm wrinkled less.
The group then measured the mechanical properties of the biofilm and also looked at how the mutant cells and their scaffolding moved as the biofilm grew. Genes that altered the scaffolding's stiffness affected how well the wrinkles formed, they found. This shows that "there's an interplay between the genetic processes and the physical aspects," says Jim Haseloff, a synthetic biologist at the University of Cambridge in United Kingdom who was not involved with the study.
"For the first time, this paper suggests a mechanism for building a structure" that involves both cell death and mechanical forces, says Ken Bayles, a microbiologist at the University of Nebraska Medical Center in Omaha who was not involved with the work. "It provides a very basic model to help guide us to understand much more complex processes."
Furthermore, says Gerard Markx, a biochemical engineer at Heriot-Watt University in the United Kingdom who was not involved with the work, the study "gives us insights into how developmental processes may work in larger multicellular organisms such as plants and animals."
Süel thinks overcrowding within a biofilm leads to the cells' demise. The scaffolding constrains the cells as they grow, and they get crammed together, much like riders on the Tokyo subway are pushed into already crowded cars. Oxygen gets depleted and wastes accumulate, triggering cell death. Meanwhile a lot of tension builds up in the scaffolding due to lateral compression. When the cells shrivel, they relieve that tension and the biofilm buckles vertically, ultimately creating a wrinkle.
By laying down cells in different densities on a laboratory dish, the researchers discovered they could make wrinkles grow in specific patterns, such as the letters of the alphabet. The technique may be useful as chemical engineers try to harness biofilms for processing biofuels and other valuable chemicals. "It's one of the few demonstrations that we can control a three-dimensional structure [emerging] from a population of living cells," Süel says.