A research team from the Nanjing Agricultural University in Nanjing, China, and the Max Planck Institute for Chemical Ecology in Jena, Germany, showed that isothiocyanates produced by cruciferous plants to fend off pests serve as oviposition cues. The plant defense substances serve as odor signals for females of the diamondback moth to lay their eggs on these plants. The scientists identified two olfactory receptors whose sole function is to detect these defense substances and to guide the moths to the ideal oviposition sites. They uncovered the molecular mechanism that explains why some insects that specialize in feeding on certain host plants are attracted by substances that are supposed to keep pests away (Current Biology, DOI: 10.1016/j.cub.2020.08.047, September 2020).
From repellent to attractant
Cruciferous
plants, such as cabbage, rape, mustard and horseradish, produce
glucosinolates. Upon mechanical damage of the plant tissues, e.g. caused
by a chewing insect, glucosinolates are hydrolyzed by the endogenous
plant enzyme myrosinase. This leads to the formation of a variety of
toxic breakdown products, mainly isothiocyanates, to defend themselves
against voracious insects. This defense mechanism is very effective
against most herbivores. The diamondback moth Plutella xylostella,
however, has evolved mechanisms of its own to outwit this defense: It
is able to feed successfully on plants of the cabbage family and make
use of the plants for its own reproductive purposes.
“We wanted
to know whether the moths use isothiocyanates as odor cues to locate
their host plants. In fact, behavioral experiments showed that three
isothiocyanates are key signals for female moths to locate and lay eggs
on cruciferous plants,” says study leader Shuang-Lin Dong from Nanjing Agricultural University.
Two olfactory receptors specialized on isothiocyanates control egg-laying
The main scientific question was, what are the molecular mechanisms on which female Plutella xylostella
moths base their choice of the oviposition site? The researchers
therefore analyzed, which olfactory receptors were highly expressed in
female moths, and studied the function of these receptors in the frog
oocytes. “With this method, we were able to investigate which odors an
individual receptor was responding to. We showed that two receptors,
OR35 and OR49, responded to the three isothiocyanates that we had
previously identified as being crucial for oviposition,” says Markus
Knaden from the Department of Evolutionary Neuroethology at the Max Planck Institute in Jena.
These
two receptors did not respond to any other plant-related odors or to
the sex pheromones of the moths. Presumably, OR35 and OR49 evolved to
detect precisely those egg-laying signals. “We were surprised that even
two receptors are specifically tuned to the isothiocyanates. The two
receptors, however, detect the isothiocyanates with different
sensitivities. We hypothesize that the more sensitive receptor could
make sure that female moths locate plants from a distance, while the
other may help to provide a more accurate detection of the
isothiocyanate concentration. This will give the female moths more
information about the substrate on which they will lay their eggs,” says
Shuang-Lin Dong.
Validation of gene function using CRISPR-Cas9 gene knockout techniques
The
researchers used the CRISPR-Cas9 genetic scissors to knock out the
genes encoding the two receptors in moths. This method is used to test
the function of a specific gene. For egg-laying assays, they used plants
of the thale cress Arabidopsis thaliana, a model plant that
belongs to the cruciferous plant family. Some of these plants were
unmodified and produced isothiocyanates that were attractive to the
moths, whereas the others were mutants that were unable to produce
isothiocyanates. When one of the two receptors was inactivated, the
moths laid considerably fewer eggs on the isothiocyanates-emitting
plants. When both receptors were knocked out, the moths were unable to
discriminate between unmodified Arabidopsis plants and the mutant
plants.
Cheaters in plant-insect interactions
In
the course of evolution, plants have developed various strategies to
defend themselves against herbivores. A crucial part of plant-insect
interaction is chemical communication. “In most cases, it is useful for a
plant to communicate to potential herbivores that its defense system is
already activated. However, there will be always someone who misuses
the communication for its own benefit, like in our case the diamondback
moth, which uses a plant defense signal as an attractant and lays eggs
and spreads on this plant,” says Markus Knaden. Finding out how these
“cheaters” outwit plant defenses and even use these defenses for their
own purposes could help improve the control of global crop pests (such
as the diamondback moth): “Our results offer various approaches to
control this pest: On the one hand, we could use the identified
isothiocyanates or other attractive substances as attractants to trap
these pests. On the other hand, we could try to develop chemical agents
to interrupt or block the perception of the isothiocyanates and thus
interfere with the females’ location of their host plants,” summarizes
Shuang-Lin Dong.
Further investigations are planned to study whether
other insects that attack cruciferous plants also use special receptors
to detect isothiocyanates and to locate the plants for oviposition. The
results may provide information on the extent to which the perception
of these odors by specialized receptors is also conserved in other