J Comp Physiol A (2005)
DOI 10.1007/s00359-005-0044-y
R EV IE W
M. Rützler Æ LJ Zwiebel
Molecular biology of insect olfaction:recent progress and conceptual
models
Received: 11 May 2005 / Revised: 3 July 2005 / Accepted: 12 July 2005

Springer-Verlag 2005
Abstract Insects have an enormous impact on global
public health as disease vectors and as agricultural e-
nablers as well as pests and olfaction is an important
sensory input to their behavior. As such it is of great
value to understand the interplay of the molecular
components of the olfactory system which, in addition
to fostering a better understanding of insect neurobiol-
ogy, may ultimately aid in devising novel intervention
strategies to reduce disease transmission or crop dam-
age. Since the first discovery of odorant receptors in
vertebrates over a decade ago, much of our view on how
the insect olfactory system might work has been derived
from observations made in vertebrates and other inver-
tebrates, such as lobsters or nematodes. Together with
the advantages of a wide range of genetic tools, the
identification of the first insect odorant receptors in
Drosophila melanogaster in 1999 paved the way for rapid
progress in unraveling the question of how olfactory
signal transduction and processing occurs in the fruitfly.
This review intends to summarize much of this progress
and to point out some areas where advances can be
expected in the near future.
Keywords Odorant binding proteins Æ Odorant
degrading enzymes Æ G protein coupled receptors Æ
OR83b/OR7 Æ Sensory neuron membrane proteins
Abbreviations AL: Antennal lobe Æ CNG: Cyclic
nucleotide gated Æ CSP: Chemosensory protein Æ DOR:
Drosophila odorant receptor Æ GRCR: G protein
coupled receptor Æ GRK: G protein coupled receptor
kinases Æ LN: Local neuron Æ OBP: Odorant binding
protein Æ ODE: Odor degrading enzymes Æ OR:
M. Rützler Æ LJ Zwiebel (&)
Department of Biological Sciences,
Program in Developmental Biology and Center
for Molecular Neuroscience, Vanderbilt University,
VU Station B 351634, Nashville, TN 37235-3582, USA
E-mail: l.zwiebel@vanderbilt.edu
Tel.: +615-343-1894
Fax: +615-936-0129
Odorant receptor Æ ORN: Olfactory receptor
neuron Æ PBP: Pheromone binding protein Æ PN:
Projection neuron Æ PBPRP: Pheromone binding
protein–related protein Æ SNMP: Sensory neuron
membrane protein Æ VA: Vaccenyl acetate
Introduction
The molecular and cellular characterization of olfactory
processes in insects has achieved an enviable rate of
growth encompassing a wide range of approaches and
systems. Indeed, one might argue that in several in-
stances, but most significantly regarding the molecular
characterization of odorant specificity, the insect olfac-
tory systems have proven to be more experimentally
robust than their mammalian counterparts. In this re-
view, we will attempt both to catalog recent advances
and to place these data within a broader biological
context. It should be noted that this is not a simple task;
indeed, the spectrum of methodologies range from bio-
informatics and genomics to real-time imagery of
olfactory signal transduction and processing. Further-
more, while the fruitfly, Drosophila melanogaster, re-
mains at the forefront of insect model systems, vigorous
experimental programs focusing on insect olfaction have
been established, which transcend the traditional limits
of ‘‘academic models‘’. These latter studies serve to
broaden our appreciation for the conservation of
olfactory mechanisms within insects as well as their di-
verse implementations. At the same time, they enhance
our understanding of processes in insects that function
as agricultural enablers and pests, as well as vectors for a
range of pathogens that cause important human diseases
such as malaria, Dengue and Yellow fever. Ultimately,
these studies may facilitate the design and implementa-
tion of novel intervention strategies directed against
critical olfactory-driven behaviors of medically and
economically important insects.
Until recently, much of our view of insect odorant
perception at the molecular level was strongly influenced
by observations that were made in vertebrates, crusta-
ceans and nematodes (Hildebrand and Shepherd 1997;
Krieger and Breer 1999). The canonical model involves a
family of heptahelical G protein coupled receptors
(GPCRs) that activate downstream effectors via het-
erotrimeric GTP binding (G) proteins that consist of
single a, b and c subunits. In general, GPCR-agonist
binding promotes GDP release from the a-subunit of the
G protein that in turn binds GTP and thereby adopts its
active state. This either triggers a structural rearrange-
ment in the heterotrimeric protein (Bunemann et al.
2003) or release of the bc-subunit (Hamm 1998; Jane-
topoulos et al. 2001). Any of the active components
Gabc.GTP, Ga.GTP or Gbc can in turn regulate a
variety of effectors such as adenylyl cyclase, phospho-
lipases or ion channels (McCudden et al. 2005) before
returning to the inactive state following GTP hydrolysis,
which is catalyzed by the GTPase domain of the a-
subunit (Markby et al. 1993). In vertebrate olfactory
signal transduction, active Gaolf stimulates adenylyl cy-
clase to synthesize the second messenger cyclic 3¢,5¢-
adenosine monophosphate (cAMP) (Jones and Reed
1989; Sinnarajah et al. 1998) while lobster olfactory
receptor neurons (ORNs) form two second messengers,
inositol 1,4,5-tris-phosphate (IP3) and cAMP in re-
sponse to individual odors (Fadool and Ache 1992;
Michel and Ache 1992; Boekhoff et al. 1994). The in-
crease in cAMP levels in vertebrates triggers the opening
of a cyclic nucleotide gated (CNG) Ca2+ channel that in
turn activates a Cl channel, generating an ORN action
potential (AP) (Firestein 2001).
It has long been assumed that the canonical model of
olfactory signal transduction would also hold in insects
where, in contrast to the shared olfactory epithelium of
most vertebrate systems, the peripheral olfactory system
is segregated in a large number of distinctive chemo-
sensory hairs known as sensilla which are found on the
antennae, maxillary palps and other sensory appendages
(Fig. 1). Indeed, several of the ‘‘usual’’ molecular sus-
pects have been identified and in part functionally
characterized in insect olfactory organs. These include
arrestins (Merrill et al. 2002, 2003, 2005), odorant
binding proteins (OBPs) (for review see Pelosi and
Maida 1995; Tegoni et al. 2004), a heterotrimeric G
protein (Laue et al. 1997) as well as a CNG (Baumann
et al. 1994; Dubin et al. 1998) and a IP3-gated ion
channel (Stengl 1994) (Fig. 1). In a study using the
cockroach, it has been demonstrated that pheromone
exposure of insect antennal preparations causes a rapid
increase in IP3 levels (Breer et al. 1990), which, in a
follow-up study, could be inhibited by pertussis toxin
(Boekhoff et al. 1990), indicating that the IP3 increase is
dependent upon either a Gai or a Gao heterotrimeric G
protein subunit. Furthermore, pheromone receptor
neuron activity of Bombyx mori could be stimulated
with fluoride ions (Laue et al. 1997) that are known to
activate heterotrimeric G proteins via binding to the a-
subunit in combination with magnesium ions (Antonny
et al. 1993). Despite this growing wealth of information,
Fig. 1 The insect antenna typically bears between several hundred
and some thousand sensillar hairs, depending on the species, which
are depicted on the left. Each sensillum contains between 1 and 4
ORNs (red and dark blue, respectively). These are bipolar neurons
which, on one end, extend a dendrite that is bathed in sensillar
lymph and interacts with odorants and on their other end project
axons that terminate in the AL, where olfactory information is
processed. ORNs are embedded in a layer of support cells (light
green) that secrete proteins such as OBPs (orange and light blue)
into the sensillum lymph. Odorants can enter the sensillum lymph
via pores in the cuticle and cross the lymph in a hitherto still
debated way that may involve OBPs as transport vehicles.
Subsequent odorant-OR binding takes place along the dendrite
of an ORN and may activate a heterotrimeric G-protein to target
one or more of many possible effectors and finally gates ion
channels thereby creating APs
it must be noted that the precise mode of insect olfactory
signal transduction remains largely obscured.
Before odorants, which are commonly hydrophobic
molecules, can be detected by any kind of membrane-
associated receptor, they must traverse through the
aqueous environment that surrounds ORNs, the
olfactory mucosa in vertebrates and the sensillum
lymph in insects. In both cases the abundant secretion
of specific proteins has been postulated to play a vital
role in this aspect of olfactory processes. These se-
creted proteins form the OBP family and are the
product of several classes of support cells that are
found together with ORNs in insect sensilla (Fig. 1).
Several classes of OBPs belonging to the lipocalin gene
family have been identified in vertebrates (Dear et al.
1991). Lipocalins typically are 160–180 amino acids
long, extracelluar, soluable proteins. They often share
less than 20% pairwise similarity but are identified by
short characteristic sequence motives. Members of the
Lipocalin gene family are generally considered to be
involved in the transport of small molecules such as
odorants or retinal, but it has become clear later that
family members have a variety of other roles such as
invertebrate cryptic coloration or prostaglandin syn-
thesis (Flower et al. 1991, 2000; Flower 1996; Fuentes-
Prior et al. 1997). Insect OBPs have been isolated from
a wide variety of species ranging from Lepidopteran
systems where they were first identified (Vogt et al.
1991) to D. melanogaster (McKenna et al. 1994; Pik-
ielny et al. 1994), Apis mellifera (Briand et al. 2001)
true bugs (Dickens et al. 1998) and more recently, the
mosquitoes Anopheles gambiae (Biessmann et al. 2002;
Xu et al. 2003), Culex quinquefasciatus (Ishida et al.
2002) and Aedes aegypti (Ishida et al. 2004). These
proteins are unrelated in sequence to their vertebrate
counterparts and hence may fulfill different functions.
The first functional characterization of one member of
this protein family, D. melanogaster OBP76a/Lush (Xu
et al. 2005), has stimulated discussions about the
contributions of these molecules in insect olfactory
signal transduction. In this review, we will attempt to
integrate these observations into a general model of
insect olfactory signal transduction.
In addition to ligand (agonist)-based activation, an
important component of sensory perception is the
cessation or reduction of signaling and several mech-
anisms have been suggested to contribute in this pro-
cess in insect olfaction. Following the activation of
cognate ORs, odorants need to be removed from the
immediate vicinity of the receptor so that fresh plumes
of odor can be discriminated. This function is carried
out by a biochemical diverse array of enzymes, which
have been collectively referred to as odor degrading
enzymes (ODEs) (Jacquin-Joly and Merlin 2004). A
number of different enzymatic activities, which are
capable of rapid chemical modification (Rybczynski
et al. 1989) or degradation of odorants (Vogt et al.
1985), have been characterized in the past and recently
two groups reported cloning of candidate molecules
for such biochemical activities (Ishida and Leal 2002;
Maibeche-Coisne et al. 2004). On the cytoplasmic side
of the ORN plasma membrane, desensitization of
GPCR-mediated signal transduction is, in part, medi-
ated through an impairment of the receptor’s ability to
activate its corresponding G protein. This process has
been extensively studied in the context of mammalian
GPCRs where it is carried out principally through the
combined activity of two classes of proteins: G protein
coupled serine/threonine receptor kinases (GRKs) and
arrestins (Freedman and Lefkowitz 1996). Phosphory-
lation of a GPCR by a GRK serves to promote the
binding of an arrestin protein, which further uncouples
the receptor from the G protein-based signaling cas-
cade (Pippig et al. 1993). Our laboratory has demon-
strated that a set of arrestin genes known to be active
in visual signal transduction (Hyde et al. 1990; Smith
et al. 1990; Alloway et al. 2000) also function in
olfactory signal transduction in D. melanogaster
(Merrill et al. 2002) and has identified several arrestin
family members expressed in olfactory tissues of An.
gambiae (Merrill et al. 2003).
Odorant receptors
While Buck and Axel (Buck and Axel 1991) used a novel
polymerase chain reaction (PCR)-based approach to
clone the first candidate ORs from rat olfactory epi-
thelium, similar approaches were unsuccessful in insects.
In what has proven to be the hallmark for the cloning of
all insect ORs, the completion of the Drosophila genome
sequence (Adams et al. 2000) was necessary for the
identification of a large family of candidate ORs in the
fly (Clyne et al. 1999). In this ground-breaking study,
the first putative D. melanogaster ORs (DORs) were
identified using a novel computer algorithm that sear-
ched for diagnostic features of the GPCR superfamily,
including hydropathy, polarity and weighted amino acid
composition of the predicted protein (Kim et al. 2000).
This approach was unique in that it did not rely on
searching for putative local or global homologies (which
might miss a divergent member of a particular family)
but rather sought to identify putative DORs by search-
ing for genes that display the physicochemical properties
derived from other established transmembrane proteins.
This report was followed by independent molecular and
bioinformatics studies (Gao and Chess 1999; Vosshall
et al. 1999) that together described members of a highly
divergent family of receptors, displaying between 10%
and 75% identity and bearing no significant homology
to any other GPCR family (Smith 1999).
Similar approaches were also used to identify the
complete repertoire of OR genes for the malaria vector
mosquito An. gambiae (Fox et al. 2002; Hill et al. 2002)
which, to date, is the only other insect whose complete
genome has been made publicly available. Nevertheless,
additional OR sequences have been identified for other
insects including the yellow fever mosquito Aedes ae-
gypti (Melo et al. 2004), the tobacco budworm Heliothis
virescens (Krieger et al. 2002, 2004) and the silk moth B.
mori (Sakurai et al. 2004; Nakagawa et al. 2005). Fur-
thermore, unpublished data has also been made avail-
able for identifying 166 ORs and 6 OR pseudogenes
from the genome of the honey bee Apis mellifera (H.
Robertson, personal communication).
Since the initial discovery of insect ORs, great pro-
gress has been made in understanding the function of
this receptor family and an increasingly clearer picture
of their contribution to olfactory coding is emerging.
Much of the recent progress has been made possible by
two major breakthroughs, both of which take advantage
of the powerful experimental toolbox available in Dro-
sophila. The first involves the introduction of a novel
approach for the ectopic expression of ORs that has
been established in D. melanogaster by the laboratory of
John Carlson (Yale University) which facilitates the
study of insect odorant receptor function in vivo (Do-
britsa et al. 2003). In this ‘‘empty neuron’’ system, a
deletion at the genomic locus of DORs 22a and 22b
(Dhalo) leads to the loss of olfactory sensitivity in the
well-defined ab3A neuron, where both receptors are
normally expressed. In order to characterize the contri-
bution of DORs 22a and 22b to the sensitivity of the
wild-type ab3A neuron, Dobritsa et al. (2003) ectopi-
cally expressed these ORs individually in the ab3A
neuron, utilizing the bipartite GAL4/UAS system
(Brand and Perrimon 1993) under control of the
endogenous DOR22a promoter region. This approach
revealed that of the two receptors DOR22a and
DOR22b, which are both expressed in the ab3A neuron,
only DOR22a conferred sensitivity to odorants, sug-
gesting that DOR22b is not functional in the ab3A
neuron. Furthermore, a resting frequency of action
potentials (APs), albeit abnormally low, for the ab3A
neuron is still detectable in Dhalo mutants, indicating
that a functional OR is not required for ORN devel-
opment.
The real strength of this approach was revealed when
Dobritsa et al. (2003) subsequently tested whether Dro-
sophila ORs that are normally not expressed in the ab3A
neuron could function in this ORN. Surprisingly, ecto-
pic expression of DOR47a in the Dhalo background re-
stored odorant sensitivity to ab3A neurons.
Importantly, ectopic expression of DOR47a shifted the
ab3A neuron’s response profile to that of the ab5B
neuron, thus, indicating that DOR47a is the functional
receptor housed in the ab5B neuron (deBruyne et al.
2001). In this manner the Carlson laboratory not only
provided compelling evidence that DORs 22a and 47b
are bona fide ORs, but more importantly, have devised a
powerful new approach for the functional characteriza-
tion of other ORs. This system was subsequently used to
deorphanize two ORs from the malaria vector mosquito
An. gambiae (AgORs) (Hallem et al. 2004a), further
demonstrating utility of this approach to functionally
characterize ORs from different insect species. In this
study, expression of AgOR1, in ab3A ORNs resulted in
specific responses to 10 3 M concentrations of 4-meth-
ylphenol (4MP). This is particularly interesting because
AgOR1 is specifically expressed in vivo in blood-feeding
female mosquitoes (Fox et al. 2001) and 4MP has also
been identified as a component of human sweat (Cork
and Park 1996). These data raise the possibility that
AgOR1 presumably acting together with other AgORs
may play a critical role in mediating the olfactory-based
preference for human hosts (anthropophily) that lie at
the heart of the high vectorial capacity of An. gambiae.
As such, it would be a prime target for chemical or ge-
netic-based intervention strategies designed to reduce
the ability of An. gambiae to transmit human malaria.
In a massive and elegant set of studies that has fun-
damentally affected our current view of odorant dis-
crimination in insects, the empty neuron approach has
subsequently been utilized to characterize the olfactory
response profile for the antenna of D. melanogaster
(Hallem et al. 2004b). Of 32 DORs that have previously
been reported to be expressed in Drosophila antennae, all
but one were expressed and characterized in this study.
Twenty-four of these were functional in this context, 13
of which conferred an odorant response profile in the
ab3A neuron that closely resembled odorant responses
of previously characterized Drosophila olfactory neurons
(deBruyne et al. 2001). Moreover, the response profiles
of all 24 functional DORs were distinct.
Importantly, these studies suggest for the first time as
to how the olfactory system of Drosophila can discrim-
inate between hundreds or thousands of odorants, with
only a relatively limited (compared to mammalian ge-
nomes) number (62) of DORs that have been detected in
the fly genome (Robertson et al. 2003). Some of the
receptors responded strongly to only one of the tested
compounds while other ORs elicited strong reactions to
a number of chemicals, in keeping with the generally
accepted combinatorial mode of OR specificity (Malnic
et al. 1999). Particular odorants, on the other hand,
activated a variable but specific subset of receptors.
These events underlie a combinatorial discrimination of
odorants, which has previously been observed in the first
center of neuronal integration, the antennal lobe (AL)
(Fiala et al. 2002; Ng et al. 2002; Wang et al. 2003).
In order to image odorant-stimulated activity across
the entire AL, Ng et al. (2002) and Wang et al. (2003)
selectively expressed fluorescent proteins that respond to
odorant stimulation with fluorescence intensity changes
that indirectly reflect Ca2+ levels in defined neuronal
subsets: ORNs, projection neurons (PNs) which relay
odorant information to higher brain centers, and in local
interneurons, which are exclusively GABAergic inhibi-
tory neurons (Jackson et al. 1990; Buchner 1991). Both
studies observed that odorant information obtained at
the periphery is similarly represented at the level of the
AL: At low odorant concentrations, some single odor-
ants did not elicit glomerular activity in the AL while
other odorants evoke activity in a small, odorant-specific
set of glomeruli. Stimulation with higher odorant con-
centrations consistently evoked activity in additional
glomeruli, which most likely reflects stimulation of
additional receptor populations in antennae as observed
by Hallem et al. (2004b). Importantly, even though at
high concentrations different odorants may activate
overlapping populations of ORNs, the individual re-
sponse profile of each ORN type observed by Hallem
et al. (2004b) is reflected as a distinct combinatorial
pattern of glomerular activity at the AL (Wang et al.
2003). Moreover, the activity patterns observed by Ng
et al. (2002) and Wang et al. (2003) were similar in
presynaptic ORNs and postsynaptic PNs. Information
transmission through glomerular relays took place in the
presence of widespread inhibitory local neuron (LN)
activity. Most odor representations by ORNs and PNs,
in fact, received simultaneous inhibition by LNs, while
in other glomeruli LN synapses were active in the ab-
sence of ORN afferent activity (Ng et al. 2002).
While the monitoring of odorant sensitive fluores-
cence intensity changes now provides superb spatial
resolution of glomerular activity in the entire AL, these
methods are unable to resolve information that may be
encoded within AP trains, which can only be monitored
by electrophysiological methods (Laurent 1999). Indeed,
neural ensemble recordings have been utilized (Chris-
tensen et al. 2000; Lei et al. 2004) to study electrical
activity of the antennal lobe in the moth Manduca sexta.
In these important experiments, an array of 16 recording
electrodes facilitates simultaneous monitoring of elec-
trical activity, thereby, combining the powerful temporal
resolution capacity of electrophysiology with spatial
information regarding glomerular activity. Furthermore,
Lei et al. (2004) observed that similar odorants cannot
be discriminated with confidence solely based on the
combinatorial, spatial glomerular activation pattern.
Temporal patterns of different ORN population, APs,
however, may be compared and integrated at the AL by
LNs (Lei et al. 2004). Such processing may shape PN
output activity to the protocerebrum (Lei et al. 2004) or
mushroombody, whereas in Drosophila, both odorant
concentrations and identities are represented as stereo-
typed patterns of neuronal activity (Wang et al. 2001,
2004).
Another important finding of Hallem et al. (2004b)
was that the resting frequency of APs of the ab3A
neuron was directly dependent on the ectopically ex-
pressed DOR and these new resting frequencies of APs
essentially recapitulated the characteristics of the native
ORN for each of the DORs studied. Resting frequencies
of APs are an important characteristic of insect ORNs
and can vary between 1 and 30 spikes per second in
Drosophila (de Bruyne et al. 1999, 2001). It has been
suggested that this frequency determines the amount of
information that can be encoded either in stimulation or
inhibition of ORN APs through odorants, both of which
are represented in the antennal lobe (de Bruyne et al.
1999, 2001; Shields and Hildebrand 2001).
For those cases of DORs that showed little or no
response, Hallem et al. (2004b) hypothesized that some
might not be normally functional in vivo as was the case
for DOR22b. In addition, these proteins could con-
ceivably respond to odorants that lie outside of the
diagnostic set of 11 compounds utilized in this study.
Furthermore, the substances used in these experiments
were chosen based on their ability to discriminate be-
tween ORN classes that were previously identified
(deBruyne et al. 2001). They included chemicals that are
known to be relevant to Drosophila behavior but did not
contain specialized compounds such as pheromones
(Hallem et al. 2004b), which are key substances in the
sexual behaviors of insects that elicit significant re-
sponses from a highly specialized subset of ORNs (Bo-
eckh and Boeckh 1979; Kaissling 1996; Clyne et al.
1997). In another novel study from the Carlson labo-
ratory (Goldman et al. 2005), it has recently been dem-
onstrated that in insects there is at least one exception to
one of the central hypotheses of vertebrate and insect
olfaction, being that one ORN can express only a single
functional OR (Ressler et al. 1993; Vassar et al. 1993;
Clyne et al. 1999; Gao and Chess 1999; Malnic et al.
1999; Vosshall et al. 1999). In this case, two odorant
receptors are co-expressed in the pb2A neuron on the
maxillary palp of Drosophila, which together account for
the odorant response profile of this ORN (Goldman
et al. 2005). If a subset of the antennal ORNs also co-
expresses more than one functional OR, this may pro-
vide a rationale for the observation that the odorant
sensitivities of several DORs ectopically expressed in the
empty neuron system could not be correlated with any
known ORN response profiles from Drosophila (Hallem
et al. 2004b).
More recently, the ‘‘empty neuron’’ approach has
been extended to encompass the larval olfactory system
in Drosophila (Kreher et al. 2005) where a set of larval
DOR proteins were examined. Here, two broad classes
of larval DORs were identified: the first class displaying
strong responses to aliphatic odorants and another
being highly sensitive to odorants that contain a benzene
ring. In addition, a single larval DOR gene (DOr67b)
displayed strong responses to both types of odorants
(Kreher et al. 2005). Importantly, by analyzing the lar-
val antennal lobe (LAL) projections derived from larval
DOR GAL4 drivers and GFP reporters, Kreher et al.
(2005) were able to show that larval ORNs expressing a
unique larval OR gene map to discrete and non-over-
lapping glomeruli within the LAL. Even more striking is
the observation that convergence of larval ORNs to a
common LAL target was never observed and, interest-
ingly, larval ORNs expressing DORs that display similar
odorant specificities map to clustered LAL glomeruli
(Kreher et al. 2005). Taken together, these observations
suggest that for Drosophila, and perhaps for other in-
sects, organization of the olfactory system is simpler in
larvae than in adults but information coding is similar.
The second breakthrough study that has sparked re-
cent progress in understanding the mechanisms under-
lying the insect olfactory apparatus was the functional
characterization of Drosophila DOR83b by the labora-
tory of Leslie Vosshall (Larsson et al. 2004). In contrast
to other ‘‘conventional’’ members of the insect OR
family that share little sequence homology and are ex-
pressed in a small subset of olfactory neurons, homologs
of DOR83b, which have been identified in several insect
orders including Diptera (Krieger et al. 2003; Melo et al.
2004; Pitts et al. 2004), Hymenoptera (Krieger et al.
2003), Lepidoptera (Krieger et al. 2003; Nakagawa et al.
2005), and Coleoptera (Krieger et al. 2003), share high
sequence conservation of more than 60% amino acid
identity. Moreover, each of these homologs are ex-
pressed in a large majority of the ORNs in each insect
(Vosshall et al. 1999; Krieger et al. 2003; Melo et al.
2004; Pitts et al. 2004; Nakagawa et al. 2005). Taking
advantage of a recent innovation that uses homologous
recombination to facilitate the targeted knockout of
genes in Drosophila (Gong and Golic 2003), Larsson
et al. observed that deletion of DOR83b from ORNs
resulted in a general insensitivity to odorants in larval
and adult stages of the fruitfly (Larsson et al. 2004).
Insight into the underlying molecular rationale of this
phenotype was gained by the observation that in
DOR83b mutants conventional DORs that are typi-
cally co-expressed with DOR83b were no longer cor-
rectly localized along the dendritic portion of ORNs.
Instead, the bulk of these DOR proteins remained in the
soma which is in agreement with the hypothesis that
DOR83b facilitates trafficking of general ORs to the
dendrite (Larsson et al. 2004). Consistent with the high
sequence conservation that is characteristic among
OR83b family members, antennal transgene expression
of respective orthologues from An. gambiae, the Medi-
terranean fruitfly Ceratitis capitata, and the corn ear-
worm moth Helicoverpa zea in Dor83b mutant flies
restored odorant sensitivity and facilitated membrane
trafficking of conventional ORs to a similar degree as
ectopic expression of DOR83b itself (Larsson et al.
2004; Jones et al. 2005). This provides direct evidence
for functional conservation of OR83b proteins in a wide
range of insects.
These results hearken back to the observation that in
many cases vertebrate GPCRs form homo and hetero-
oligomeric complexes, which contribute to GPCR func-
tion in several ways (Terrillon and Bouvier 2004). It has
been suggested (Reddy and Corley 1998) that formation
of oligomeric complexes is important for the escape of
some molecules from the endoplasmic reticulum, which
could explain the retention in the soma of general ORs in
the absence of an OR83b family protein. In this light it is
worthwhile to note that heterologous expression systems
for many GPCRs and in particular, a large number of
chemoreceptors have traditionally failed to correctly
traffic these proteins to the plasma membrane leading to
a significant impediment to the functional characteriza-
tion of candidate ORs (McClintock et al. 1997; Gimel-
brant et al. 1999, 2001). Accordingly, there has been
considerable speculation that other genes are expressed
in chemosensory neurons whose products aid in the
membrane trafficking of ORs. Such molecules have
subsequently been identified in nematodes (Dwyer et al.
1998) and vertebrates (Hague et al. 2004). The results of
Larsson et al. (2004) suggest that DOR83b and its ho-
mologues may play a similar role in plasma membrane
trafficking of conventional OR proteins.
Indeed, oligomer formation of DOR83b family pro-
teins and conventional insect ORs has since been ob-
served in two independent studies. In the first set of
experiments, DOR83b was expressed in Human
Embryonic Kidney (HEK) 293 cells along with two
conventional ORs, DOR22a and 43a, respectively
(Neuhaus et al. 2005). Examining bioluminescence res-
onance energy transfer (BRET) between fluorescence-
tagged versions of each of these receptors, which can be
used to monitor physical proximity between molecules
(Xu et al. 1999), ligand-independent formation of het-
ero-oligomeric complexes between DOR83b and
DOR22a as well as DOR83b and DOR43a was observed
(Neuhaus et al. 2005). Similarly, formation of the
respective homo-oligomeric complexes was also ob-
served (Neuhaus et al. 2005). Importantly, co-expression
of DOR 43a along with DOR83b dramatically enhanced
cyclohexanone responses in transfected cells (Neuhaus
et al. 2005), a known agonist of DOR43a (Stortkuhl and
Kettler 2001; Wetzel et al. 2001; Wang et al. 2003;
Hallem et al. 2004b). In contrast to the in vivo obser-
vation of anosmia in DOR83b-deficient flies (Larsson
et al. 2004), HEK293 transfection with either DOR22 or
DOR43 alone also conferred sensitivity to their respec-
tive cognate agonists, albeit at greatly reduced sensitivity
and in tenfold fewer cells (Neuhaus et al. 2005).
In another recent study, DOR83b was similarly
shown to facilitate the function of DOR47a in Xenopus
laevis oocytes (Nakagawa et al. 2005). In a similar
fashion, the effect of two DOR83b homologues, BmOR2
and HR2 from the moths B. mori and H. virescens
(Krieger et al. 2003), were examined. In both cases, these
OR83b family proteins increased plasma membrane
localization of the B. mori pheromone receptor BmOR1
(Sakurai et al. 2004), providing further evidence that
DOR83b family receptors may play a general and, in-
deed, essential role in the trafficking of conventional
ORs in vivo (Nakagawa et al. 2005). In a previous
study, this group recorded ligand-dependent responses
derived from Ca2+ sensitive Cl channels in Xenopus
oocyte voltage clamp studies only when BmGaq was co-
expressed along with only BmOR1 and not with 83b
family protein (Sakurai et al. 2004), which supports
previously obtained in vivo results that insect ORs
function as bone fide GPCRs (Boekhoff et al. 1990;
Laue et al. 1997). Surprisingly, in a follow-up study, an
additional current in response to Bombykol, a BmOR1
agonist, was observed when BmOR2 was co-expressed
along with BmOR1 even in the absence of BmGaq
(Nakagawa et al. 2005). Further characterization of this
response (Nakagawa et al. 2005) implicated the
involvement of a channel that preferentially carried
monovalent cations and was inhibited by ruthenium red,
which is a non-selective ryanodine receptor antagonist
(Zucchi and Ronca-Testoni 1997). Interestingly, while
Xenopus oocytes do not express ryanodine receptors
(McPherson et al. 1992), it has been suggested that
ruthenium red can also target IP3 receptor (Ikeda and
Maruyama 2001). Similar responses were also recorded
when BmOR1 was co-expressed with HR2 and
DOR83b, indicating that this receptor family may, in
addition to its function in trafficking of general ORs,
also contribute to signal transduction in an enigmatic
way. One possibility is that a complex of one DOR83b
family protein and one conventional OR are required to
fully activate one heterotrimeric G protein. In fact, evi-
dence for such a mode of activation has been observed
for refolded leukotriene B4 receptor that was reconsti-
tuted in lipid vesicles along with G protein (Baneres and
Parello 2003).
Odorant binding proteins
An important implication of the studies involving ecto-
pic or heterologous expression of conventional ORs and
pheromone receptors is the suggestion that these pro-
teins can discriminate biologically relevant concentra-
tions of their cognate ligands and effectively initiate
olfactory signal transduction in the absence of OBPs or,
in more specialized ORNs, pheromone binding proteins
(PBPs). This comes despite a large body of literature
confirming that OBPs and PBPs are widely and robustly
expressed in insect olfactory organs and an equally large
variety of hypotheses concerning their function. These
include roles in removal or inactivation of odorants
(Steinbrecht and Müller 1971; Vogt and Riddiford 1981;
Ziegelberger 1995), solubilization of hydrophobic
odorants (Wojtasek and Leal 1999; Sandler et al. 2000),
concentration of odorants in the sensillum lymph Pelosi
(1996) by acting as filters to screen out subsets of
odorants (Pelosi 1994) or transportation of odorants
through the lymph to the olfactory neurons to possibly
act as co-initiators of the signal transduction process
(Krieger and Breer 1999). Furthermore, several OBP
and PBP structures have been resolved both alone and in
combination with various ligands (Tegoni et al. 2004).
In some cases, specific binding of biologically relevant
molecules has been observed for PBPs (Bette et al. 2002;
Mohl et al. 2002; Riviere et al. 2003) and another class
of molecules, the chemosensory proteins (CSPs), that
has been suggested to interact with odorants in the
sensillum lymph (Briand et al. 2002; Lartigue et al.
2002). On the other hand, others have found that PBPs
can bind a wide range of chemicals with high affinity
(Campanacci et al. 2001).
Not surprisingly, there is evidence that this class of
proteins may in fact carry out more than one of the
proposed set of OBP functions. For example, Drosophila
OBP19a, which was originally termed Pheromone
Binding Protein Related Protein (PBPRP) 2, has been
localized in two studies by immuno-electron microscopy
(Park et al. 2000; Shanbhag et al. 2001b) to diverse areas
of chemosensory organs of the fruitfly. In the antennae
and the maxillary palpi, PBPRP2 is localized to the
extracellular space that is situated between the cuticle
and the underlying epidermal cells (Park et al. 2000).
Additionally, PBPRP2 labeling was observed in a subset
of coeloconic sensilla on the antennal surface. These
sensilla are unique among olfactory hairs in that they are
double walled and contain two separate liquid-filled
cavities (Shanbhag et al. 1999; Park et al. 2000). Of
these, only the inner cavity is in contact with the den-
drite of the sensory neuron and PBPRP2 labeling was
restricted to the outer cavity. Therefore, it is unlikely
that PBPRP2 comes in contact with the dendrite and the
signal transduction machinery (Park et al. 2000).
Moreover, PBPRP2 was identified in more than 90% of
taste bristles on the labial palp and in most of the tarsal
taste bristles where its expression was confined to the
outer sensillum lymph and the lumen of the sensory hair,
neither of which is in contact with the dendrite of the
sensory cell. Shanbag et al. (2001b) suggested that all the
above localizations of OBP19a may indicate its function
in detoxification of odorants rather than in interactions
with elements of the signal transduction apparatus. In
contrast to these instances, where a role for OBP19a as a
chemical transporter from the sensillum surface to the
dendrite or as a receptor interaction partner can be ex-
cluded, the protein was found in close association with
the dendrites of the receptor cells in all taste pegs and
hence may function as a carrier of tastant molecules in
this system (Shanbhag et al. 2001b). Furthermore, while
the structure of taste pegs is certainly atypical for con-
tact chemoreceptor sensilla, a recent report has charac-
terized the expression of Drosophila Gr5a, a gustatory
receptor sensitive to trehalose, in a subset of taste pegs
suggesting these sensilla indeed have a specific chemo-
sensory function (Thorne et al. 2004).
Also confounding is a recent functional evidence
suggesting that at least one OBP can fulfill variable
functions in different neurons. Drosophila OBP76a,
which was initially named lush based on a mutant phe-
notype conferring insensitivity to high concentrations of
alcohols, is expressed in the sensillum lymph that sur-
rounds the dendrite in all three types of trichodic sen-
silla, T1, T2 and T3, on D. melanogaster antennae (Kim
et al. 1998). Of these, the T1 sensilla house neurons that
are narrowly tuned to 11-cis vaccenyl acetate (VA)
(Clyne et al. 1997) while T2 sensilla can be grouped into
two functional classes T2A and T2B, the former con-
taining neurons that are inhibited by 1-hexanol while the
latter are inhibited by high concentrations of ethanol
and butanol (Xu et al. 2005). Electrophysiological
recordings from T1 neurons typically show a spontane-
ous activity of 1 AP per second and upon application of
VA, wild-type T1 sensilla respond with a robust burst of
APs. Remarkably, in lush mutants this sensitivity to VA
is completely lost and the spontaneous activity drops
approximately 400-fold (Xu et al. 2005) which is con-
sistent with a role for OBP76a/ lush not only as a VA
carrier but also as a component of ORN signal trans-
duction. In one model proposed by Xu et al. (2005) to
describe this observation, OBP76a/ lush contributes to
the activation of a receptor on the dendritic membrane
of the sensory neuron and VA would serve to stabilize
an active conformation of OBP76a. This active confor-
mation could also manifest spontaneously albeit at a
significantly lower frequency, thereby explaining the
infrequent spontaneous APs in wild-type T1 neurons as
well as an almost lack of them in lush mutants. Fur-
thermore, in an elegant follow-up experiment, Xu et al.
(2005) were able to restore spontaneous activity to T1
neurons by directly infusing recombinant OBP76a/
LUSH into the sensillum lymph from the recording
pipette tip. This type of rescue effectively excludes a
developmental requirement of OBP76a in T1 neuron
maturation as the reason for loss of spontaneous APs.
In addition to OBP76a/LUSH, at least two other
OBPs are secreted into the lymph of T1 sensilla
(Shanbhag et al. 2001a), which are unusual in that they
only contain 1 neuron (Shanbhag et al. 1999). Since the
additional OBPs in the T1 lymph cannot compensate for
loss of spontaneous APs observed in lush mutants, they
are likely to function differently from the hypothesized
involvement of OBP76a in signal transduction. It is
possible that these roles may include activities such as
odorant detoxification or solubilization.
In contrast to T1 neurons, T2 neurons exhibited no
loss of spontaneous AP in lush mutants of D. melanog-
aster (Xu et al. 2005). Intriguingly, a more subtle effect
was observed in T2 sensilla that could explain the pre-
viously observed insensitivity of lush mutants to high
concentrations of alcohols (Kim et al. 1998). Upon
application of 100% ethanol or 10% butanol through
the air stream, the spontaneous rate of APs in T2B
neurons, which are housed in a subset of T2 sensilla, was
inhibited. Importantly, this inhibition was lost in lush
mutants (Xu et al. 2005), which again points to a specific
involvement of OBP76a/LUSH in signal transduction
and suggests that alcohols may confer an inhibitory
conformation upon OBP76a/LUSH.
A specific involvement of OBPs in OR activation is
furthermore indicated by experiments of Pophof (Pop-
hof 2002, 2004). Two PBPs that are expressed together
in one sensillum of the moth Antheraea polyphemus, each
preferentially interacts with a cognate pheromone in
binding assays (Maida et al. 2000). In agreement with
the exhibited specificity, pre-binding of a PBP and its
cognate pheromone and infusion into the lymph sur-
rounding the pheromone responsive sensilla elicit re-
sponses from only one of the two ORNs (Pophof 2002,
2004). When each pheromone was pre-bound to the
respective non-cognate PBP and infused into the sen-
sillum lymph, a response in one of the two neurons was
dependent on the cognate PBP but indifferent to the
pheromone (Pophof 2002).
Conclusions/current models of insect olfactory signal
transduction
In light of recent progress in deorphanization of insect
ORs in heterologous expression systems (Wetzel et al.
2001; Sakurai et al. 2004; Nakagawa et al. 2005;
Neuhaus et al. 2005), it is clear that these receptors
can function to a substantial degree in the absence of
their endogenous OBPs. Moreover, many insect ORs
can be expressed ectopically in the context of a foreign
OBP environment without significant impairment of
their function (Wang et al. 2003; Hallem et al. 2004b).
While comparing the novel results regarding lush by
Xu et al. (2005) with those obtained for insect ORs in
heterologous expression systems, we recognize two
possibilities that would account for these effects on
olfactory signal transduction that both embrace a set
of underlying basic elements. The first premise, now
firmly established, is that insect ORs are bona fide
receptors for a large range of odorant molecules and
are fully capable of interfacing with all the elements of
classical GPCR signal transduction pathways. Next,
the extreme conservation of Or83b family members
(Krieger et al. 2003; Pitts et al. 2004; Jones et al. 2005)
along with the elegant functional analyses carried out
in Drosophila (Larsson et al. 2004) strongly implicate
that these proteins play a central role in insect olfac-
tory signal transduction. Furthermore, the essential
roles for these proteins in maintaining functionality
through protein–protein interactions with conventional
ORs in heterologous expression systems (Nakagawa
et al. 2005; Neuhaus et al. 2005) provide a compelling
rationale for placing them at the heart of the trans-
duction mechanism. Indeed, we favor the role of Or83
proteins as scaffolding components that not only
facilitate trafficking and placement of conventional
ORs within the ORN plasma membrane but also help
to maintain interactions with arrestins, G proteins and
other signaling components (Fig. 2).
We consider it highly unlikely that membrane-bound
ORs are recognizing odorants in vivo that are freely in
solution in the sensillum lymph. In this regard we rec-
ognize studies wherein the in vivo half-life of the moth
pheromone bombykol was calculated to be approxi-
mately 4 min (Kasang and Kaissling 1972) as well as
others utilizing in vitro moth sensillar preparations
shown to contain highly active ODEs that were shown
to degrade or chemically modify physiologically relevant
concentrations of odorants within millisecond time-
frames (Vogt et al. 1985; Rybczynski et al. 1989). This
incongruity was recognized by Vogt et al. (1985) who
described significant protection of pheromone from
ODEs by OBPs thus indicating a role of OBPs as
essential odorant shelters within olfactory sensilla.
It is reasonable to imagine that these experiments
may indeed point to the primary function of both OBPs
and ODEs. The insect chemosensory system has evolved
to provide these organisms with vital information about
their chemical environment. This environment can con-
tain beneficial substances for the organism such as food
but may also contain harmful chemicals, which are
generally to be excluded by the integument. This barrier,
however, has to be selectively permeable in order to
develop a system for chemical perception. Therefore, an
odorant delivery system, which nonspecifically loaded
chemical cargo upon entry and transferred it to a specific
location where these chemicals could be evaluated, may
have evolved to minimize detrimental effects on cells
that are directly exposed to the environment. Further-
more, in order to afford protection from chemicals that
escaped the delivery system as well as for the removal of
odorants after presentation to the specific recognition
system (i.e. ORs), ODEs capable of metabolizing these
substances into less harmful forms are expressed in
chemosensory sensilla.
In the first model (Fig. 2, red odorant), OBP76a/
LUSH represents an exceptional case in which an OBP/
agonist complex is directly and specifically recognized by
a conventional OR. In contrast, the majority of insect
ORs either would not bind odorants in the context of an
OBP at all or, alternatively, could not discriminate be-
tween different OBPs, each with high affinity for broad
classes of odorants but with low specificity for any
particular odorant within their receptive range. In this
model, the OR still maintains a high degree of discrim-
Fig. 2 Hypothetical model incorporating recent insights about
molecular interactions in the lumen and at the dendritic membrane
of an insect ORN. Odorants entering through cuticular pores are
immediately loaded onto OBPs that transport chemicals to
conventional ORs (ORx) and also protect them from degradation
by ODEs (yellow). Transport of odorants is directed by a specific
OBP receptor that is either constituted by (1) the conventional OR
(interacting with the red odorant/ OBP) or (2) by a different
molecule (SNMP?; interacting with the black odorant/ blue OBP),
which may physically interact with the conventional and/or 83b
family OR. SNMPs are candidate molecules that may function as
OBP receptors. Conventional ORs physically interact with a highly
conserved 83b family OR which is expressed in a majority of
ORNs. OR83b family proteins facilitate trafficking of conventional
ORs to the dendritic membrane and may contribute to signal
transduction. A complex that consists of a conventional OR, an
Or83b family protein and possibly additional molecules, may be
required to fully activate a heterotrimeric G-protein. Little is
known about the signal transduction events and ion channels that
are involved in the generation of APs in insects. Possible G protein
effectors involve phospholipases such as PLCb
ination for its odorant, regardless of the OBP context in
which it is ‘‘presented.’’
In the second model (Fig. 2, black odorant), the re-
sults obtained for the lush mutant, the only OBP mutant
functionally characterized, are not exceptional but in
fact, represent the fundamental paradigm for OBP
function in insect olfactory signal transduction where
OBPs make a specific and requisite contribution. Fur-
thermore, in this model we hypothesize that conven-
tional ORs do not directly interact with OBPs and a
distinct OBP receptor exists in ORNs (Fig. 2, black
odorant). This OBP receptor may act in concert with the
conventional OR by directing OBPs to the sites of spe-
cific chemical evaluation. This hypothesis could be tes-
ted by genetic studies in Drosophila that combine
deletion or expression knockdown of each ab3-sensillum
OBP in a Dhalo background where the ‘‘empty’’ ab3a
ORN lacks any endogenous DORs but is able to sup-
port the ectopic functional expression of multiple DORs
and at least 2AgORs in a sensitivity range that closely
resembled sensitivity of the native ORN of these DORs
(Hallem et al. 2004a, 2004b). If any of the Dobp Dhalo
double mutants shows a similar lack of spontaneous APs
as has been described for the lush mutant in T1 trichoid
sensilla (Xu et al. 2005), thus differing from the Dhalo
phenotype, which still generates spontaneous APs de-
spite a lack of any functional OR (Dobritsa et al. 2003),
a discrete receptor for each odorant and OBP exists.
Here a signal transduction cascade that proceeds from
OBPs to conventional ORs through G protein interme-
diates would be inconsistent with two differing pheno-
types for the ab3 OBP and Dhalo mutants. It should be
noted, however, that APs in the empty ab3A neuron
occur as random bursts (Dobritsa et al. 2003), possibly
indicating a role of conventional ORs in synchronizing
spontaneous APs. In this context we note that DOR83b
mutants are also devoid of spontaneous APs (Larsson
et al. 2004), suggesting that DOR83b could act down-
stream of OBPs to generate spontaneous APs in ORNs
and could therefore constitute a general OBP receptor.
Alternatively, a role as a separate OBP receptor has
previously been suggested for another enigmatic family
of insect olfactory proteins, the Sensory neuron mem-
brane proteins (SNMPs) (Rogers et al. 1997, 2001;
Pophof 2002; Jacquin-Joly and Merlin 2004) (Fig. 2,
black odorant). Indeed a number of the expected char-
acteristics for hypothetical OBP receptors are fulfilled by
SNMPs. This protein family is related to the CD36
family of proteins (Rogers et al. 1997), which is known
to bind protein-ligands (Tandon et al. 1989; Abumrad
et al. 1993; Asch et al. 1993) and comprises 14 members
in D. melanogaster (Rogers et al. 2001); hence interac-
tions with a wide range of OBPs are conceivable. Fur-
thermore, some SNMPs are specifically expressed in the
dendrite and soma of ORNs (Rogers et al. 1997) and
their onset of expression during development parallels
expression of OBPs (Rogers et al. 1997, 2001). It is also
interesting to note that Snmp-1, the first characterized
member of this protein family, was only localized in one
neuron of each sensillum where it was expressed (Rogers
et al. 1997). This could indicate a specific role in sorting
a subset of the OBP content of each sensillum to only
one neuron. In the only experiment that examined the
function of SNMPs thus far, a photoaffinity analog of a
moth pheromone was utilized to label sensillar extracts
of B. mori (Vogt et al. 1988). The subsequent identifi-
cation of an SNMP as one of the labeled proteins pro-
vided evidence for binding of an odorant to these
molecules (Rogers et al. 1997). Nevertheless, it seems
likely that careful analyses of Drosophila SNMP mutants
will be needed to gain a better understanding of a pos-
sible SNMP contribution in olfactory signal transduc-
tion. Given the large number of SNMP family members
in the Drosophila genome, these will not be straightfor-
ward experiments.
Another important question is whether such a model
could agree with the in vivo observations of Hallem
et al. (2004a, 2004b) who showed that many DORs and
indeed AgORs can function in alternative receptor
neurons and in many cases, retain their native odorant
response profile. In this instance it is possible that the
OBP receptor discussed above is present in the ‘‘empty’’
ab3A neuron and hence could still interact stochastically
with its cognate OBP to generate spontaneous APs as
suggested by Xu et al. (2005). If the function of such a
receptor is merely to attract and dock the OBPs, which
are required in vivo to transport odorants in close
proximity to conventional ORs, then the binding spec-
trum of the endogenous ab3 sensillum OBPs may define
as to which odorants can be presented to ectopically
expressed receptors. In this context, studies carried out
with individual OBPs showing a varying degree of bona
fide odorant binding capacity (Campanacci et al. 2001;
Bette et al. 2002; Mohl et al. 2002; Riviere et al. 2003) as
well as the variable OBP composition in different types
of sensilla (Pikielny et al. 1994) would effectively lead to
preselection of odorants that differentiate between those
that could be transported through the sensillum lymph
from those that cannot. As it happens, the ab3 sensillum
may be particularly favorable for ectopic or expression
because it contains a set of endogenous OBPs with a
broad range of odorant specificities. This, in turn,
should lead to a subset of ORs that would be refractory
to study in the empty neuron system. Indeed, it should
be noted that the study of Hallem et al. (2004a) and
Kreher et al. (2005) that used the ‘‘empty neuron’’ sys-
tem employed a limited panel of odorant ligands, and
more importantly consistently observed that a small
subset of DORs tested did not respond to any odorants
in these studies.
Hallem et al. (2004a) demonstrated that some
ectopically expressed DORs conferred sensitivity to 10–6
dilutions of some cognate odorants to the ab3A neuron
and compared those odorant response profiles to the
respective native neurons after stimulation with odor-
ants at 10–2 dilution. It would be interesting to see if the
odorant response profiles conferred by ectopic ORs in
the ab3A neuron and the response profiles of their native
ORNs are as similar at lower odorant concentrations.
Increased odorant sensitivity in the native neurons could
reflect the contribution of the OBP makeup of the sen-
sillum lymph. Regardless of the particulars, it is likely
that the response profile of each individual ORN reflects
a diverse spectrum of olfactory components, each of
which contributing towards the robust sensitivity and
output characteristics of these remarkable chemosensory
units.
Acknowledgements We wish to especially thank Dr. Jonathan
Bohbot for expert illustrations as well as thank Dr. H.W. Kwon,
Jason Pitts and H. Wegner for critical reading of the manuscript.
This work received financial support from the NIH (DC04692/
AI56402 to L.J.Z) and through a Max Kade Post-Doctoral Re-
search Exchange grant (to M. R.).
References
Abumrad NA, el-Maghrabi MR, Amri EZ, Lopez E, Grimaldi PA
(1993) Cloning of a rat adipocyte membrane protein implicated
in binding or transport of long-chain fatty acids that is induced
during preadipocyte differentiation. Homology with human
CD36. J Biol Chem 268:17665–17668
Adams MD, Celniker SE, Holt RA, Evans CA, Gocayne JD,
Amanatides PG, Scherer SE, Li PW, Hoskins RA, Galle RF,
George RA, Lewis SE, Richards S, Ashburner M, Henderson
SN, Sutton GG, Wortman JR, Yandell MD, Zhang Q, Chen
LX, Brandon RC, Rogers YH, Blazej RG, Champe M, Pfeiffer
BD, Wan KH, Doyle C, Baxter EG, Helt G, Nelson CR, Gabor
GL, Abril JF, Agbayani A, An HJ, Andrews-Pfannkoch C,
Baldwin D, Ballew RM, Basu A, Baxendale J, Bayraktaroglu L,
 Beasley EM, Beeson KY, Benos PV, Berman BP, Bhandari D,
Bolshakov S, Borkova D, Botchan MR, Bouck J, Brokstein P,
Brottier P, Burtis KC, Busam DA, Butler H, Cadieu E, Center
A, Chandra I, Cherry JM, Cawley S, Dahlke C, Davenport LB,
Davies P, de Pablos B, Delcher A, Deng Z, Mays AD, Dew I,
Dietz SM, Dodson K, Doup LE, Downes M, Dugan-Rocha S,
Dunkov BC, Dunn P, Durbin KJ, Evangelista CC, Ferraz C,
Ferriera S, Fleischmann W, Fosler C, Gabrielian AE, Garg NS,
Gelbart WM, Glasser K, Glodek A, Gong F, Gorrell JH, Gu Z,
Guan P, Harris M, Harris NL, Harvey D, Heiman TJ, Her-
nandez JR, Houck J, Hostin D, Houston KA, Howland TJ,
Wei MH, Ibegwam C et al (2000) The genome sequence of
Drosophila melanogaster. Science 287:2185–195
Alloway PG, Howard L and Dolph PJ (2000) The formation of
stable rhodopsin-arrestin complexes induces apoptosis and
photoreceptor cell degeneration. Neuron 28:129–138
Antonny B, Sukumar M, Bigay J, Chabre M and Higashijima T
(1993) The mechanism of aluminum-independent G-protein
activation by fluoride and magnesium. 31P NMR spectroscopy
and fluorescence kinetic studies. J Biol Chem 268:2393–2402
Asch AS, Liu I, Briccetti FM, Barnwell JW, Kwakye-Berko F,
Dokun A, Goldberger J and Pernambuco M (1993) Analysis of
CD36 binding domains: ligand specificity controlled by
dephosphorylation of an ectodomain. Science 262:1436–1440
Baneres JL and Parello J (2003) Structure-based analysis of GPCR
function: evidence for a novel pentameric assembly between the
dimeric leukotriene B4 receptor BLT1 and the G-protein. J Mol
Biol 329:815–829
Baumann A, Frings S, Godde M, Seifert R and Kaupp UB (1994)
Primary structure and functional expression of a Drosophila
cyclic nucleotide-gated channel present in eyes and antennae.
EMBO Journal 13:5040–5050
Bette S, Breer H and Krieger J (2002) Probing a pheromone
binding protein of the silkmoth Antheraea polyphemus by
endogenous tryptophan fluorescence. Insect Biochem Mol Biol
32:241–246
Biessmann H, Walter MF, Dimitratos S and Woods D (2002)
Isolation of cDNA clones encoding putative odorant binding
proteins from the antennae of the malaria-transmitting mos-
quito, Anopheles gambiae. Insect Mol Biol 11:123–132
Boeckh J, Boeckh V (1979) Threshold and odor specificity of
pheromone-sensitive nerurons in the deutocerebrum of Anthe-
raea pernyi and A. polyphemus (Saturnidae). J Comp Physiol
132:235–242
Boekhoff I, Raming K, Breer H (1990) Pheromone-induced stim-
ulation of inositol-trisphosphate formation in insect antennae is
mediated by G-proteins. J Comp Physiol B 160:99–103
Boekhoff I, Michel WC, Breer H, Ache BW (1994) Single odors
differentially stimulate dual second messenger pathways in
lobster olfactory receptor cells. J Neurosci 14:3304–3309
Brand AH, Perrimon N (1993) Targeted gene expression as a
means of altering cell fates and generating dominant pheno-
types. Development 118:401–415
Breer H, Boekhoff I, Tareilus E (1990) Rapid kinetics of second
messenger formation in olfactory transduction. Nature 345:65–
68
Briand L, Nespoulous C, Huet JC, Takahashi M, Pernollet JC
(2001) Ligand binding and physico-chemical properties of
ASP2, a recombinant odorant-binding protein from honeybee
(Apis mellifera L.). Eur J Biochem 268:752–760
Briand L, Swasdipan N, Nespoulous C, Bezirard V, Blon F, Huet
JC, Ebert P, Penollet JC (2002) Characterization of a chemo-
sensory protein (ASP3c) from honeybee (Apis mellifera L.) as a
brood pheromone carrier. Eur J Biochem 269:4586–4596
de Bruyne M, Clyne PJ, Carlson JR (1999) Odor coding in a model
olfactory organ: the Drosophila maxillary palp. J Neurosci
19:4520–4532
de Bruyne M, Foster K, Carlson J (2001) Odor coding in the
Drosophila antenna. Neuron 30:537–552
Buchner E (1991) Genes expressed in the adult brain of Drosophila
and effects of their mutations on behavior: a survey of transmit-
ter- and second messenger-related genes. J Neurogenet 7:153–192
Buck L, Axel R (1991) A novel multigene family may encode
odorant receptors: a molecular basis for odor recognition. Cell
65:175–187
Bunemann M, Frank M, Lohse MJ (2003) Gi protein activation in
intact cells involves subunit rearrangement rather than disso-
ciation. Proc Natl Acad Sci U S A 100:16077–16082
Campanacci V, Krieger J, Bette S, Sturgis JN, Lartigue A, Cam-
billau C, Breer H, Tegoni M (2001) Revisiting the specificity of
Mamestra brassicae and Antheraea polyphemus pheromone-
binding proteins with a fluorescence binding assay. J Biol Chem
276:20078–20084
Christensen TA, Pawlowski VM, Lei H, Hildebrand JG (2000)
Multi-unit recordings reveal context-dependent modulation of
synchrony in odor-specific neural ensembles. Nat Neurosci
3:927–931
Clyne P, Grant A, O’Connell R, Carlson JR (1997) Odorant re-
sponse of individual sensilla on the Drosophila antenna. Invert
Neurosci 3:127–135
Clyne PJ, Warr CG, Freeman MR, Lessing D, Kim J, Carlson JR
(1999) A novel family of divergent seven-transmembrane pro-
teins: candidate odorant receptors in Drosophila. Neuron
22:327–338
Cork A, Park KC (1996) Identification of electrophysiological-ac-
tive compounds for the malaria mosquito, Anopheles gambiae,
in human sweat extracts. Med Veterinary Entomol 10:269–276
Dear TN, Boehm T, Keverne EB, Rabbitts TH (1991) Novel genes
for potential ligand-binding proteins in subregions of the
olfactory mucosa. EMBO J 10:2813–2819
Dickens JC, Callahan FE, Wergin WP, Murphy CA, Vogt RG
(1998) Odorant-binding proteins of true bugs. Generic speci-
ficity, sexual dimorphism, and association with subsets of
chemosensory sensilla. Ann N Y Acad Sci 855:306–310
Dobritsa AA, van der Goes van Naters W, Warr CG, Steinbrecht
RA, Carlson JR (2003) Integrating the molecular and cellular
basis of odor coding in the Drosophila antenna. Neuron 37:827–
841
Dubin AE, Liles MM, Seligman F, Le T, Tolli J, Harris GL (1998)
Involvement of genes encoding a K+ channel (ether a go-go)
and a Na+ channel (smellblind) in Drosophila olfaction. Ann
NY Acad Sci 855:212–222
Dwyer ND, Troemel ER, Sengupta P, Bargmann CI (1998)
Odorant receptor localization to olfactory cilia is mediated by
ODR-4, a novel membrane-associated protein. Cell 93:455–466
Fadool DA, Ache BW (1992) Plasma membrane inositol 1,4,5-
trisphosphate-activated channels mediate signal transduction in
lobster olfactory receptor neurons. Neuron 9:907–918
Fiala A, Spall T, Diegelmann S, Eisermann B, Sachse S, Devaud
JM, Buchner E, Galizia CG (2002) Genetically expressed ca-
meleon in Drosophila melanogaster is used to visualize olfactory
information in projection neurons. Curr Biol 12:1877–1884
Firestein S (2001) How the olfactory system makes sense of scents.
Nature 413:211–218
Flower DR (1996) The lipocalin protein family: structure and
function. Biochem J 318 (Pt 1):1–14
Flower DR, North AC, Attwood TK (1991) Mouse oncogene
protein 24p3 is a member of the lipocalin protein family. Bio-
chem Biophys Res Commun 180:69–74
Flower DR, North AC, Sansom CE (2000) The lipocalin protein
family: structural and sequence overview. Biochim Biophys
Acta 1482:9–24
Fox AN, Pitts RJ, Robertson HM, Carlson JR, Zwiebel LJ (2001)
Candidate odorant receptors from the malaria vector mosquito
Anopheles gambiae and evidence of down-regulation in response
to blood feeding. Proc Natl Acad Sci U S A 98:14693–14697
Fox AN, Pitts RJ, Zwiebel LJ (2002) A cluster of candidate
odorant receptors from the malaria vector mosquito, Anopheles
gambiae. Chem Senses 27:453–459
Freedman NJ, Lefkowitz RJ (1996) Desensitization of G protein-
coupled receptors. Recent Prog Horm Res 51:319–353
Fuentes-Prior P, Noeske-Jungblut C, Donner P, Schleuning WD,
Huber R, Bode W (1997) Structure of the thrombin complex
with triabin, a lipocalin-like exosite-binding inhibitor derived
 from a triatomine bug. Proc Natl Acad Sci U S A 94:11845–
11850
Gao Q, Chess A (1999) Identification of candidate Drosophila
olfactory receptors from genomic DNA sequence. Genomics
60:31–39
Gimelbrant AA, Stoss TD, Landers TM, McClintock TS (1999)
Truncation releases olfactory receptors from the endoplasmic
reticulum of heterologous cells. J Neurochem 72:2301–2311
Gimelbrant AA, Haley SL, McClintock TS (2001) Olfactory
receptor trafficking involves conserved regulatory steps. J Biol
Chem 276:7285–7290
Goldman AL, Van der Goes van Naters W, Lessing D, Warr CG,
Carlson JR (2005) Coexpression of two functional odor
receptors in one neuron. Neuron 45:661–666
Gong WJ, Golic KG (2003) Ends-out, or replacement, gene tar-
geting in Drosophila. Proc Natl Acad Sci U S A 100:2556–2561
Hague C, Uberti MA, Chen Z, Bush CF, Jones SV, Ressler KJ,
Hall RA, Minneman KP (2004) Olfactory receptor surface
expression is driven by association with the beta2-adrenergic
receptor. Proc Natl Acad Sci U S A 101:13672–13676
Hallem E, Fox AN, Zwiebel LJ, Carlson JR (2004a) A mosquito
odorant receptor tuned to a component of human sweat. Nat-
ure 427:212–213
Hallem E, Ho MG, Carlson JR (2004b) The molecular basis of
odor coding in the Drosophila antenna. Cell 117:965–979
Hamm HE (1998) The many faces of G protein signaling. J Biol
Chem 273:669–672
Hildebrand JG, Shepherd GM (1997) Mechanisms of olfactory
discrimination: converging evidence for common principles
across phyla. Annu Rev Neurosci 20:595–631
Hill CA, Fox AN, Pitts RJ, Kent LB, Tan PL, Chrystal MA,
Cravchik A, Collins FH, Robertson HM, Zwiebel LJ (2002) G
protein-coupled receptors in Anopheles gambiae. Science
298:176–178
Hyde DR, Mecklenburg KL, Pollock JA, Vihtelic TS, Benzer S
(1990) Twenty Drosophila visual system cDNA clones: one is a
homolog of human arrestin. Proc Natl Acad Sci U S A 87:1008–
1012
Ikeda M, Maruyama Y (2001) Inhibitory effects of ruthenium red
on inositol 1,4, 5-trisphosphate-induced responses in rat
megakaryocytes. Biochem Pharmacol 61:7–13
Ishida Y, Leal WS (2002) Cloning of putative odorant-degrading
enzyme and integumental esterase cDNAs from the wild silk-
moth, Antheraea polyphemus. Insect Biochem Mol Biol
32:1775–1780
Ishida Y, Cornel AJ, Leal WS (2002) Identification and cloning of a
female antenna-specific odorant-binding protein in the mos-
quito Culex quinquefasciatus. J Chem Ecol 28:867–871
Ishida Y, Chen AM, Tsuruda JM, Cornel AJ, Debboun M, Leal
WS (2004) Intriguing olfactory proteins from the yellow fever
mosquito, Aedes aegypti. Naturwissenschaften 91:426–431
Jackson FR, Newby LM, Kulkarni SJ (1990) Drosophila GAB-
Aergic systems: sequence and expression of glutamic acid
decarboxylase. J Neurochem 54:1068–1078
Jacquin-Joly E, Merlin C (2004) Insect olfactory receptors: con-
tributions of molecular biology to chemical ecology. J Chem
Ecol 30:2359–2397
Janetopoulos C, Jin T, Devreotes P (2001) Receptor-mediated
activation of heterotrimeric G-proteins in living cells. Science
291:2408–2811
Jones DT, Reed RR (1989) Golf: an olfactory neuron specific-G
protein involved in odorant signal transduction. Science
244:790–795
Jones WD, Nguyen TA, Kloss B, Lee KJ, Vosshall LB (2005)
Functional conservation of an insect odorant receptor gene
across 250 million years of evolution. Curr Biol 15:R119–R121
Kaissling KE (1996) Peripheral mechanisms of pheromone recep-
tion in moths. Chem Senses 21:257–268
Kasang G, Kaissling KE (1972) Kinetic studies of transduction in
olfactory receptors of Bombyx mori. In: Schneider D (eds)
International symposia on olfaction and taste. Vol. 4, Ver-
lagsgesellschaft, F. R. G. Stuttgart, pp 200–206
Kim MS, Repp A, Smith DP (1998) LUSH odorant-binding pro-
tein mediates chemosensory responses to alcohols in Drosophila
melanogaster. Genetics 150:711–721
Kim J, Moriyama EN, Warr CG, Clyne PJ, Carlson JR (2000)
Identification of novel multi-transmembrane proteins from
genomic databases using quasi-periodic structural properties.
Bioinformatics 16:767–775
Kreher SA, Kwon JY, Carlson JR (2005) The molecular basis of
odor coding in the Drosophila larva. Neuron 46:445–456
Krieger J, Breer H (1999) Olfactory reception in invertebrates.
Science 286:720–723
Krieger J, Raming K, Dewer YM, Bette S, Conzelmann S and
Breer H (2002) A divergent gene family encoding candidate
olfactory receptors of the moth Heliothis virescens. Eur J
Neurosci 16:619–628
Krieger J, Klink O, Mohl C, Raming K, Breer H (2003) A candi-
date olfactory receptor subtype highly conserved across differ-
ent insect orders. J Comp Physiol A Neuroethol Sens Neural
Behav Physiol 189:519–526
Krieger J, Grosse-Wilde E, Gohl T, Dewer YM, Raming K, Breer
H (2004) Genes encoding candidate pheromone receptors in a
moth (Heliothis virescens). Proc Natl Acad Sci U S A
101:11845–11850
Larsson MC, Domingos AI, Jones WD, Chiappe ME, Amrein H,
Vosshall LB (2004) Or83b encodes a broadly expressed odorant
receptor essential for Drosophila olfaction. Neuron 43:703–714
Lartigue A, Campanacci V, Roussel A, Larsson AM, Jones TA,
Tegoni M, Cambillau C (2002) X-ray structure and ligand
binding study of a moth chemosensory protein. J Biol Chem
277:32094–32098
Laue M, Maida R, Redkozubov A (1997) G-protein activation,
identification and immunolocalization in pheromone-sensitive
sensilla trichodea of moths. Cell Tissue Res 288:149–158
Laurent G (1999) A systems perspective on early olfactory coding.
Science 286:723–728
Lei H, Christensen TA, Hildebrand JG (2004) Spatial and temporal
organization of ensemble representations for different odor
classes in the moth antennal lobe. J Neurosci 24:11108–11119
Maibeche-Coisne M, Merlin C, Francois MC, Queguiner I,
Porcheron P, Jacquin-Joly E (2004) Putative odorant-degrading
esterase cDNA from the moth Mamestra brassicae: cloning and
expression patterns in male and female antennae. Chem Senses
29:381–390
Maida R, Krieger J, Gebauer T, Lange U, Ziegelberger G (2000)
Three pheromone-binding proteins in olfactory sensilla of the
two silkmoth species Antheraea polyphemus and Antheraea
pernyi. Eur J Biochem 267:2899–2908
Malnic B, Hirono J, Sato T, Buck LB (1999) Combinatorial
receptor codes for odors. Cell 96:713–723
Markby DW, Onrust R, Bourne HR (1993) Separate GTP binding
and GTPase activating domains of a G alpha subunit. Science
262:1895–1901
McClintock TS, Landers TS, Gimelbrandt AA, Fuller LZ, Jackson
BA, Jayawickreme CK, Lerner MR (1997) Functional expres-
sion of olfactory-adrenergic receptor chimeras and intracellular
retention of heterologously expressed olfactory receptors. Mol
Brain Res 48:270–278
McCudden CR, Hains MD, Kimple RJ, Siderovski DP, Willard FS
(2005) G-protein signaling: back to the future. Cell Mol Life Sci
62:551–577
McKenna MP, Hekmat Scafe DS, Gaines P, Carlson JR (1994)
Putative Drosophila pheromone-binding proteins expressed in a
subregion of the olfactory system. J Biol Chem 269:16340–
16347
McPherson SM, McPherson PS, Mathews L, Campbell KP, Longo
FJ (1992) Cortical localization of a calcium release channel in
sea urchin eggs. J Cell Biol 116:1111–1121
Melo AC, Rutzler M, Pitts RJ, Zwiebel LJ (2004) Identification of
a chemosensory receptor from the yellow fever mosquito, Aedes
aegypti, that is highly conserved and expressed in olfactory and
gustatory organs. Chem Senses 29:403–410
Merrill CE, Riesgo-Escovar J, Pitts RJ, Kafatos FC, Carlson JR,
Zwiebel LJ (2002) Visual arrestins in olfactory pathways of
Drosophila and the malaria vector mosquito Anopheles gambiae.
Proc Natl Acad Sci U S A 99:1633–1638
Merrill CE, Pitts RJ, Zwiebel LJ (2003) Molecular characterization
of arrestin family members in the malaria vector mosquito,
Anopheles gambiae. Insect Mol Biol 12:641–650
Merrill CE, Sherertz T, Walker W, Zwiebel LJ (2005) Odorant
specific requirements for arrestin function in Drosophila olfac-
tion. J Neurobiol 63(1):15–28
Michel WC, Ache BW (1992) Cyclic nucleotides mediate an odor-
evoked potassium conductance in lobster olfactory receptor
cells. J Neurosci 12:3979–3984
Mohl C, Breer H, Krieger J (2002) Species-specific pheromonal
compounds induce distinct conformational changes of phero-
mone binding protein subtypes from Antheraea polyphemus.
Invert Neurosci 4:165–174
Nakagawa T, Sakurai T, Nishioka T, Touhara K (2005) Insect sex-
pheromone signals mediated by specific combinations of
olfactory receptors. Science 307:1638–1642
Neuhaus EM, Gisselmann G, Zhang W, Dooley R, Stortkuhl K,
Hatt H (2005) Odorant receptor heterodimerization in the
olfactory system of Drosophila melanogaster. Nat Neurosci
8:15–17
Ng M, Roorda RD, Lima SQ, Zemelman BV, Morcillo P, Mie-
senbock G (2002) Transmission of olfactory information be-
tween three populations of neurons in the antennal lobe of the
fly. Neuron 36:463–474
Park SK, Shanbhag SR, Wang Q, Hasan G, Steinbrecht RA,
Pikielny CW (2000) Expression patterns of two putative odor-
ant-binding proteins in the olfactory organs of Drosophila
melanogaster have different implications for their functions.
Cell Tissue Res 300:181–192
Pelosi P (1994) Odorant-binding proteins. Crit Rev Biochem Mol
Biol 29:199–228
Pelosi P (1996) Perireceptor events in olfaction. J Neurobiol 30:3–19
Pelosi P, Maida R (1995) Odorant-binding proteins in insects.
Comp Biochem Physiol B Biochem Mol Biol 111:503–514
Pikielny CW, Hasan G, Rouyer F, Rosbash M (1994) Members of
a family of Drosophila putative odorant-binding proteins
are expressed in different subsets of olfactory hairs. Neuron
12:35–49
Pippig S, Andexinger S, Daniel K, Puzicha M, Caron MG, Lef-
kowitz RJ, Lohse MJ (1993) Overexpression of beta-arrestin
and beta-adrenergic receptor kinase augment desensitization of
beta 2-adrenergic receptors. J Biol Chem 268:3201–3208
Pitts RJ, Fox AN, Zwiebel LJ (2004) A highly conserved candidate
chemoreceptor expressed in both olfactory and gustatory tis-
sues in the malaria vector, Anopheles gambiae. Proc Natl Acad
Sci U S A 101:5058–5063
Pophof B (2002) Moth pheromone binding proteins contribute to
the excitation of olfactory receptor cells. Naturwissenschaften
89:515–518
Pophof B (2004) Pheromone-binding proteins contribute to the
activation of olfactory receptor neurons in the silkmoths An-
theraea polyphemus and Bombyx mori. Chem Senses 29:117–125
Reddy PS, Corley RB (1998) Assembly, sorting, and exit of olig-
omeric proteins from the endoplasmic reticulum. Bioessays
20:546–554
Ressler KJ, Sullivan SL, Buck LB (1993) A zonal organization of
odorant receptor gene expression in the olfactory epithelium.
Cell 73:597–609
Riviere S, Lartigue A, Quennedey B, Campanacci V, Farine JP,
Tegoni M, Cambillau C, Brossut R (2003) A pheromone-
binding protein from the cockroach Leucophaea maderae:
cloning, expression and pheromone binding. Biochem J
371:573–579
Robertson HM, Warr CG, Carlson JR (2003) Molecular evolution
of the insect chemoreceptor gene superfamily in Drosophila
melanogaster. Proc Natl Acad Sci U S A 100 Suppl 2:14537–
14542
Rogers ME, Sun M, Lerner MR, Vogt RG (1997) Snmp-1, a novel
membrane protein of olfactory neurons of the silk moth An-
theraea polyphemus with homology to the CD36 family of
membrane proteins. J Biol Chem 272:14792–14799
Rogers ME, Krieger J, Vogt RG (2001) Antennal SNMPs (sensory
neuron membrane proteins) of Lepidoptera define a unique
family of invertebrate CD36-like proteins. J Neurobiol 49:47–61
Rybczynski R, Reagan J, Lerner MR (1989) A pheromone-
degrading aldehyde oxidase in the antennae of the moth
Manduca sexta. J Neurosci 9:1341–1353
Sakurai T, Nakagawa T, Mitsuno H, Mori H, Endo Y, Tanoue S,
Yasukochi Y, Touhara K, Nishioka T (2004) Identification and
functional characterization of a sex pheromone receptor in the
silkmoth Bombyx mori. Proc Natl Acad Sci U S A 101:16653–
16658
Sandler BH, Nikonova L, Leal WS, Clardy J (2000) Sexual
attraction in the silkworm moth: structure of the pheromone-
binding-protein-bombykol complex. Chem Biol 7:143–151
Shanbhag SR, Müller B, Steinbrech RA (1999) Atlas of olfactory
organs of Drosophila melanogaster 1.Types, external organiza-
tion, innervation and distribution of olfactory sensilla. Int J
Insect Morphol Embryol 28:377–397
Shanbhag SR, Hekmat-Scafe D, Kim MS, Park SK, Carlson JR,
Pikielny C, Smith DP, Steinbrecht RA (2001a) Expression
mosaic of odorant-binding proteins in Drosophila olfactory
organs. Microsc Res Tech 55:297–306
Shanbhag SR, Park SK, Pikielny CW, Steinbrecht RA (2001b)
Gustatory organs of Drosophila melanogaster: fine structure and
expression of the putative odorant-binding protein PBPRP2.
Cell Tissue Res 304:423–437
Shields VD, Hildebrand JG (2001) Recent advances in insect
olfaction, specifically regarding the morphology and sensory
physiology of antennal sensilla of the female sphinx moth
Manduca sexta. Microsc Res Tech 55:307–329
Sinnarajah S, Ezeh PI, Pathirana S, Moss AG, Morrison EE,
Vodyanoy V (1998) Inhibition and enhancement of odorant-
induced cAMP accumulation in rat olfactory cilia by antibodies
directed against Gas/olf- and Gai-protein subunits. FEBS Lett
426:377–380
Smith DP (1999) Drosophila odor receptors revealed. Neuron
22:203–204
Smith DP, Shieh BH, Zuker CS (1990) Isolation and structure of
an arrestin gene from Drosophila. Proc Natl Acad Sci U S A
87:1003–1007
Steinbrecht RA, Müller B (1971) On the stimulus conducting
structures in insect olfactory receptors. Z Zellforsch Mikrosk
Anat 117:570–575
Stengl M (1994) Inositol-trisphosphate-dependent calcium currents
precede cation currents in insect olfactory receptor neurons
in vitro. J Comp Physiol [A] 174:187–194
Stortkuhl KF, Kettler R (2001) Functional analysis of an olfactory
receptor in Drosophila melanogater. Proc Natl Acad Sci U S A
98:9381–9385
Tandon NN, Lipsky RH, Burgess WH, Jamieson GA (1989) Iso-
lation and characterization of platelet glycoprotein IV (CD36).
J Biol Chem 264:7570–7575
Tegoni M, Campanacci V, Cambillau C (2004) Structural aspects
of sexual attraction and chemical communication in insects.
Trends Biochem Sci 29:257–264
Terrillon S, Bouvier M (2004) Roles of G-protein-coupled receptor
dimerization. EMBO Rep 5:30–34
Thorne N, Chromey C, Bray S, Amrein H (2004) Taste perception
and coding in Drosophila. Curr Biol 14:1065–1079
Vassar R, Ngai J, Axel R (1993) Spatial segregation of odorant
receptor expression in the mammalian olfactory epithelium.
Cell 74:309–318
Vogt RG, Riddiford LM (1981) Pheromone binding and inacti-
vation by moth antennae. Nature 293:161–163
Vogt RG, Riddiford LM, Prestwich GD (1985) Kinetic properties
of a sex pheromone-degrading enzyme: the sensillar esterase of
Antheraea polyphemus. Proc Natl Acad Sci U S A 82:8827–8831
Vogt RG, Prestwich GD, Riddiford LM (1988) Sex pheromone
receptor proteins. Visualization using a radiolabeled photoaf-
finity analog. J Biol Chem 263:3952–3959
Vogt RG, Rybczynski R, Lerner MR (1991) Molecular cloning
and sequencing of general odorant-binding proteins GOBP1
and GOBP2 from the tobacco hawk moth Manduca sexta:
comparisons with other insect OBPs and their signal peptides.
J Neurosci 11:2972–2984
Vosshall LB, Amrein H, Morozov PS, Rzhetsky A, Axel R (1999)
A spatial map of olfactory receptor expression in the Drosophila
antenna. Cell 96:725–736
Wang Y, Wright NJ, Guo H, Xie Z, Svoboda K, Malinow R,
Smith DP, Zhong Y (2001) Genetic manipulation of the
odor-evoked distributed neural activity in the Drosophila
mushroom body. Neuron 29:267–276
Wang JW, Wong AM, Flores J, Vosshall LB, Axel R (2003) Two-
photon calcium imaging reveals an odor-evoked map of activity
in the fly brain. Cell 112:271–282
Wang Y, Guo HF, Pologruto TA, Hannan F, Hakker I, Svo-
boda K, Zhong Y (2004) Stereotyped odor-evoked activity in
the mushroom body of Drosophila revealed by green fluo-
rescent protein-based Ca2+ imaging. J Neurosci 24:6507–
6514
Wetzel CH, Behrendt H, Gisselmann G, Storkuhl KF, Hovemann
B, Hatt H (2001) Functional expression and characterization of
a Drosophila odorant receptor in a heterologous cell system.
PNAS 98:9377–9380
Wojtasek H, Leal WS (1999) Conformational change in the pher-
omone-binding protein from Bombyx mori induced by pH and
by interaction with membranes. J Biol Chem 274:30950–30956
Xu Y, Piston DW, Johnson CH (1999) A bioluminescence reso-
nance energy transfer (BRET) system: application to interacting
circadian clock proteins. Proc Natl Acad Sci U S A 96:151–156
Xu PX, Zwiebel LJ and Smith DP (2003) Identification of a distinct
family of genes encoding atypical odorant-binding proteins in
the malaria vector mosquito, Anopheles gambiae. Insect Mol
Biol 12:549–560
Xu P, Atkinson R, Jones DN, Smith DP (2005) Drosophila OBP
LUSH is required for activity of pheromone-sensitive neurons.
Neuron 45:193–200
Ziegelberger G (1995) Redox-shift of the pheromone-binding pro-
tein in the silkmoth Antheraea polyphemus. Eur J Biochem
232:706–711
Zucchi R, Ronca-Testoni S (1997) The sarcoplasmic reticulum
Ca2+ channel/ryanodine receptor: modulation by endogenous
effectors, drugs and disease states. Pharmacol Rev 49:1–51