Issue 20 November 2012 - Max Planck Institute for Chemical Ecology

PULS/CE 20
Public Understanding of Life Sciences / Chemical Ecology
Newsletter November 2012
Elucidating Insecticide Resistance
Cotton bollworm larvae have evolved a novel enzyme from the group of so-called
P450 monooxygenases; the enzyme makes them resistant to pyrethroids.
The gene encoding the enzyme, a chimera, combines parts from two precursor
genes … p. 3
How Plants Make Cocaine
Scientists have discovered a key reaction in cocaine formation in the coca plant. The
enzyme they identified is shown to belong to the aldo-keto-reductase protein family.
The discovery of the first enzyme in the pathway sheds new light on the evolution of
alkaloid formation ... p. 4
Behavioral Assays in the “Flywalk”
Flies process attractive and deterrent odors in different brain areas.
Newly developed analytic device “Flywalk” allows accurate studies of
insect behavior to be made … p. 5
PULS/CE 20
2
Newsletter November 2012 | Editorial
Subterranean Research
Unit Starts Operation
Dear Readers!
Since its completion on the Beutenberg Campus
in 2001, the Max Planck Institute for Chemical
Ecology has consisted of two major building units:
the main building with offices and laboratories,
and the connected greenhouse complex on the
east side. Because of its artificial illumination,
the greenhouse can be seen from the opposite
hillside especially during the long and dark winter
months.
members of the department to investigate the
neurophysiology of insects and to monitor their
behavior under different experimental and replicatable environmental conditions.
Whether the institute will get further units in the
future is not clear yet. There is certainly no shortage of ideas. However, Beutenberg Campus is
running out of space, because the valley around
Jena, in which the prosperous town spreads in
such a picturesque way, has its natural limits: the
hills.
Wishing you a pleasantly cool as well as a sunny
winter!
Over the last few months, the main building has
received a new neighboring module on the west
side, which is now almost fully operative: the
“Schneiderhaus”. Unlike the greenhouse, this
new annex is almost invisible from the outside,
because it is completely subterranean. Around
500 m2 of floor space have been created underground. The building has been named after the
founder of modern olfactory physiology, Dietrich
Schneider (1919-2008).
The managing director of the institute, Bill Hansson, also head of the Department of Evolutionary
Neuroethology, is extremely satisfied. Two wind
tunnel facilities for studying the odor-regulated
behavior of moths, flies, and other insects have
recently commenced operation (see pictures), and
two so-called “Flywalk” systems have already
contributed to interesting results, presented as a
research highlight on page 5 in this issue. These
facilities, together with several growth chambers
for insect rearing and preparation rooms, allow
Jan-W. Kellmann
The new building has been named after Dietrich Schneider, the founder of modern olfactory physiology and thus a
pioneer in chemical ecology.
Left: The flight behavior of a moth is recorded in the
wind tunnel facility. Above: A fly is targeting the source
of an odor in the wind tunnel model. Graphic: Alexandra
Schmidt/MPI-CE.
PULS/CE
20
3
PULS/CE 203
Research Highlight | Newsletter November 2012
Solupta nusae volut abo.
Dolut omnimi, sunt
officimusant od eriamen
Original Publication:
Joußen, N., Agnolet, S., Lorenz, S.,
Schöne, S. E., Ellinger, R., Schneider,
B., Heckel, D. G. (2012). Resistance
of Australian Helicoverpa armigera
to fenvalerate is due to the chimeric
P450 enzyme CYP337B3. Proceedings
of the National Academy of Sciences
of the United States of America, 109,
15206-15211.
Elucidating Insecticide Resistance
Pyrethroids are synthetic substances based on
compounds of the natural insecticide pyrethrum.
They have been successfully applied in fruit, vegetable and crop farming for decades. In cotton
bollworms, however, since 1983 the development
of resistance to the particularly effective pyrethroid fenvalerate has been observed. In 1998,
David Heckel, who became head of the Department of Entomology in 2004, mapped the location of the resistance gene in the genome of the
bollworm.
Nicole Joußen, a scientist in David Heckel’s department, has studied the Helicoverpa armigera
strain “TWB”, known to be resistant to fenvalerate. She was able to identify the P450 monooxygenase which is mediating the pyrethroid
resistance in this strain by discovering that out
of seven P450 enzymes only one, CYP337B3,
hydroxylates the fenvalerate molecule to 4‘-hydroxyfenvalerate. This chemical reaction increases resistance to this toxin 42-fold: A LD50 value
of 1.9 µg of fenvalerate was measured in resistant TWB larvae, whereas half of the non-resistant caterpillars died after intake of only 0.04 µg
of the toxin. The CYP337B3 gene was formed in a
process geneticists call “unequal crossing-over”.
If similar DNA sequences, for example, transposable elements, come into contact with one another during the division of cell nuclei, novel gene
combinations occur. As a consequence, some
of the genetic information is lost on one DNA
strand, and new genetic information is inserted
and sometimes even doubled on the other strand.
This natural process is important for the evolution
of gene families, as Nicole Joußen observed in
the case of the CYP337B3 gene.
Above: Larvae of the cotton bollworm
(Helicoverpa armigera) are dreaded
pests all over the world. They have a
very wide host range: About 200 different plant species are known as potential food for the voracious insect. Nearly 30% of all globally used insecticides
− Bt toxins as well as pyrethroids − are
applied to protect cotton and other
crops against the bollworm.
Below: Nicole Joußen feeding cotton
bollworm moths.
For the first time, these results reveal an insecticide resistance mediating mutation caused
by a crossing-over event. The scientists studied
CYP337B3 and found that the gene consists of
parts of two other P450 genes, B1 and B2, encoding enzymes, neither of which can detoxify
fenvalerate. The unique combination of parts of
these precursor genes in the chimeric B3 gene is
responsible for the ability of CYP337B3 to bind,
hydroxylate, and finally detoxify the insecticide.
Resistance to insecticides is a natural event that
cannot be stopped. However, a careful and moderate use of pesticides together with other plant
protection measures, such as crop rotation, can
considerably slow the process. [AO/JWK]
Photos: Nicole Joußen, Angela Overmeyer, MPI -CE.
PULS/CE 20
4
Newsletter November 2012 | Research Highlight
How Plants Make Cocaine
Original Publication:
Jirschitzka, J., Schmidt, G., Reichelt,
M., Schneider, B., Gershenzon, J.,
D‘Auria, J. (2012). Plant tropane
alkaloid biosynthesis evolved
independently in the Solanaceae and
Erythroxylaceae. Proceedings of the
National Academy of Sciences of the
United States of America, 109(26),
10304-10309.
Native tribes in South America have been cultivating coca plants (Erythroxylum coca) and chewing its leaves for at least 8000 years for their
stimulant and hunger-suppressing properties.
Cocaine, which is accumulated in the coca leaves, is one of the most commonly used (and abused) plant-derived drugs in the world. However,
we have almost no modern information on how
plants produce this complex alkaloid. Alkaloids
constitute a very large group of natural nitrogencontaining compounds with diverse effects on
the human organism.
Right above: Coca plant (Erythroxylum
coca) and the molecular structure of
cocaine (grey: carbon, blue: nitrogen,
red: oxygen, white: hydrogen). Cocaine can account for up to 10% of the
dry weight of the immature coca leaf, a
phenomenal amount for the accumulation of any one particular alkaloid.
Below: The members of the project
group Biochemistry and Evolution of
Tropane Alkaloid Biosynthesis: Teresa
Docimo, Gregor Schmidt, John D’Auria,
Jan Jirschitzka. Photos: MPI-CE
Plant-produced alkaloids are used as toxins, stimulants, pharmaceuticals or recreational drugs,
including caffeine, nicotine, morphine, atropine
and cocaine. Atropine, used to dilate the pupils of
the eye, and the addictive drug cocaine are both
tropane alkaloids which possess two distinctive,
interconnecting five- and seven-membered rings.
Although the formation of cocaine has not been
investigated in the last 40 years, the biosynthesis of the related tropane alkaloid, atropine, from
belladonna is well established. In the penultimate step, a ketone is reduced to an alcohol
residue. This key reaction is catalyzed by an enzyme of the short-chain dehydrogenase/reductase
(SDR) protein family in belladonna. To find the corresponding enzyme in cocaine biosynthesis, Jan
Jirschitzka, a PhD student in the group of John
D’Auria, searched the genome of the coca plant
for SDR-like proteins. However, none of the SDR
genes that he cloned and expressed showed any
activity for the key reaction in cocaine formation.
So he used a more classical approach: he identified the cocaine-synthesizing enzyme activity in
extracts from coca leaves, purified the responsible protein, isolated the polypeptide, and, after
partially sequencing it, cloned the corresponding
gene. Surprisingly, the enzyme reaction analogous to that in atropine synthesis − the conversion of the keto group into an alcohol residue − is
catalyzed by a completely different enzyme in
coca plants than in belladonna: by the aldo-keto
reductase (AKR) named methylecgonone reductase (MecgoR). The second surprising result is
that the MecgoR gene, as well as the protein,
is highly active in the very young leaves of coca
plants but not in its roots. Atropine, on the other
hand, is synthesized exclusively in the roots of
belladonna, from where it is transported into the
plant‘s green organs. Based on these results, the
researchers conclude that the tropane alkaloid
pathways in coca and belladonna evolved completely independently. [AO/JWK]
PULS/CE
20
5
PULS/CE 205
Research Highlight | Newsletter November 2012
Behavioral Assays in the Flywalk
In collaboration with colleagues from Portugal
and Spain, researchers from the Department of
Evolutionary Neuroethology have developed an
apparatus that automatically applies odors to
an airstream, while simultaneously filming and
analyzing the behavior of insects. The system is
called the “Flywalk” and consists of glass tubes,
airstream regulators, and a video camera. The reactions of 15 flies to up to eight different odorant
signals can be tested at the same time. “Flywalk”
allows the use of not only single odorants but also
odor mixtures. In addition, odor pulses of varying
length and concentration can be given. Its high
throughput and the long automated measurement
periods − the insects can stay in the “Flywalk” tubes for up to eight hours − facilitate the statistical
analysis of the results.
Experiments with fruit flies demonstrated that
unlike males, females were more attracted by
typical food odors, such as ethyl acetate. This
behavior seems to reflect the search for the best
oviposition place in order to make sure that the
larval offspring will find sufficient food after hatching. Flies’ responses to deterrent odors, such as
benzaldehyde, were identical in both males and
females. Only males, however, responded positively to the odor of unmated females: If the odor
of virgin females was introduced into the “Flywalk” tubes, males moved upwind. Females that
have mated are surrounded by cis-vaccenyl acetate, a deterrent which renders them unattractive
to males willing to mate: a male marks the female
during copulation with cis-vaccenyl acetate to
prevent further fertilization by other males and
thus makes sure that his genes are spread.
In an additional study, scientists scrutinized the
tiny brains of fruit flies. Using specific activity
markers, they measured certain nerve cells, the
so-called projection neurons; these neurons are
located in the antennal lobe, the olfactory center
of flies. Experiments performed with six highly
attractive and six highly deterrent odors, selected out of 110 different and tested compounds,
revealed that attractive and deterrent odors are
processed in specific brain regions of the flies.
Drosophila fly in a glass Flywalk tube.
The newly developed analytic device is
used for accurate studies of insect behavior. Photo: Markus Knaden, MPI-CE
Original Publications:
Such specificity has also been shown in mice
and humans. The function of an insect brain thus
resembles that of a mammalian brain more than
previously thought. Because the activity of projection neurons reflects a kind of “interpretation”
of incoming odorant signals, the assessments
“good” or “bad” which ultimately regulate the
flies’ behavior seem to be represented early in
the insects’ brains. [AO/JWK]
Knaden, M., Strutz, A., Ahsan, J.,
Sachse, S., Hansson, B. (2012). Spatial
representation of odorant valence in an
insect brain. Cell Reports, 1, 392-399.
Steck, K., Veit, D., Grandy, R., Badia,
S. B. i., Mathews, Z., Verschure, P.,
Hansson, B., Knaden, M. (2012). A
high-throughput behavioral paradigm
for Drosophila olfaction - the Flywalk.
Nature Scientific Reports, 2: 361.
Two views inside the brain (here: antennal lobe) of a fruit fly while it is smelling; left: active glomeruli
(represented by bright colors) after stimulation with a deterrent odor (linalool); right: active glomeruli
after application of an attractant (3-methylthio-1-propanol). This shows that deterrent responses are
generated in lateral areas of the brain, whereas attractant responses are generated in medial areas.
Imaging: Antonia Strutz, MPI-CE.
PULS/CE 20
6
Newsletter November 2012 | IMPRS Project
The specialization of pea aphids is thought to
arise principally from their diet. Although phloem
sap generally contains amino acids and a high
concentration of sugars, a large range of secondary metabolites accumulates whose content
varies according to plant species. Some legume
secondary metabolites, for example saponins in
alfalfa and alkaloids in white lupine, strongly influence pea aphid performance. However, the role
of host plant chemistry on the specificity of pea
aphid host races has not been studied.
Pea aphids (Acyrthosiphon pisum) sucking the phloem sap from a pea stalk.
Photo: By courtesy of Jan-Peter Kasper.
Aphid Warning!
Although most of the insect herbivores under
study in our institute feed by chewing on plant
material, herbivores that suck plant fluids, such as
aphids, whiteflies, thrips and leafhoppers, are a
prominent group of pest insects that cause enormous damage to natural and cultivated plants.
Carlos Fernando Sánchez Arcos from
Colombia is a PhD student in the International Max Planck Research School
(IMPRS). For his dissertation in the
Department of Biochemistry (project
group of Grit Kunert), he studies the
defense chemistry of legumes and the
specificity of pea aphid host races.
Foto: MPI-CE
We are investigating the pea aphid (Acyrthosiphon pisum), an organism whose genome has
been fully sequenced, to understand the effects
of feeding specialization on different host plants.
Collectively, pea aphids feed on a range of host
plants in the legume family, but host races exist
that exhibit a clear preference for specific plants
such as alfalfa, red clover, or pea. This can be considered a first step towards speciation since the
host fidelity of races leads to assortative mating,
and assortative mating reduces gene flow among
host races.
The aim of my project is to identify the chemical compounds in the phloem sap of alfalfa, red
clover, pea and broad bean which determine the
pattern and the maintenance of host plant usage
by pea aphid host races. We use an untargeted
approach to determine the metabolic differences
among the phloem sap of these plant species as
well as the differences among honeydew metabolic profiles of four pea aphid host races reared
on each legume host. Then, we isolate and purify
the most relevant metabolites before evaluating
the response of the aphids to these metabolites
in artificial diets using the electrical penetration
graph (EPG) technique. EPG measures feeding behavior. The information obtained will allow us to
establish the impact of legume chemistry on the
pattern of host plant preference by pea aphids, as
well as the possible use of these metabolites as
new strategies to reduce the infestation of commercial crops.
Carlos Fernando Sánchez Arcos
PULS/CE 20
7
News | Newsletter November 2012
Field Study Published in the New Scientific Journal eLife Reveals
that Biological Pest Control Increases Plant Fitness
Researchers from the Department of Molecular
Ecology demonstrated that the release of volatiles to attract the enemies of herbivores, i.e.
predators of insect eggs and larvae, not only
controls insect pests but also increases the
reproduction of infested plants. For integrated
pest management practices, this means that
these natural plant defenses are able to increase
agricultural yields in an environmentally friendly
manner. These results were published as part of
the launch of the new open-access journal
eLife.
Original Publication:
Schuman, M., Barthel, K., Baldwin, I. T. (2012). Herbivory-induced volatiles function as defenses increasing
fitness of the native plant Nicotiana attenuata in nature.
eLife, 1: e00007.
A big-eyed bug (Geocoris spp.) is
attracted by green leaf volatiles and
starts sucking a tobacco hornworm
egg. By executing this kind of biocontrol, the tobacco plant rids itself of
herbivores. Copyright: Merit Motion
Pictures, Winnipeg, Manitoba, Canada
Humidity Increases Odor Perception in Terrestrial Hermit Crabs
Scientists from the Department of Evolutionary
Neuroethology found that the olfactory system
in hermit crabs is still underdeveloped compared
to that of vinegar flies. While flies have a very
sensitive sense of smell and are able to identify
various odor molecules in the air, crabs recognize
only a few odors. Humidity significantly enhances
both the electrical signals that are induced in the
crabs’ antennal neurons as well as the crabs’ corresponding behavioral responses to the odorants.
Exploring the molecular biology of olfaction in
land crabs and flies thus offers insights into the
evolution of the olfactory sense during the transition from life in water to life on land.
Original Publication:
Krång, A.-S., Knaden, M., Steck, K., Hansson, B. (2012).
Transition from sea to land: olfactory function and cons-
Coenobita clypeatus hermit crab using
traints in the terrestrial hermit crab Coenobita clypeatus.
a discarded snail shell for protection:
Proceedings of the Royal Society of London. Series B:
Both pairs of antennae are clearly
Biological Sciences. 279 (1742), 3510-3519.
visible. Photo: Katrin Groh, MPI-CE
Tobacco Plants Advertise Their Defensive Readiness to Attackers
Original Publication:
Kallenbach, M., Bonaventure, G., Gilar-
A few minutes after an herbivore attack, plants
produce jasmonic acid, a hormone which activates their defenses. The result is that toxic substances such as nicotine or digestion inhibitors
accumulate in the leaves. Scientists from the Department of Molecular Ecology have now found
that attacking leafhoppers can evaluate whether
Nicotiana attenuata plants are ready to activate
their defenses when attacked. If jasmonate-
signaling is immediately activated after attack to
the plants, leafhoppers desist from further feeding and start testing other plants. If this hormonal
signaling system is dysfunctional, the herbivores
start their attack. Hence, these leafhoppers can
actually be used by scientists as “bloodhounds”
in field experiments to locate plants hidden in
natural populations which are naturally defective
in their jasmonate signaling.
doni, P. A., Wissgott, A., Baldwin, I. T.
(2012). Empoasca leafhoppers attack
wild tobacco plants in a jasmonatedependent manner and identify jasmonate mutants in natural populations.
Proceedings of the National Academy
of Sciences of the United States of
America, 109(24), E1548-E1557.
PULS/CE 20
8
Newsletter November 2012 | News & Events
David G. Heckel on Insecticide Resistance 50 Years after
Carson’s Bestseller Silent Spring was Published
David Heckel Photo: Norbert Michalke
50 years after Rachel Carson published her famous book Silent Spring, which marked the birth
of the environmental movement, David G. Heckel
provides an overview in the journal Science on
the current state of resistances pest insects
have developed against insecticides. In the past
50 years more than 450 arthropod species have
become resistant to at least one pesticide. However, a paradigm shift in dealing with this global
problem has also occurred.
Original Publication:
Heckel, D. H. (2012). Insecticide resistance after Silent
Spring. Science, 337, 1612-1614.
Bill S. Hansson Elected Member of the Academia Europaea
Bill S. Hansson, director of the Department
of Evolutionary Neuroethology and currently
managing director of the MPI-CE, was elected
member of the Academia Europaea. This European scientific academy was founded in 1988
by scientists from the UK, France, Italy, Sweden,
Germany and the Netherlands as a panBill Hansson
European body that expresses the opinions
and ideas of scientists from across Europe.
The academy has more than 2000 members,
including over 40 Nobel Laureates.
http://www.acadeuro.org/
Photo: Norbert Michalke
Upcoming Events:
From October 19 until December 14, 2012, the Max Planck Institute for Chemical Ecology will host
an exhibit of watercolor paintings by Leipzig artist Hans-Joachim Wiesner, from his picture cycles
NATURE & FORM and ANALOGIES & REDUCTIONS. The exhibit is open to the public on Mondays,
12:00 - 04:00 p.m., and Tuesdays to Fridays, 1:00 - 3:00 p.m. With this art exhibition, the MPI is continuing a tradition which is on Beutenberg Campus associated with the name Hans Knöll: The head of the
former Central Institute for Microbiology and Experimental Therapy (ZIMET) initiated the ZIMET art
collection to forge a connection between art and science.
On Thursday, April 25, 2013, Beutenberg Campus Jena will organize the fourth Forsche-SchülerTag. The MPI for Chemical Ecology invites all school-kids from 10th grade up to a close encounter with
research on interactions between plants, insects and microbes, with the opportunity to perform experiments with our scientists in their labs. A detailed program will be available in February 2013.
© Danny Kessler
www.ice.mpg.de
Impressum: PULS/CE is published semi-annually and can be downloaded free of charge on the homepage of the MPI for Chemical Ecology and is distributed electronically as PDF to subscribers. A print version will be sent on request.
Editor: MPI-CE, Jena • Managing Director: Prof. Dr. Bill S. Hansson (viSdP). Editorial Staff: Dr. Jan-W. Kellmann,
Research Coordination • Angela Overmeyer, M.A., Information and Communication • Emily Wheeler, Editing
ISSN: 2191-7507 (Print), 2191-7639 (Online)