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Small mutation changes brain freeze to hot foot

cule that functions as a sensor of pain."

In a study appearing online early May 8 in the journal Neuron, Grandl's team focused on TRPA1, an ion channel best known as a sensor for pain caused by environmental irritants and pungent chemicals, such as mustard oil, the active compound found in wasabi.

Grandl's colleagues, postdoctoral fellow Sairam Jabba and research technician Raman Goyal, investigated whether single-point mutations could change cold-activated mouse TRPA1 into heat-activated. They formed this hypothesis because, in some other animals, including Drosophila fruit-flies and rattlesnakes, TRPA1 is naturally heat-activated.

To identify these structures, the team created a library of 12,000 mutant clones of the cold-activated mouse TRPA1 ion channel and randomly inserted one or two point mutations into each clone. After placing single clones into the individual slots of a 384-well plate and heating it from 25 degrees Celsius to 45 C in a matter of seconds, they were able to measure the thermal sensitivity of each mutant protein.

This screening pinpointed seven clones that showed strong activation when exposed to heat. Gene sequencing of these clones revealed 12 mutations that could potentially be responsible for changing the mouse TRPA1 from cold-activated to heat-activated. Out of these 12 mutations, Jabba and Goyal identified three mutations powerful enough to individually make that switch in TRPA1.

The mutations all turned out to be located within a single small domain of the ion channel protein known as ankyrin repeat six, indicating this domain plays a role in determining cold or heat activation. Ankyrin repeats are often responsible for managing protein-to-protein interactions, but their precise function in TRPA1 had not been previously known.

Interestingly, these single-point mutations didn't change the ion channels' responses to chemicals, such as mustard oil.

"This was

Contact: Karl Bates
Duke University

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