A MOLECULAR machine that can attach different-coloured glowing molecules to different types of damaged DNA is promising biologists a powerful new way to track how cells die.
Cell suicide, or apoptosis, is a crucial part of the developmental process in multicellular organisms. It is thought that many cancers start when apoptosis fails and cells refuse to die off as they should.
Early in apoptosis, enzymes shred a dying cell’s DNA into thousands of fragments. Looking for this damage is a common way to recognise apoptotic cells. But there is a complication: the DNA can be shredded in two ways, and to understand apoptosis properly, biologists need to be able to tell in which way it was cut up. One way leaves a phosphate group on the so-called five-prime (5) end, while the other leaves the 5 end carrying a hydroxyl group. Standard lab tests can only detect the phosphate-loaded cut; detecting the less common hydroxyl-carrying break is much tougher.
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Now Vladimir Didenko and his colleagues at Baylor College of Medicine in Houston, Texas, and at the Sloan-Kettering Institute in New York have developed a convenient way to attach different fluorescent labels to each type of DNA fracture. Their method marries a natural DNA-rearranging enzyme called topoisomerase with a synthetic single-stranded DNA fragment to which they attach two fluorescent molecules: red on one end and green on the other.
In the presence of damaged DNA, the topoisomerase cleaves the synthetic DNA strand and attaches the green-labelled half to a 5′ hydroxyl end. The remaining synthetic DNA fragment, with its red label, can then be attached to any 5′ phosphate breaks in the sample using a separate enzyme, called DNA ligase.
When Didenko’s team tested the detector on apoptotic tissue samples from the thymus gland of a rat, they found both types of DNA damage were present. And because, unlike existing techniques, the detector puts only one fluorescent label on each break, the intensity of the fluorescent signal is directly proportional to the number of breaks, making them easy to count. The technique makes it possible to track DNA degradation during apoptosis very precisely, Didenko says.
Harnessing natural molecules, which have evolved over billions of years, may be the way to make nanotechnology a reality, say the researchers. They dub their detector a “semi-artificial machine” (Nano Letters, DOI: 10.1021/n1048357e).
Thomas Schneider, a molecular biologist at the National Cancer Institute in Frederick, Maryland, calls it “a cyborg-like device”. Schneider is developing a molecular rotor based on myosin, a protein responsible for muscle contraction. Like Didenko he believes that using biological molecules can avoid some of the problems that tend to trip up nano-engineers when they try to scale down larger machines, such as friction and viscosity.
“Using biological molecules can avoid problems that tend to trip up nano-engineers when they try to scale down machines”
Letting nature do most of the work is the best approach, Schneider says. “It’s easier to steal things from nature. This thing already has its precision pre-built for you,” he adds. “Maybe this is the beginning of some spectacular new technology. I can’t tell you – it’s way too early to know.”
