How the monarch butterfly evolved its resistance to toxic milkweed

The beloved black and orange wings of the monarch butterfly are more than just beautiful. They warn potential predators that this insect is poisonous to eat—a trait acquired from the butterfly feeding on the toxic milkweed plant. Now, two research teams have independently discovered how this iconic insect became immune to its poisonous diet, and they have shown how three genetic mutations are key—by making those same changes in a fruit fly.

“They are impressive pieces of research,” says Marcus Kronforst, an evolutionary biologist at the University of Chicago in Illinois who was not involved with either group. He notes that these studies are rare examples where researchers verified the mutations responsible for a trait by making them in another species.

Milkweed plants produce compounds called cardiac glycosides, which disrupt molecular pumps that control the proper flow of ions in and out of cells. Monarch butterflies and other consumers of the plant, however, have evolved versions of these pumps that leave the animals unaffected. To find what changes these milkweed eaters had in common, Noah Whiteman, an evolutionary biologist at the University of California, Berkeley, and his colleagues recently matched up the gene for this molecular pump in 21 insects, including monarchs, that tolerate the plant to varying degrees.

They found three mutations that changed three amino acids in the protein pump. By looking at the distribution of these changes across the insect family tree, Whiteman and colleagues were able to predict which ones came first—and it turned out that the order in which they evolved mattered. They then tried to replay the tape of evolution by making combinations of those changes in fruit flies using the gene-editing tool CRISPR.

A single mutation predicted to have arisen in the monarch’s ancestor made the fruit fly only a little resistant to the toxins in milkweed. But when that change was combined with a second one, the fruit fly was far better protected, Whiteman and his colleagues report today in Nature . And when they added the third mutation, the fruit fly thrived on milkweed as well as monarchs. The flies even retained some of the toxin in their bodies—as monarchs do to make themselves toxic to predators. “It’s just mind-blowing that the amazing ability of the butterfly to survive on these harsh chemicals comes down to just those three amino acids in the protein,” Kronforst says.

The findings confirm what evolutionary geneticist Peter Andolfatto of Columbia University and his colleagues reported 27 August in eLife when they engineered fruit flies with a different editing approach, sometimes making mutations in both copies of the pump’s gene and sometimes making them in only one copy. The order in which the mutations were introduced was also critical, Andolfatto notes. In one sequence, the mutations produced healthy, milkweed-tolerant insects. In others, the flies had neurological defects, he and his colleagues reported.

His team also studied what happened when the fruit fly had just a single copy of the ion pump gene with either one, two, or three mutations. The team found that just a single copy with the initial mutation confers some glycoside resistance, a property that may have enabled the change to persist long enough for other, more beneficial mutations to occur.

“Hopefully [this work] will serve as a reminder that the genomic context in which a mutation occurs is important,” says Hopi Hoekstra, an evolutionary biologist at Harvard University who was not involved with the work. All in all, “It’s just a beautifully complete story,” she adds.