Beans use an immune receptor to call in airstrikes on caterpillars

For decades, scientists have understood that plants can release volatile organic compounds—essentially airborne chemical signals—to attract the natural enemies of the things that eat them, like caterpillars. What we didn’t know was exactly how a plant translates the physical act of being eaten into a specific, predator-summoning distress signal.

“[One] thing we didn’t know is how the plant detects the caterpillar in the first place,” says Adam Steinbrenner, a biologist at the University of Washington. Now, after years of experimenting with common bean plants in the lab and in the agricultural fields of Oaxaca, Mexico, Steinbrenner’s team pinpointed a single immune receptor that orchestrates its anti-caterpillar defense system.

Drooling caterpillars

When an herbivorous insect like a caterpillar feeds on a plant, it introduces its saliva straight into the plant’s damaged tissues. This saliva contains biological clues called HAMPs: herbivore-associated molecular patterns. One of the HAMPs molecules is a peptide called inceptin, and there’s an 11-amino acid fragment of inceptin named In11, as well. Both of them turn out to be a fragment of the ATP synthase found in chloroplasts—basically a piece of one of the plant’s own proteins. As the caterpillar ingests the leaf, its gut enzymes chop up the plant’s cellular engines and their pieces, including In11, are regurgitated back onto the leaf’s surface, albeit at extremely small concentrations.

Over millions of years, plants like the common bean have evolved a specialized cell-surface receptor called the inceptin receptor just to detect In11. When this receptor interacts with In11, it sets off a signaling cascade in the plant’s cells, initiating immune responses. Proving that this specific receptor is responsible for releasing predator-summoning signals, though, was extremely tricky. “We were excited to do that, but we needed the perfect comparison plants—plants lacking the receptor versus ones that have the intact receptor,” Steinbrenner says.

The problem was that common bean plants are notoriously difficult to genetically modify, so the usual modern techniques like gene silencing were off the table. Picking an easier-to-modify plant was off the table, too. “We were sort of limited to bean because this receptor we were studying is only present in certain bean species,” Steinbrenner explains. To get around it, his team had to introduce the modifications they needed the old-fashioned way—through selective breeding.

Breeding siblings

The first step was to find a common bean plant with a muted In11 receptor. What the team needed was a natural mutant that was unable to detect the caterpillar’s saliva. They screened a massive panel of Mesoamerican beans, looking for varieties that failed to produce ethylene gas, a classic plant stress indicator, when exposed to In11. Out of 89 varieties tested, they found two that completely ignored the peptide. Of these two, they picked a Honduran strain called W6 13807.

When the researchers sequenced the genome of this insensitive bean, they found it had a naturally occurring 103-base-pair deletion in the gene that encodes the inceptin receptor. This mutation, they found, deletes a crucial chunk of the receptor, resulting in a truncated, non-functional protein.

To test the effect of this dysfunctional receptor on the plant’s defenses, the team began breeding the plants for their experiment. Through a series of genetic crosses and backcrosses between the mutant and a standard bean variant that was responsive to In11, they created sibling plants that were nearly identical genetically except for the presence or absence of the functional inceptin receptor. “We were just being breeders and that took several years”, Steinbrenner recalls.

When these two siblings were put side by side in the lab and in the field, it turned out the consequences of having a broken inceptin alarm were rather grave for the bean plants.

The cost of silence

First, the researchers examined direct defenses—the chemical and physical changes the plant undergoes to make its leaves less palatable for caterpillars and thus hamper their growth. When caterpillars fed on the mutant beans with inactive inceptin receptors, though, they had a field day. Over a five-day feeding period, their growth rate was over 70 percent higher than on the plants with a functional receptor.

More detailed analysis revealed exactly why this was the case. In plants that could detect the In11 peptide, a feeding caterpillar triggered the rapid up-regulation of 527 genes, including the ones responsible for anti-herbivore defenses. The plants that were oblivious to the In11 in the caterpillar spit failed to mount this targeted response. Instead, they reacted as if they were just being mechanically wounded by the wind or a passing animal. Without the receptor, they entirely missed that a live, hungry insect was actively eating them.

Another consequence for In11 insensitive beans was that they were unable to summon predatory wasps.

Calling air support

When a normal bean plant detects In11, it begins synthesizing and emitting a highly specific blend of volatile organic chemicals. To a predatory wasp, this blend of scents signals not just “a plant is damaged,” but specifically “a caterpillar is actively feeding here right now.” Lab tests showed that the plants without the active inceptin receptor failed to emit this volatile blend when exposed to either the synthetic In11 peptide or actual caterpillar oral secretions.

To see how much this lack of chemical signaling mattered in the wild, the researchers packed up their sibling bean lines and headed to an experimental agricultural field in Oaxaca, Mexico. There, they placed pairs of bean plants—one with the active receptor and one without it—out in the open. They treated the plants with either water, caterpillar oral secretions, or In11. Then, they attached live sentinel caterpillars to the leaves and sat back to watch what happened.

It turned out local predatory wasps were highly active in the field, but they weren’t searching randomly. Driven by the airborne chemical cues, the wasps disproportionately targeted the plants that had functional inceptin receptors. The plants treated with In11, or caterpillar spit were sending out their chemical distress signals into the wind, and the wasps were coming in to attack and remove the caterpillars in response to the call.