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Study reveals how bacteria build barriers

From the April 25, 2005,Princeton Weekly Bulletin.

Scientists at Princeton and Harvard universities have identified genes responsible for building a protective membrane around bacterial cells, a finding that helps solve a long-standing mystery and may lead to new antibiotic drugs.

A major class of bacteria, including E. coli, cholera and many others, guard themselves against antibiotics and other threats with a membrane of fat and proteins. Each E. coli bacterium, for example, is a single cell surrounded by a barrier, like a castle surrounded by an outer wall. Biologists have long puzzled over how bacteria build this outer membrane, because cells do not usually make structures outside their own bodies; the cell’s fuel and genetic instructions are inside the cell.

Researchers led by biologist Thomas Silhavy at Princeton and chemist Daniel Kahne at Harvard found a clever way to pinpoint the construction mechanism. They started with bacteria that were known to have defects in their outer membranes and subjected them to various antibiotics, which destroyed nearly all the bacteria. There were rare survivors, however, and the researchers knew these bacteria must have a special ability to repair their outer membranes.

Identifying the genes that made these cells different led to clues about how the membrane is built, including a network of five genes that appear to orchestrate the overall process. “If you could design a drug that would inhibit that process, then you could kill the bacteria,” said Silhavy.

The discovery also revealed a general laboratory technique for identifying other genes responsible for making major cell parts, known as organelles, which are surrounded by membranes, said Silhavy.

The researchers published their results in two separate papers in the April 22 issue of Cell. In addition to Silhavy and Kahne, authors of the papers are Juliana Malinverni, Natividad Ruiz and Brian Falcone at Princeton and Tao Wu and Seokhee Kim at Harvard.

“Many scientists study bacteria to better understand and to improve antibiotics, but this research does the opposite — it uses antibiotics to probe the inner workings of bacteria,” said Bert Shapiro, a cell biologist at the National Institute of General Medical Sciences, which supported the work. “A deeper understanding of the biochemical pathways in bacteria, such as that provided by this research, could in turn lead to new classes of antibiotics. Then the research will have come full circle.”

For Silhavy, who is Princeton’s Warner-Lambert Parke-Davis Professor of Molecular Biology, a deep understanding of the basic functioning of E. coli has been a longstanding goal. A member of the Princeton faculty since 1984, Silhavy has focused the work in his lab on how chemical signals are processed within bacteria and how proteins move and create structures in the cells. “The reason I do this work is because I just want to know how it works,” he said. “Once we understand the basic mechanisms — how the outer membrane is put together — useful, practical things will stem from that.”

For their latest findings, researchers in Silhavy’s lab started with a series of chemical compounds that killed the bacteria if they slipped through a leaky outer membrane. They discovered that each compound required the bacteria to come up with a slightly different set of membrane repairs in order to survive, allowing careful analysis of each repair mechanism. The researchers believe their strategy will help unlock the secrets of many cell compartments, because any chemical that harms a cell by slipping past a membrane is likely to elicit observable changes to the membrane as cells try to block the chemical.

Silhavy and colleagues are now looking to complete the picture of outer-membrane construction by finding the genes responsible for moving the machinery from the center of the cell to the outer membrane. “A very important piece of the puzzle is missing: the delivery men, as it were,” he said. “We will now try hard to find the delivery men.”

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