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OCR for page 87
Colloquium
Molecular genetics and evolution of pheromone
biosynthesis in Lepidoptera
Wendell L. Roelofs*t and Alejandro P. Rooney'
*New York State Agricultural Experiment Station, Cornell University, Geneva, NY 14456; and "National Center for Agricultural Utilization Research,
Agricultural Research Service, U.S. Department of Agriculture, Peoria, IL 61604
Contributed byWendell L. Roelofs, June 18, 2003
A great diversity of pheromone structures are used by moth species
(Insecta: Lepidoptera) for long-distance mating signals. The signal/
response channel seems to be narrow for each species, and a major
conundrum is how signal divergence has occurred in the face of
strong selection pressures against small changes in the signal.
Observations of various closely related and morphologically similar
species that use pheromone components biosynthesized by dif-
ferent enzymes and biosynthetic routes underscore the question as
to how major jumps in the biosynthetic routes could have evolved
with a mate recognition system that is based on responses to a
specific blend of chemicals. Research on the desaturases used in the
pheromone biosynthetic pathway for various moth species has
revealed that one way to make a major shift in the pheromone
blend is by activation of a different desaturase from mRNA that
already exists in the pheromone gland. Data will be presented to
support the hypothesis that this process was used in the evolution
of the Asian corn borer, Ostrinia furnacalis species. In that context,
moth sex-pheromone desaturase genes seem to be evolving under
a birth-and-death process. According to this model of multigene
family evolution, some genes are maintained in the genome for
long periods of time, whereas others become deleted or lose their
functionality, and new genes are created through gene duplica-
tion. This mode of evolution seems to play a role in moth specia-
tion, as exemplified by the case of the Asian corn borer and
European corn borer, Ostrinia nubilalis species.
Chemical communication systems in insects have provided ex-
citing challenges to researchers in chemistry, biochemistry,
physiology, ecology, genetics, and behavior for over four decades.
Much of this research has been focused on moths in the order
Lepidoptera, which is the second largest insect order with well over
a hundred thousand described species. Most of the hundreds of
species studied have been found to use a long-distance chemical
communication system for attracting mates (www.nysaes.
cornell.edu/fst/faculty/acree/pheronet/index.html). Initially,
pheromone components were characterized, and behavior medi-
ated by these chemical cues was studied. However, increased
knowledge of the precise blends used by different species only
raised more questions on many aspects of the communication
system. Underlying these questions was the fundamental curiosity
about the extensive radiation seen in moths and what role phero-
mones played in the speciation process. How did sex pheromones
evolve and could changes in this mating system give rise to isolated
populations that become new species? Studies on pheromone
biosynthetic pathways provided basic information on this issue, but
the use of molecular techniques in the postgenomics era has
become essential in addressing these questions.
Pheromone Biosynthetic Pathways
By the 1980s, maIly moth pheromone components had been
characterized, and a majority were acetates, alcohols, or alde-
www.pnas.org/cgi/doi/ ~ 0. ~ 073/pnas.1233767 ~ 00
hydes with long hydrocarbon chains (10-18C) containing 1-3
double bonds with variable positions and geometric configura-
tions (1~. Studies with labeled precursors were carried out to
determine whether the pheromone components were produced
via fatty acid precursors and what regulated the chain length and
double bond positions. Data on the red-banded leafroller moth,
Argyrotuenia velutinana, showed that the two major pheromone
components, (Z)-1 1-tetradecenyl acetate (Z1 1-14:0Ac) and
(E)-11-tetradecenyl acetate (E11-14:0Ac) were produced with
/~11 desaturation of myristic acid (14:Acid), followed by reduc-
tion and acetylation (2~. Data on the cabbage looper, Trichop-
lusia ni, showed that the main pheromone component, (Z)-7-
dodecenyl acetate (Z7-12:0Ac), was produced by 1~11
desaturation of palmitic acid (16:Acid) followed by chain short-
ening to Z9-14:Acid and then to Z7-12:Acid, with subsequent
reduction and acetylation (3~.
As biosynthetic pathways were defined for more moth species,
it became obvious that biochemical pathways ~nvolving two key
enzymatic steps, desaturation and chain shortening, had evolved
in the terminal segments (pheromone gland) of female moths.
The limited chain-shortening steps added diversity by generating
cascades of compounds of different chain lengths with double
bond positions two carbons closer to the functional group with
each round of chain shortening. It was soon found, however, that
the integral membrane desaturases were able to add d~versity by
evolving unique substrate specificities, as well as regio- and
stereospecificities. Thus, although many species were found to
use rare Z\11 desaturases that produced Z11-16:Acid, mixtures
of Z11/E11-14:Acid, or only E11-14:Acid, other species were
found to use different desaturases. Primitive moth species in
New Zealand are particularly interesting because the complex of
morphologically similar spec~es in the Planotortrix and Ctenop-
seutis genera produce pheromone components that involve at
least /~5, A7, A9, and All desaturases, either alone or in
combination (4~. How did these closely related species evolve
such diverse desaturases, and how was it possible to mutate from
one strongly stabilized pheromone system to another that used
pheromone components with different double bond positions?
Desaturase Genes
The biosynthetic enzymes in the moth pheromone glands could
have evolved from genes involved in normal fatty acid metab-
olism, but it was soon recognized (5) that the resolution powers
This paper results from the Arthur M. Sackier Colioquium of the Nationai Academy of
Sciences, "Chemical Communication in a Post-Genomic Worid," held January 17-19, 2003,
at the Arnold and Mabel Beckman Center of the National Academies of Science and
Engineering in Irvine, CA.
Abbreviations: ACB, Asian corn borer; ECB, European corn borer; Z14-16:Acid, (Z)-14-
hexadecenoic acid tfa~y acids and acetates of other configurations, chain lengths, and
position of unsaturation are named similarly).
tTo whom correspondence should be addressecI. E-mail: WLR1@cornell.edu.
PNAS 1 August 5, 2003 1 vol. 100 1 no. 16 1 9179-9184
OCR for page 88
of data on biosynthetic pathways for phylogenetic reconstruction
are limited and do not necessarily impart evidence of evolution-
ary direction. A move to the molecular level was dictated in an
attempt to determine homologies among the metabolic /~9
desaturases and sex-pheromone desaturases.
A collaborative effort in the early l990s was initiated on the
gene that encodes a /~11 desaturase in the sex-pheromone gland
of female T. nit Pheromone mRNA was isolated and reverse
transcribed to make cDNA, which was used as template in PCR
reactions with degenerate primers designed from conserved
areas of rat and mouse acyl-CoA /~9 desaturases (6, 7~. Candi-
date desaturase clones were obtained but had to be assayed for
functionality. This proved to be difficult with an in vitro recon-
stitutive biochemical assay because integral membrane desatu-
rases were shown to use a complex consisting of the desaturase
protein, NADH-cytochrome b5 reductase (a flavoprotein), and
cytochrome b5 (a hemoprotein). However, a discovery that the
yeast OLE1 desaturase can be functionally replaced in viva by the
rat desaturase (7) led the way to the development of yeast
expression systems that could be used for the functional assay of
pheromone desaturases (8~.
The structures of 1~9 and All desaturase-encoding cDNAs
from T. ni and the corn earworm moth, Helicoverpa zea, were
first characterized and reported (8-10~. Research on other
selected species resulted in the characterization of structures of
other pheromone desaturases with different regio- and ste-
reospecificities, including Z/E11, E11, Z10, Z9, and Z/E14
desaturases (8-15~. Cloning studies using mRNA isolated from
fat bodies and pheromone glands revealed that there were at
least two classes of /~9 desaturases in the Lepidoptera. One
produced a mixture containing palmitoleic acid (Z9-16:Acid) >
oleic acid (Z9-18:Acid), and the other with a reverse ratio of
Z9-18 > Z9-16. It also became obvious that there were
desaturase genes present in the pheromone gland that were not
functioning to produce unsaturated fatty acid product and some
desaturase clones that showed no activity in any of the available
functional assays. These results provided some insights into how
this multigene family evolved and how these genes could play a
role in the speciation process.
Corn Borer Speciation
Research on the European corn borer (ECB), Ostrinia nubilalis,
has shown that pheromone production (female) and response
(male) are not genetically linked (16), which provides the basis
for models or the evolution of new pheromones based on
asymmetric tracking (17, 18) and includes examples wherein a
large mutational effect in female pheromone production is
subsequently tracked by male response. Desaturase cloning
studies with ECB surprisingly revealed a way that the large
change in female pheromone production could be effected. The
studies (15) showed that the ECB pheromone gland contained
mRNA for three different desaturases, but unsaturated products
were only present in the gland from one of them, the All that
makes precursors for the pheromone components, Z11-/E11-
14:0Acs (19) (Fig. 1~. Another one was a /\9 desaturase, which
is commonly present in pheromone glands, and the third was
found to be a A14 desaturase. The presence of the A 14 desaturase
is significant because it is the desaturase used by the Asian corn
borer (ACB), O. furnacalis (20), to produce its unusual mixture
of Z/E12-14:0Acs pheromone components (21) (Fig. 1~.
The ACB is the only Ostrinia species in the world known to use
the A14 desaturase, with all others using a 1~11 desaturase (22~.
Research on mRNA from ACB pheromone glands showed that
they had the same three desaturase clones as ECB, only in this
case the only unsaturated product was that from the 1~14
desaturase. A sudden switch in pheromone components from
those produced by i\ 11 desaturation to the products of a different
biosynthetic pathway involving A14 desaturation could help
9180 1 www.pnas.org/cgi/doi/ 10.1 073/pnas. 1233767100
16:Acid o
ACB Pheromone
~14 desaturation
-
-
-
ECB Pheromone
-
_
l Oxidation
H _' I' ~ —~—~(OH
ZJE1 416: Acid
1 Oxidation
A12-14: Acid
7A F
reduction and ulceration
Z 8 E
14: Acid
1 ~11-desaturation
611-14: Acid
~12-14: OAc
^11-14: OAc
Fig. 1. Pheromone biosynthetic pathways for ACB and ECB from hexade-
canoic acid (16:Acid) and proceeding through different routes to the 14-
carbon acetate pheromone components.
explain how the ACB population was derived about a million
years ago. Studies of several other genes involved in the chemical
communication system of ACB and ECB also seem to be
unchanged, supporting the recent derivation of ACB. A single
pheromone binding protein in male antennae has an identical
structure in ACB and ECB males (23), and the reductase enzyme
in ACB and ECB has similar specificities with a preference for
Z1 1-14:Acid, even though ACB needs to reduce Z/E12-
14:Acids (24~.
The activation of a nonfunctional A14 desaturase in the
pheromone gland of an ancestral Ostrinza species would provide
the mutational shift in pheromone production needed to initiate
evolution of this chemical communication system, but would
there be any males to respond to this pheromone blend? A
screening of ECB males in flight-tunnel assays (25) showed that
there are rare males that exhibit a broad range of responses that
include flight to the ACB pheromone blend. This was surprising
because ECB males would not be expected to respond to 1~12
acetates and also because the ACB blend of 1~12 acetates
contains 33% of the E isomer (33% of E11 acetate is antagonistic
to this race of ECB males) (25~. However, the rare males
exhibited complete upwind flights to both the ECB (97:3 mix of
All acetates) and ACB blend (2:1 mix of /~12 acetates). The
existence of these rare males supports the possibility for a major
shift in pheromone blend being tracked by male response and fits
well with simulation models (26), showing that sudden major
switches in pheromone blend and male response seem more
likely than accumulation of small changes.
Discovery of the 1~14 desaturase in ECB gives rise to many
more questions on the origin of this gene and how long it has
been carried in moths. Research in the postgenomic era is
required to answer these questions.
Evolutionary Mechanisms
In a previous study (15), we identified several unique aspects of
insect desaturase multigene family evolution. The first was the
discovery that this family originated before the split among
Lepidoptera, Diptera, and Orthoptera, which is estimated to
have occurred ~350 million years ago (27~. Second, we found
that the family is composed of at least four gene clusters,
evolving at disparate rates that are correlated with the function
of each group (15~. For instance, the AZ9 (16 > 18) group has
evolved the slowest and contains metabolic desaturases, which
Roelofs and Rooney
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Representative terms from entire chapter:
pheromone components
~O[UVE11 AF441861
~OnUaE1 1 AF441221
BmO AF182405
POCZ10 AY017379
~_EPOE11 AY049741
_ ~ AVeaE1 1 AF416738
95 CrOZ/E11 AF518014
_ _ HZeZ1 1 AF272342
Tn jZ1 1 AF035375
~ Crc3NF AFS1~18
10
_ ^10,11
,.° ~ O!UZIE1 4 AY062023
^1 ~OnUZIE14 AF441220
-.—1 Dme CG1 5531 —a14
Dme CG9743
Dme CG9747
Dme CG8630
~ AdOZ9 AF338465
~q DSIZ9 ~2~848
! MdoZ9 AF417841
. ~ Dme CG7923
Dme CG5925
AVeZ9 AF243046
99 CrOZ9 AF518017
~ POCZ~ AP268276
.~. EPOZg AY061988
~OfUZ9 AY057862
~On~Z9 AF243047
HZeZ9 AF27~43
_OfUZ~ AYQS7863
_ ~OnUZg AF430246
_ EDOZ9 AF402775
I_Bmo AF182406
741~ HzeZ9 AF272345
- . TnIZ9 AF038050
Tkk U03281
Fig. 2. Phylogeny of the desaturase genes of various insect species Only
species for which complete sequences were available were used. The tree was
reconstructed from JTE amino acid distances (28) using the maximum likeli-
hood method as implemented in the PROML computer program in the PHYLIP
software package (http://evolution.genetics.washington.edu/phylip.html).
Numbers along branches indicate bootstrap support from 1,500 replicates.
The accession numbers for sequences are given after the species abbreviation,
as per ref. 15. In the case of D. melanogaster, the gene names are given as they
are listed in the Drosophila genome database (www.fruitfly.org). The tree is
rooted with the desaturase gene from the tick (Amb/yomma americanum),
and the log likelihood was -14,878.90361.
presumably represent the ancestral function of this gene family.
The AZ9 (18 > 16) and 1`l0,ll groups are composed of
sex-pheromone desaturase genes and evolved at faster rates,
which is coincident with a change in function from metabolism
to reproduction. The A14 group is made up of the fastest-
evolving sequences (lS) that are nonfunctional (e.g., li14 of O.
nubilalis), function in sex-pheromone biosynthesis (e.g., A14 of
0. furnacalis), or are yet to be functionally characterized.
Interestingly, we found (lS) that several dipteran sequences
cluster within this group (Fig. 2~. However, until we know the
function of the proteins these genes encode, we cannot be certain
on the basis of phylogenetic data alone, whether or not these
represent new groups distinct from the sequences classified in
the lL14 group.
Perhaps the most interesting finding of these studies ~s tnat
reciprocal desaturase gene nonfunctionalization in O. furnacalis
and O. nubilalis has resulted in a change in pheromone compo-
sition between these species. In this case, the A14 gene of O.
furnacalis makes a functional desaturase gene product but the
/`Z/Ell does not, whereas the opposite is true in O. nubilalis.
Because the ORFs of the AZ/Ell sequences are identical and
the ORFs of the /~14 differ by two amino acids in the Ostrinia
species, the reciprocal nonfunctionalization event must have
been relatively recent [i.e., at the time of their speciation, which
is inferred to have occurred ~l million years ago (R. Harrison,
personal communication), assuming a lepidopteran mtDNA
substitution rate of 2% per million years (29~. There are at least
two possible mechanisms to account for the nonfunctionality of
these genes. First, they may have recently become pseudogenes.
The paradigm pseudogene is one that is no longer transcribed,
possesses numerous nucleotide insertions or deletions resulting
in frameshift mutations, and is highly divergent from its func-
tional counterpart~s) (30~. What is unusual about the Ostrinia
nonfunctional genes is that they are well conserved between the
two species (Fig. 2~. However, recent pseudogenization could
Roelofs and Rooney
account for this. In addition, there are several other cases in
which pseudogenes have been shown to be conserved and even
possess intact ORFs. For example, the bacteria species Hae-
mophilus aegyptius and Haemophilus influenzue are sister taxa
that diverged very recently, and they each possess hap pseudo-
genes that are identical in sequence (31~. In this case, not enough
time has elapsed for substitutions to have occurred in either
species' pseudogene. Highly conserved pseudogene sequences
may also result among paralogues that duplicated recently, as has
been shown to occur among the three aquaporin pseudogenes of
humans (99% sequence similarity) (32) and the human desatu-
_ A9~16~18) rase functional/pseudogene pair (95% sequence similarity) (33~.
There are even numerous examples in which a pseudogene
produces an mRNA transcript but not a translated protein (e.g.,
refs. 34-37~.
Another possible explanation for the nonfunctionality of
_ '`9~18>16, certain Ostrinia genes is that there is an epigenetic mechanism
repressing their transcription and/or expression (38~. For ex-
ample, the genes nanos and pumilio interact to control the
translational repression of genes involved in embryonic pattern-
ing and spermatogenesis-oogenesis switching (39~. However, in
cells in which the nanos protein is not required, it is translation-
ally repressed through the action of at least two proteins known
as Smaug and Bicaudal (reviewed in ref. 40~. Such mechanisms
for translational repression are found throughout eukaryotes,
suggesting that a similar one might be involved in the control of
moth sex-pheromone desaturase genes, although whether or not
such a mechanism can account for the situation with Ostrinia
remains to be shown.
Nevertheless, sequence conservation and/or preservation of
an intact ORF do not necessarily ensure the functionality of a
gene. The only reliable way to determine whether a protein-
coding gene with an intact ORF is functional is to determine
whether a functional protein product is made. Our work on the
Ostrinia genes shows that in no instance has a protein product or
predicted sex pheromone been detected in an experimental assay
(15~. The next step, which we are currently pursuing, is to
determine how the genes became nonfunctional In this regard,
we draw attention to a study on the desat2 gene of Drosophila
melanogaster, which showed that this gene has been nonfunc-
tionalized in the Canton-S strain (41~. In this case, a mutation in
the promoter has inactivated the desat2 gene, rendering it a
recently formed pseudogene. Follow-up studies showed that this
nonfunctionalization of the desat2 gene has occurred in other
Drosophila races and has led to an incipient speciation event
among those with a functional desat2 versus those with a
nonfunctional desat2 (~42~. This is because the functional desat2
races produce a different pheromone complement than the
nonfunctional desat2 races, leading to a change in mating
response. In addition to explaining how sex-pheromone desatu-
rase gene nonfunctionalization has occurred in Drosophila, these
studies reinforce our contention that desaturase gene nonfunc-
tionalization can lead to speciation in insects (15~. One question
that remains is how gene duplication has played into the picture.
It is dear from the phylogenetic tree shown in Fig. 2 that there
have been various points of desaturase gene duplication during
the course of insect evolutionary history. The first duplication
event gave rise to the Al0,ll group followed by subsequent
duplications that gave rise to the /~14 and 1~9 (18 > 16) groups.
As we stated previously (lS), we believe that the ancestral gene
that gave rise to these groups was a Z9 gene from the /~9 (16 >
18) group based on the fact that (i) these genes function in
metabolism in much the same way as in other animals and (
~ -
New Pheromone
Blend
t
Gene Duplication, Loss,
or Nonfunctionalization
Lift in Detects\
System? )
\:Male Response:/
Fig. 3. Proposed birth-and-death mechanism for speciation in O. furnacalis
and nubitalis. Here, gene duplication, loss, or nonfunctionalization leads to a
shift in pheromone blend (female stimulus), which, if it leads to speciation, is
accompanied by a shift in the detection system (male response), which is also
brought about through gene duplication, loss, or nonfunctionalization.
and gave rise to the lepidopteran A9 (18 > 16) group (Fig. 1).
Similarly, the ancestral dipteran Z9 gene seems to have dupli-
cated sometime after the dipteran-lepidopteran divergence fol-
lowed by another duplication event, thus resulting in three
dipteran paralogs that cluster within the 1\9 (16 > 18) group (Fig.
1~. Likewise, a nonfunctional gene (CroNF) seems to have
duplicated from the ancestral moth Z11 gene, long before
Choristoneura rosaceana diverged from the other moth species
shown in the A10,11 group.
The above results, along with those of previous studies (15, 41,
42), indicate that gene duplication, gene loss, and pseudogene
formation have influenced the evolution of desaturase genes in
moths and flies. These processes are characteristic of the birth-
and-death model of multigene family evolution (43-464. Ac-
cording to the birth-and-death model, multigene families are
created through gene duplication, which gives rise to new
member genes, but some genes become deleted from the genome
or degenerate into pseudogenes, whereas others are maintained.
As a result, the members of a multigene family subject to
birth-and-death evolution will evolve more or less indepen-
dently, and different paralogs will be shared by different species
or lineages. Thus, in a phylogenetic analysis of a multigene family
including member genes from relatively closely related species,
we should see the following observations: (i) sequences will
cluster by gene or duplication order and not by species; (ii) low
levels of sequence homogeneity between genes will be observed
(especially at noncoding sites), except in the case of recent
duplicates; and (iii) evidence of gene loss/deletion or pseudo-
gene formation will be apparent. As we have already shown, all
of these occur in the insect desaturase multigene family. In
addition, we believe that the male moth pheromone-receptor
system, which is composed of olfactory receptor loci (47, 48),
may also be subject to a birth-and-death model of evolution,
perhaps as a result of coevolution with the desaturase multigene
family. A large and diverse olfactory receptor multigene family
would provide an adaptive advantage for male moths, allowing
for the rapid evolution of male response to female pheromone
blends (Fig. 3~. Although such studies have not yet been con-
ducted on moths, we predict that high levels of olfactory receptor
gene duplication, gene loss, and pseudogenization will be found
for these species on the basis of what has been found in other
animals. For instance, a family of at least 60 olfactory receptor
loci have been identified in D. melanogaster (49), and in verte-
9182 1 www.pnas.org/cgi/doi/, 0.4 073/pnas. 3 233767 100
-
//Major Shift in~
Signaling System
female Stimulu:/
~ Aga CG55982
80~1_ Aga CG55979
1 82 ~AgaCG55981
_ Dme CG9747
_ "gal ~~84!UU
65 _=CAga CG55969
_ Dme CG9743
'°° ~ Dme CG15531
_ ~ Aga CG55934
At_ Dme C<;8630
Aga CG47953
. . —Dme CG7923
74 Dme CG5925
B7 ~ - Aga CG50841
MdoZ9 AF417841
7~_ DsiZ9 AJ245748
i°°. DmeZ9 CG5887
TickU0328
20
Fig. 4. Phylogeny of the desaturase genes of flies. The tree was recon-
structed as in Fia. 1. Species and Gene abbreviations are as Der Fia. 2. The tree
_ . _ , ,~ _
is rooted with the desaturase gene from the tick (A. americanum), and the loo
... ... . _ . . . . .
.
likelihood was—9,899.33096. Note that only partial sequences for the A.
gambiae genes were available at the time this study was completed.
brates and Caenorhabditis elegans hundreds to thousands of
olfactory receptor loci, including many pseudogenes, have been
found (50, 51~.
Future Directions: Genomics and Beyond
One particularly interesting avenue for future research involves
studying how desaturase genes diversify once they duplicate. One
possibility is that the genes originated under a subfunctionaliza-
tion model (52-55) of gene family diversification (56~. Under the
subfunctionalization model, a generalized multifunctional gene/
protein duplicate gives rise to one or more paralogs. The
paralogous genes subsequently diverge and specialize in differ-
ent functions that were previously all carried out by the ancestral,
generalized multifunctional gene/protein. The problem is that
direct evidence confirming multifunctionality of the ancestral/
progenitor moth desaturase gene is lacking. An alternative to the
subfunctionalization model is one in which sex-pheromone genes
acquired their functions through rapid evolution subsequent to
gene duplication via positive Darwinian selection (e.g., ref. 57~.
The ancient origin of insect desaturase genes and their increased
substitution rates (15), which are highly pronounced in some
cases (e.g., the A14 and A10,11 group genes in Fig. 2), are highly
suggestive that nucleotide substitutions have become saturated.
This obviously confounds attempts to detect positive Darwinian
selection. For example, the estimate of synonymous substitution
(57) between the O. filrnacalis Z9 (16 > 18) and Z/E14 genes is
1.44 + 0.44 and is 2.96 + 1.20 between the Z/E11 and Z/E14
genes. Clearly, these values are well above the saturation level
(58, 59~. On the basis of our current state of knowledge, it is not
possible to determine which of the above models best fits the
insect desaturases. Further studies aimed at determining
whether the desaturases of basal insects (e.g., Collembola) are
multifunctional may help to answer this question.
Another promising area of research is on the genomics of
desaturase genes. Such studies are important for understanding
the underlying basis of insect reproduction in terms of behavior
as well as biochemistry. Consequently, until we obtain informa-
tion on the number of desaturase genes and their genomic
organization in various insect lineages, our understanding of the
core elements of insect reproduction will be incomplete. Fur-
thermore, such studies will provide insights into the evolution of
insect desaturases. For example, now that their complete ge-
Roelofs and Rooney
A
~ ~ ~ ~ , -
IC(;7g23~ 1 ~ ~ ~747 - Gt55~9743 ~ ICG - 25ICG~71 '~301
-
.. f , j ,~., ~ ......
.'
_""-- ~ ~ ~ ~ _- ~
3_ i=—_
Fig. 5. Genomic map of desaturase genes in D. melanogaster chromosome
3 (A) and A. gambiae chromosome 2 (B). The vertical lines bisecting the D.
melanogaster chromosome represent the division between the left and right
arms of chromosome 3. Arrows above genes represent the direction of tran-
scription. Dotted lines connect orthologous counterparts between species, on
the basis of clustering patterns depicted in Fig. 4.
nome sequences are available, we know that the D. melanogaster
and Anopheles gambiae genomes possess seven and eight desatu-
rase genes, respectively (Figs. 4 and 5~. By combining informa-
tion from phylogeny (Fig. 4) and genomic organization (Fig. S),
we can make inferences regarding several aspects of desaturase
evolution, genomics, and even gene function in dipterans. For
instance, CG50841 of A. gambiae clusters significantly with the
D. melanogaster genes CG5925 (desat2) and CG5887 (desatl).
Because both of these genes are Z9 desaturases (Fig. 2), we
predict that the A. gambiae CG50841 is also a Z9 desaturase.
The above approach can also be used to deduce the history of
desaturase gene duplication and genomic organization. For exam-
ple, the dotted lines connecting genes in Fig. 5 represent inferred
orthologous relationships. Thus, D. melanogaster CG9747 and the
ancestor of A. gambiae genes CG55981, CG55979, and CG55982
share an orthologous relationship. One piece of evidence for this
comes from our phylogenetic analysis (Fig. 4), in which these genes
cluster together with relatively high statistical support. Another
piece of evidence comes from the conserved genomic organization
of the fruit fly and mosquito desaturase genes (Fig. 5~. In this case,
the genomic location and direction of transcription of inferred
orthologous gene pairs is conserved relative to other gene pairs. By
using this line of reasoning in light of our results, we can also infer
that there were four rounds of gene duplication that took place
before the divergence of mosquitoes and flies. These events gave
rise to (i) D. melanogaster CG8630 andA. gambiae CG47953, (ii) the
ancestor of D. melanogaster CG5925/CG5887 and A. gambiae
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Roelofs and Rooney
CGS0841, (iii) D. melanogaster CG9743 end A. gambiae CG55969,
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whatever modelts) are uncovered will reflect this functional and
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Roelofs and Rooney
l`<
Arthur M. Sack/er
_ C O L L O Q U I A
~ OF THE NATIO NAL ACADEMY OF SCIENCES
Chemical Communication in a Post-Genomic World
January 17-19, 2003
Arnold and Mabel Beckman Center, Irvine, CA
Organized by May R. Berenbaum and Gene E. Robinson
Program
Friclay, January 17
Welcome Reception and Registration
Saturday, January 18
Genomics of Chemical Attraction: Genomics of Olfaction
Cori Bargmann, University of California, San Francisco
Genomics of Olfaction in Caenorhabditis elegant
John Carlson, Yale University
Genomics of Olfaction in Drosophila melanogaster
Linda Buck, Fred Hutchinson Cancer Research Center
Genomics of Olfactory Receptors in Rodents
Gene Robinson, University of Illinois, Urbana-Champaign
Pheromone Regulation of Division of Labor in Honey Bee Colonies: From Behavior to
Gene Expression Profiles in the Brain
Genomics of Chemical Attractions: Molecular Genetics, Evolution, and
Biochemistry of Attractant Signaling
Hugh Robertson, University of Illinois, Urbana~hampaign
Molecular Evolution of the Insect and Nematode Chemoreceptor Superfamilies
Wendell Roelofs, Cornell University
Molecular Genetics of Pheromone Biosynthesis in Lepidoptera
John Hildebrand, University of Arizona
Central Processing of Chemosensory Signals: Anticipating Post-Genomic Opportunities
Nancy Moran, University of Arizona
Interactions of Insects with Bacterial Symbionts
Thomas Eisner, Cornell University
Can We Survive Without Natural Products Chemistry?
Jerrold Meinwald, Cornell University
The Role of Natural Products Chemistry in a Post-Genomic World
Sunday, January 19
Molecular Genetics/Genomics of Chemical Repulsion and Defense:
Predator/Prey and Pathogen/Host Interactions
Baldomero Olivera, University of Utah
Venomous Cone Snails, Specialists in Neuropharmacology: Reconstructing an
Evolutionary History of Drug Development
Bonnie Bassler, Princeton University
How Bacteria Talk to Each Other
Sir David Hopwood, John Innes Centre, Norwich Research Park
Streptomyces Secondary Metabolites: Much More than Weapons of Mass Destruction
Klaus Hahlbrock, Max Planck Institute
Transcriptional Reprogramming and Secondary Metabolite Accumulation During
Plant/Pathogen Interactions
Molecular Genetics/Genomics of Chemical Repulsion and Defense: Plant/lnsect
Interactions
Ian Baldwin, Max Planck Institute
Allopolyploid Speciation and Its Effects on Ecological Adaptations: Plant-Herbivore
Interactions in Nicotiana Native to North America
Clarence Ryan, Washington State University
Signaling for Herbivore Resistance in Solaneceae Species: Systemins, Prosystemins, and
the Systemin Receptor
Thomas Mitchell-Olds, Max Planck Institute
Genomics of Plant/lnsect Interactions in Arabidopsis
May Berenbaum, University of Illinois, Urbana-Champaign
Cytochrome P450s in Insects An Embarrassment of Riches
