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'Genetic network' guards against lethal DNA damage

lled synthetic fitness or lethality interactions. Synthetic lethality is a phenomenon in which two mutations that are not individually lethal cause cell death when combined. Specifically, the Hopkins team identified 4,956 interactions among 875 genes involved in DNA repair, DNA replication, the halting of replication and cell cycle progression by "checkpoints" so that damaged DNA can undergo repair, and responses to oxidative stress necessary for reducing the intracellular levels of highly reactive molecules that bind to and damage DNA.

The yeast has about 6,000 genes, of which about 1,000 are essential to survival and 5,000 are not, Boeke said. Specifically, 1,000 of the 5,000 non-essential genes are important enough so that the yeast grows slowly if any one of them is absent. And any of the 4,000 other genes can be deleted from the cell without interfering with the cell's growth.

A major goal of the Hopkins team is to determine which of the non-essential genes interact with each other, said Boeke. All such pair-wise combinations of the 5,000 non-essential genes in the yeast genome would require about 25 million tests, he added. In the current study, 74 genes were tested in pair-wise combination with the 5,000 non-essential genes, a feat approximately equivalent to 370,000 gene-pair tests.

The Hopkins team used a technology known as dSLAM (heterozygote diploid-based synthetic lethality analyzed by microarray) to look at the effects of 5,000 different double mutations on cell fitness in a single experiment. With this technology, only 5,000 tests would be required to map the 25 million pair-wise combinations, greatly speeding the work.

The dSLAM strategy is somewhat like pulling out parts of a radio at random to see what happens, Boeke said.

"With yeast, as with a radio, you might rip out part A or part B and find that the radio still works; but if you pull out both parts and the radio dies you would learn that A and B can comp
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Source:Johns Hopkins Medical Institutions


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