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Development 1994 Supplement, 209-215 (1994)
Printed in Great Britain @ The Company of Biologists Limited 1994
209
The evolving role of Hox genes in arthropods
Michael Akam, Michalis Averof, James Castelli-Gair, Rachel Daw€s, Francesco Falciani and David Ferrier
Wellcome/CRC lnstitute and Department of Genetics, Tennis Court Road, Cambridge, CB2 1QR, UK
SUMMARY
Comparisons between Hox genes in different arthropods
suggest that the diversity of Antennapedia-class homeotic
genes present in modern insects had already arisen before
the divergence of insects and crustaceans, probably during
the Cambrian. Hox gene duplications are therefore
unlikely to have occurred concomitantly with trunk
segment diversification in the lineage leading to insects.
Available data suggest that domains of homeotic gene
expression are also generally conserved among insects, but
changes in Hox gene regulation may have played a significant role in segment diversification. Differences that have
been documented alter specific aspects of Hox gene reguIation within segments and correlate with alterations in
segment morphology rather than overt homeotic transformations.
The Drosophila Hox cluster contains several homeobox
genes that are not homeotic genes - bicoidrfushi-tnrazu and
INTRODUCTION
In Drosophila, the differences between the trunk segments are
controlled by the homeotic genes of the Antennapedia and
Bithorax gene complexes (Lewis, 1978; S6nchez-Herrero et
al., 1985; Kaufman et a1., 1990). This paper is concerned with
the extent to which changes affecting these Hox genes may
have contributed to the evolution of body form and developmental mechanism in the arthropods, and in particular the
insects. We begin by considering the timing of gene duplications that gave rise to the Insect Hox cluster*, in relation to the
origin of invertebrate phyla and arthropod classes. Then we
consider how changes in the regulation of Hox genes may have
contributed to the diversification of the insects. Finally, we
present evidence that the selective constraints on some genes
in the Hox clusters appear to have been relaxed in some insect
lineages, allowing them to diverge relatively rapidly, both in
terms of sequence and role in development.
*Here we shall use the term Hox genes to refer generically to the homeobox genes
contained within the arthropod homeotic gene clusters, as well as their vertebrate homologues. The term Hom/Hox, never euphonic, is now redundant. The epithet Hox should
no longer be applied to other classes of homeobox genes (Scott, 1992). We include within
the 'Hox' designation the non-homeotic genes of the DrosophilaHox clusters (bcd, ftz,
zen). The final section of this manuscript provides some justification for this usage. We
use the term Antennapedia-class (Antp-class) Hox genes more restrictively, to refer to
that subset of the Hox genes that have a homeobox conforming to the Antp-class
consensus defined by Scott et al (1989). This includes all the central genes of the Hox
clusters, but excludes the labial, proboscipedia, Deformed and Abdominal-B classes (vertebrate paralogy groups l-4, 9-13).
zen. The role of these genes during early development has
been studied in some detail. It appears to be without
parallel among the vertebrate Hox genes. No well
'
conserved homologues of these genes have been found in
other taxa, suggesting that they are evolving faster than the
homeotic genes. Relatively divergent Antp-class genes
isolated from other insects are probably homologues of
fushi-tarazu, but these are almost unrecognisable outside of
their homeodomains, and have accumulated approximately
10 times as many changes in their homeodomains as have
in the same comparisons. They show
conserved patterns of expression in the nervous system, but
not during early development.
homeotic genes
Key words: homeobox, homeotic gene, tagmosis, insect development
evolution
HOX GENE DUPLICATIONS AND THE ORIGIN OF
ARTHROPOD BODY PLANS
An explicit
hypothesis relating Hox genes
change was articulated by E. B. Lewis
paragraph of his classic paper (Lewis , 1978):
to evolutionary
in the opening
"During the evolution of the fly, two major groups of genes
must have evolved: 'leg suppressing' genes which removed
legs from abdominal segments of millipede like ancestors,
followed by 'haltere-promoting' genes which suppressed the
second pair of wings of four-winged ancestors...
During evolution a tandem ar-ray of redundant genes presumably diversified by mutation to produce this complex."
(the Bithorax Complex).
In essence, Lewis proposed that the evolutionary history of
the homeotic gene clusters would be related to the evolution
of morphology in the arthropod lineage, with increasing
complexity of the gene cluster underlying increasing complexity of the body plan. No doubt he had in mind a scheme
proposed by Snodgrass (1935), where the archetypal insect is
derived from a superficially myriapod-like ancestor in which
all the segments between the mouthparts and the anal
segment were similar. In the parlance of arthropod morphologists, such an animal is homonomous; all the segments of
the trunk form a single tagma, or body region. This contrasts
with the regional specialisation of segment form and function
210
M. Akam and others
W
that constitutes tagmosis, and characterises most arthropod
classes.
Snodgrass' scheme assumes
that in the
proposed
insect/myriapod lineage, a homonomous trunk is the primitive
state, and tagmosis a derived character. At first sight this
conflicts with the observation that most representatives of the
other major group of mandibulate arthropods - the Crustacea
- also display clear trunk tagmosis, and typically have distinct
thorax and abdomen (Schram, 1986). However, conventional
phylogenies argue that these analogous states have been
acquired independently in the two lineages (Brusca and
Brusca, 1990): homonomous Crustacea exist (the remipedes),
w
ffi
Insects
Crustaceans
@
and are assumed to reflect the condition ancestral to this whole
class (Fig. 1).
It is not obvious why a homonomous, myriapod-like animal
would need the array of homeotic genes that specify trunk
ffi
in Drosophila, most particularly the set of
distinct Antp-class Hox genes that specify the thoracic and
segment diversity
abdominal segments to be different from one another (Antennapedia (Antp), Ultrabithorax (Ubx) and abdominal-A (abdA)). Indeed, we previously argued that the duplication leading
to these particular genes may have occuffed relatively late, in
association with the origin of the thorax/abdomen distinction
in insects (Akam et al., 1988).
One test of this model is to compare the structure and
expression of Antp-class Hox genes in crustaceans and insects.
If the mechanism to specify thorax and abdomen arose independently in these two lineages, we should see signs of this
when we examine the molecular machinery. The first step in
this comparison has been achieved (Averof and Akam, 1993).
In a study of Hox gene sequences in the branchiopod crustacean Artemia, we have shown that specific homologues of
Antp, Ubx and abd-A, as well as Sex Combs Reduced (Scr) and
Deformed
@fA are
present
in
Crustacea. Other, more
divergent Hox gene classes (labial, proboscipedia, AbdominalB) are known to be of even more ancient origin (Fig. 2;
Schubert et al., 1993). We conclude that the existing diversity
of homeotic genes in insects (or at least Drosophila) derives
from gene duplications that occurred prior to the insect/crustacean split.
Arthropods recognisable as crustaceans are well documented among Cambrian fossils (Robison and Kaesler, 1987),
including probable Branchiopods from the lower Cambrian,
550 Myr before present (Butterfield, 1994). In contrast, the
earliest insect fossils appear significantly later, during the
Devonian, 400 Myr before present (Jeram et al., 1990). These
insects are likely to derive from an early crustacean-mandibulate clade, in which case the final diversification of the Antpclass Hox genes may have occurred within this arthropodan
lineage. However, at least one of the relevant gene duplication
events occurred earlier, before the separation of annelid and
arthropod lineages: leeches possess a Ubx/abd-A type gene
(Lox2), distinct from other Antp class sequences (Lox I, Lox5;
Wysocka-Diller
et &1., 1989;
Nardelli-Haefliger
and
Shankland, 1992).
These findings argue against the strong version of the Lewis
hypothesis - that the acquisition of trunk tagmosis in insects
was directly correlated with the origin of new Hox genes.
However, they do not rule out the possibility that significant
changes in the regulation of pre-existing Hox genes may have
played a crucial part in this transition. Further work will be
#*
Insects
w
_aF
Crustaceans
Fig. 1. 'Conventional' and 'alternative' hypotheses concerning the
origin of trunk tagmosis in relation to arthropod phylogeny.
(A) Conventional view depicting the common ancestor of insects and
crustaceans with a homonomous trunk. Thorax/abdomen tagmosis is
assumed to have arisen independently in the two lineages.
Myriapods, assumed to constitute a sister group to the insects, and
remipedes within the Crustacea, retain a homonomous trunk
reflecting the primitive condition for this whole lineage (B) An
alternative scenario in which insects and Crustacea derive from a
common ancestor that already displays well differentiated trunk
segments (perhaps controlled by the differential expression of
multiple Antp-class Hox genes, which we infer to have been present
in this ancestor). Those myriapods (some or all) that constitute a
sister group to the insects, and remipedes among the Crustacea,
would then show a secondarily simplified trunk. Note that the
monophyly of the myriapods is uncertain, and the position of some
myriapods within this lineage has recently been challenged:
Sequence data (Turbeville et al., l99l; Ballard et al., 1992; Averof,
1994) and patterns of neural development (Whitington et al., l99l)
suggest that many myriapods may form an outgroup to the whole
lineage depicted here.
required to establish if and how the Antp class Hox genes are
used to define trunk segment identities in Crustacea, and in
homonomous arthropods (e.g. myriapods).
Even before these data are available, it is perhaps worth
questioning the validity of one assumption on which the Lewis
model is based, for the argument illuminates one contribution
that developmental data may make to evolutionary discussions.
This is the assumption that homonomy is always primitive. As
developmental geneticists, we would emphasise that there is an
enorrnous asymmetry between the invention of a complex
feature (e.g. the transition from homonomy to heteronomy) and
its loss (heteronomy+homonomy). An animal that has no
Hox genes in arthropods 211
ChOrdateS
(Amphioxus, human, mouse)
t 2 3 4 5 6 7 8 9 l0 il t2
tIt
rll
123
4
Annelids
(6-8)(e-13)
13
(Helobdeua, Hirudo)
TTTIIIMT
I-;i t- I
Lx5 Lx2
III
t23
Crustaceans
(Ailemia)
trtrtrIIIIINT
IIIIIIIINI
t z 34
Scr
Dfd Scr Hx I Ant Ubx abA
(4)
fi:*on,
ubx abA
[l[,
InSeCtS
lab pb
(I
(o"osophila, Tribolium, Schistocerca)
zen Dfd Scr
) (2) (3?) (4)
ftz Ant Ubx abA
Dax
AbB
(9- 13)
Fig.2.The minimal inferred complexity of Hox gene clusters in the lineage leading to the insects. Panels on the right summarise the diversity
of Hox genes described from insects, Crustacea, annelids (Class Hirudinea, leeches) and chordates(Amphioxus, and several vertebrates; a
single 'complete' chordate cluster is illustrated, based on Amphioxus data; Garcia-Fernandez and Holland, 1994). Genes characterised by at
least the full homeobox sequence are shown as coloured boxes. Open boxes in the annelid and crustacean panels are used for genes that are
assumed to exist in these taxa, but have not yet been characterised adequately. Identical colours are used for genes that can be assigned unique
homologues (orthologues) within the clusters of different taxa. Similar but non-identical colours are used for genes where orthology
relationships are unclear. Boxes are joined only where linkage has been established in some member of the taxon. The assumed phylogenetic
relationships of these four taxa are not disputed. Panels on the left show the minimal complexity of the Hox cluster in each of the stem groups
that is implied by considering this phylogeny together with the assumed orthology relationships. A single Antp-class ancestor is shown for the
basal stem group (though Scr and the genes of vertebrate paralogy group 5 may derive from a second ancestral Antp-class sequence present at
the base of this lineage. The origin of these genes is not clear; Btirglin,1994). DistinctAntp-like and Ubx/abd-A-like genes are shared by
arthropods and annelids. All homeotic gene classes are shared by insects and Crustacea. The inclusion of a group 3 homologue in the arthropod
lineages is based on unpublished data (see text), and on the isolation of a homeobox PCR fragment that has residues diagnostic for class 3 Hox
genes from Limulus, a chelicerate arthropod (Cartwright et al., 1993). Gene symbols have been abbreviated as follows: Lx, Lox (leech Hox)
genes; Ant, Antp; abd, Ubx/abd-Arelated; ab-A, abd-A; Ab-B, Abd-B. (References: Chordates: Biirglin, 1994; Garcia-Fernandezand Holland,
1994. Annelids: Loxl, Aisemberg and Macagno, 1994); Inx2,Wysocka-Diller et al., 1989; Nardelli-Haefliger and Shankland, 1992; Lox 5,6,7 ,
Shankland et al., 1991. Crustaceans: Averof and Akam, 1993; see also legend to Fig.4. Insects: For a summary of Drosophila gene sequences,
see Biirglin, 1994; for other insects, see legend to Fig. 4.)
Fig. 3. Segment specification involves precise spatial regulation of
homeotic genes in Drosophila. A region of the Drosophila embryo at
extended germ band stage (maxillary to second abdominal segments)
has been stained for protein produced by the Ubx gene (black). At
this stage, homeotic genes are regulated in parasegmental
domains.Ubx plays no role in segment specification in parasegment 4
(PS4; posterior first thoracic segment/anterior second). Near
ubiquitous expression of Ubx characterises parasegment 6 (PS6,
T3p/A I a), and can be shown to specify PS6 by experimental
manipulation (Gonzalez-Reyes et al., 1990; Mann and Hogness,
1990). Spatially regulated expression of Ubx characterises
parasegment 5 (PS5, T2ptT3A), and is necessary for its correct
specification. In this preparation, counter staining for the product of a
distal-less reporter gene (Vachon et al., 1992) marks the developing
leg discs of the thoracic segments (brown). Scale, l0 pm.
212
M. Akam and others
developmental mechanism to make segments different must
invent both the mechanism to specify the differences, and also
the machinery to elaborate distinct, adaptive segment types.
Loss of heteronomy can be achieved by what seem to us to be
much simpler processes - by turning off the mechanism that
specifies segments to be different, or simply by eliminating
some tagma from the adult body plan altogether.
We do not seriously doubt the hypothesis that the first
metamerically segmented ancestors of the arthropods were
homonomous - though there is at present no way of knowing
where to place that ancestor on the phylogenetic tree. We do
question whether homonomy is generally a primitive trait
among extant arthropods. In an environment where it is useful
for all segments to have legs, this adaptive change may be relatively easy to achieve. A single Bx-C mutation will make a
'myriapod' of a fly (albeit not a viable one; Lewis, 1978), but
we cannot conceive of the reverse transition occurring in a
single step.
An analogous situation pertains to the origin and loss of
insect wings. It takes many genes to make a wing (Williams
and Carroll , 1993) yet, in Drosophila, we know of several
single gene mutations that will suppress wing formation in an
otherwise viable fly (wingless, apterous, vestigial; Lindsley
and Zimm, 1992). Among the insects, where the phylogenetic
context is much less ambiguous, wings are believed to have
evolved only once; we do not hesitate to invoke convergence
repeatedly to explain the origin of wingless taxa at many phylogenetic levels among the pterygotes, from whole orders (e.g.
fleas, lice) to individual species (Kristensen, I99l).
If heteronomy is hard to achieve de novo, but useful and
easy to modify once invented, we should expect to see two very
different modes of evolution: clades that generate a diversity
of taxa which retain essentially homonomous body plans, and
clades that exploit heteronomy to filI many niches. Trilobites
may exemplify the first mode; despite their abundance and
taxonomic diversity throughout the Palaeozorc, virtually all
trilobite families retain largely homonomous trunk segments
(Eldridge, 1977). The Crustacea present a very different
picture, with a Palaeozoic radiation giving rise to a huge
diversity of body plans (Cisne, 1974; Schram, 1986; Briggs et
&1., 1992).
HOMEOTIC GENE REGULATION AND
EVOLUTIONARY CHANGE WITHIN INSECTS
Hox gene duplications may have been a necessary precondition for segment diversification within the insects, but the
results summarised above suggest that they were not an
immediate driving force for its appearance. There is some
evidence, however, that changes in the regulation of the Hox
genes have occurred within insects, and may have contributed
to morphological evolution. These changes are to be
seen
against a broadly conserved role for many Hox genes, at least
as can be inferred from the phenotypes of mutations
in the Hox clusters of beetles and moths (Beeman et a1., 1993;
Tazima, 1964; Booker and Truman, 1989; Ueno et al. , 1992),
and from the patterns of expression of Hox genes in these and
other insects (see below).
Most data are available for abd-A. In each of four species
where the expression of abd-A has been examined
in so far
(Drosophila, Karch et al., 1990; Macias et aI.,1990; Manduca,
Nagy et al. , I99I; Tribolium, Stuart et al. , 1993; Schistocerca,
Tear et al., 1990), the gene is expressed throughout much of
the abdomen, with no evidence for expression in the head or
thorax. Moreover, in each case, the gene shows an abrupt
anterior boundary of expression within the first abdominal
segment. Where tested, this lies at the parasegmental boundary
defined by engrailed expression, suggesting that, at least in this
case, the precise co-ordination of segmentation and homeotic
gene expression
is
maintained
in
insects as diverse
Orthopteru and Diptera (Tear et al., 1990).
In
some other cases, the domains
as
of homeotic gene
expression are broadly conserved, but the precise limits of
expression are not the same as those in Drosophila. For
example, the Abd-B genes in Tribolium and Schistocerca
appear to be expressed in the epidermis of only the most
posterior abdominal segments (from A8p back; Kelsh et al.,
1993; J. He & R. Denell, personal communication), whereas
in Drosophila Abd-B is also expressed in the more anterior
abdominal segments, A5p-A8a. This anterior expression of the
gene, which is necessary for normal development, is regulated
quite independently of that in the more posterior abdomen: It
derives from a separate promoter and is activated through
different regulatory elements (Boulet et a1., 199I). Altered regulation of the Abd-B gene may have played a role in the
evolution of the modified abdomen that is characteristic of the
higher Diptera.
Comparisons between Schistocerca and Drosophila reveal
one clear case of 'molecular heterochrony'. In Drosophila,
abd-A is normally never expressed in the ninth or more
posterior abdominal segments (parasegment l4).All of these
segments fuse
with A8 to form the 'terminalia'. In Schisto-
cerca, abd-A is expressed at high levels in A.9 and A10 during
early embryogenesis, but this expression is rapidly lost, at the
same time that Abd-B expression extends anteriorly (Tear et
&1., 1990; Kelsh et al., 1994). It seems most likely that this is
due to the direct repression of the abd-A gene by Abd-B
protein, for this same interaction has been demonstrated in
Drosophila.
In Abd-B mutants of
Drosophila,, abd-A
expression extends posteriorly into ,A.9 (Macias et al., 1990),
and this segment now develops characteristics of the more
anterior abdomen, including a denticle belt (S6nchez-Herrero
et &1., 1985). Thus a regulatory interaction that occurs as a
temporal sequence after segment formation in Schistocerca,
appears to be 'hard wired' at an earlier stage in Drosophila
development, and is only apparent in mutants.
The two cases above suggest that alterations in the timing
or extent of homeotic gene expression may be used to modify
segment development even though they do not result in
striking 'homeotic' transformations. However, there ate a
of observations in the entomological literature sug-
number
gestive of homeotic transformations that have been fixed in the
course of evolution. One that we are unlikely to be able to study
in detail concerns the Monura - an extinct group of insects that
flourished in the Carboniferous (Kukalova-Peck, 1991). These
insects, in common with a number of other early groups, had
reduced but well-formed legs (with terminal claws) on each
abdominal segment A1 to A10: thorax-abdomen tagmosis was
not complete. However, in contrast with all other known
insects, in the Monura, these legs were also present on the
terminal abdominal segment, in place of the filiform cerci that
Hox genes in
are the usual appendages of A1 1. The presumed sister group
(Paleodictyoptera) and outgroups (Thysanura, Symphyla) to
the Monura had cerci, so it is possiUte that a homeotic transformation of cerci to leglets occurred in this particular order.
Alternatively, of course, this 'simple' character state may
represent a retained primitive feature that has been lost several
times in other groups.
The case of the Strepsipteran haltere is more amenable to
experimental analysis. The Strepsiptera ate a small order of
parasitic insects. The adult females are neotenrc, retaining larval
morphology. It is the males that are interesting for our pu{poses
(Whiting and Wheeler, 1994). They have only a single pair of
wings, but these-are developed from the third thoracic segment,
not the second, as in the Diptera. The dorsal appendages of the
second thoracic segment are reduced to halteres, which in many
details resemble the halteres derived from the third thoracic
segment of Diptera. Until recently, this resemblance has been
to
ascribed
convergence, for the Strepsiptera have been
assumed to be only distantly related to the Diptera. Ribosomal
DNA sequence data now provides strong evidence that the
Strepsiptera are in fact the sister group to the Diptera. This
prompted Whiting and Wheeler to suggest that the Strepsipteran
haltere is indeed homologous to the haltere of Diptera, and that
this developmental pathway has been adopted in T2 of the
Strepsiptera, presumably by a change in the segmental regulation of Ubx. This hypothesis is open to test, using a recently
described antibody that cross-reacts with Ubx protein in a wide
range of insect species (Kelsh et al., 1994).
However, we should be cautious in making such predictions.
This dramatic modification of adult segment morphology may
depend entirely on the regulation of genes acting downstream
of the homeotics. Even if it does involve modification of
homeotic gene expression, it may involve only their regulation
in a subset of tissues at a particular stage in development (e.g.
in the developing imaginal discs). It is not necessary to assume
that evolutionary change affecting the homeotic genes must
involve major, saltatory change in the morphology of whole
segments. It may much more frequently proceed by incremental changes affecting levels of expression, or by the alteration
of those enhancer elements within their promoters that control
activity in particular structures within a segment.
This view is supported by careful studies of the role of one
homeotic gene, Ubx, rn Drosophila (J. Castelli-Gair and M.
Akam, unpublished data). The Ubx gene is responsible for
defining the identities of two quite different (para)segments,
parasegment 5 (T3) and parasegment 6 (A1). This differential
function does not appear to depend on the segment-specific
expression of different protein variants (Busturia et al., 1990).
Rather it seems to involve the precise temporal and spatial regulation of the gene in parasegment 5 (Fig. 3). Examples of how
changes in the precise pattern of within-segment regulation
may have contributed to insect segment diversity are provided
by the work of Carroll et al. (this volume). They show, for
example, that the loss of expression of abd-A specifically
within some limb buds may allow the development of larval
abdominal prolegs in Lepidoptera.
HOX GENES THAT GOT AWAY
Up to this point, we have taken for granted that Hox
genes
arthropods
213
cloned from any insect can be identified without question as
the speciflc homologue (orthologue) of one of the Hox cluster
genes defined rn Drosophila. In general, this is the case. With
few exceptions, the homeodomains are almost identical, and
gene-specific sequences around
the
homeodomain and
extending upstream to the hexapeptide motif (Biirglin, 1994)
are sufficiently similar to make identification unambiguous.
This is true, however, only for homologues of the 'homeotic'
genes within the Drosophila clusters (labial, proboscipedia,
Dfd, Scr, Antp, Ubx, abd-A and Abd-B). Well-conserved homologues have not been isolated for the several homeobox genes
of the Antp complex that have 'anomalous' roles in development. There are four such genes rn Drosophila melanogaster
- bicoid, fushi-taruzu and the two zen genes.
Bicoid is only expressed maternally. It encodes a gradient
morphogen
in the early (syncytial) embryo (Driever
and
Niisslein-Volhard, 1988). Fushi-taruzu (ftz) is a 'pair-rule' segmentation gene (Kaufman et al., 1990). It is re-used during
neurogenesis to specify the identity of certain neurons in each
trunk segment (Doe et al., 1988). Zenl is required very early
for the specification of the dorsal pattern elements in the
embryo (Ferguson and Anderson, 1991). None of these roles
is analogous to that of the typical homeotic genes, or to those
of the Hox genes in vertebrates, so far as we know. The
existence of these genes within the insect Hox clusters has been
something of an enigma, but isolation of several 'divergent'
Hox genes from other insects throws some light on their origin.
The best studied of these are putative homologues of fushitaraztt, isolated from Tribolium (Brown et al.,1994) and Schistocerca (Dawes et al., 1994). Neither of these genes shows
extensive similarity to ftz (or any other Hox gene) outside of
the homeobox, and even within the homeobox they are much
more divergent than is typical for the other Hox genes (Fig. a).
However, both of these genes are closely linked to the Hox
cluster, and both show a pattern of expression in the nervous
system very similar
to that of
the fushi-tarazu gene in
Drosophila.
Outside of the homeodomain, short conserved peptides are
shared uniquely between Drosophila ftz and the Tribolium ftzlike gene, and between the Tribolium and Schistocerca genes.
In the latter case, the conserved region is centred on a YPWM
sequence just upstream of the homeodomain. This so-called
hexapeptide motif is characteristic of the homeotic Hox genes,
and generally well conserved between distant species, often
showing extended similarity between insect and vertebrate
genes of the same class. Its presence in the Schistocerca and
Tribolium genes strongly suggests that, on structural grounds,
these should be regarded as 'typical' Hox genes. However,
note that no such motif is conserved in the Drosophila ftz gene.
In contrast to the conserved expression of ftz in the nervous
system, these putativ e ftz homologues show different patterns
of expression during early development. Drosophila ftz is
expressed in parasegmental stripes at two-segment intervals,
and is needed for the formation of alternate segment boundaries (Lawrence and Johnston,
1
989). ln Tribolium, the ftzgene
is expressed in less well resolved stripes, and is apparently not
needed for making the correct number of segments (Stuart et
dI., I99l; Brown et al., 1994). The Schistocerca gene is
expressed in a posterior domain of the early embryo, but never
in stripes (Dawes et al., 1994); its function in early development is unknown.
214
M. Akam and others
A
homeotic
O
+
bicoid
ftz-related
amlno
acid
A
diffs
in
Hbox
A
A
VS
D. mel.
AA
A
A
A
A
A
uugltso
HS(DCD
o'oo.:1
A6_Tg3
6;;!-'
=rli]
Fo
have been unsuccessful (I. Dawson, S. Frenk and M. A.,
unpublished results).
We think that these genes may be derived, Iike ftz, from Hox
genes which,
cocoo
+==:1_=
EEqds
H.
A
Because the changes in the homeobox are confined to regions
other than those that contact DNA, we suggest that it may be
the lower complexity of protein-protein interactions that allows
ftz to diverge more rapidly than the canonical homeotic genes.
The other 'non-homeotic' genes of the Drosophila Ant-C are
in some ways analogous to ftz. Their homeoboxes are very
divergent, with respect to all other non-Dipteran homeobox
genes; they lack any recognisable hexapeptide motif; comparison between Dipteran species shows that the bicoid homeobox
is diverging rapidly (Sommer and Tautz, l99I; Schroder and
Sander , 1993). Activity capable of rescuing bicoid mutants can
be recovered from the eggs of other cyclorapphan Diptera, but
was not detected in honeybee, or even lower Diptera (Schroder
and Sander, 1993). Searches for bicoid genes in other insects
5(D
)4
(D(a
ts.V
-D9
Fr
o.ts
;r
+
CD
-F!
v)
Oc
oF
ii
(D
Fig. 4. Relative divergence of homeotic and other Hox cluster
homeodomains within Arthropods. Species for which Hox gene
sequences are available have been grouped according to approximate
phylogenetic distance from Drosophila melanogaster. Rough
estimates of divergence times are given below, based on
immunological similarity (within Diptera; Beverley and Wilson,
1984)) or on the first appearance of other taxonomic groups in the
fossil record (Kukalova-Peck, 1991 ). Other Drosophila subgenera,
(40-60 My): D. hydeiftz (Jost and D. Maier, personal
communication; EMBL accessio n X7 9494); other Dipteran
famlly/suborder, (100-200My): Musca bicoid (n.b.6 changes in 44
residues of homeodomain sequenced (Sommer and Tautz, I99l);
Aedes abd-A (Eggleston, 1992); other Endopterygote insects
(250My): Bombyx (Lepidoptera) Ubx, abd-A (Ueno et al. , 1992);
Manduca (Lepidoptera) abd-A (Nagy et al. , I99I); Apis
(Hymenoptera) Antp, Scr, Dfd (Fleig et al., 1988; Walldorf et al.,
1989); Tribolium (Coleoptera) abd-A (Stuart et al. , 1993), ftz (Brown
et al. , 1994). Schistocerca (Orthoptera, approx. 300My) abd-A (Tear
et aI.,1990), Abd-B (Kelsh et al., 1993), Scr (Akam et al., 1988),
Dax (ftz-related gene (Dawes et al., 199D; crustacean: Artemia
(Anostraca,550 My; Averof and Akam, 1993). The assignment of
the divergent Artemia gene AfiIxl to theftz homology group is
tentative, but is consistent with the fast divergence of these genes.
For comparison, the divergence between some annelid or vertebrate
Hox sequences and their closest Drosophila match are shown on the
right.
We believe that all of these ftz-related genes derive from a
single Antp-class Hox gene that acquired novel functions in
arthropod development. They are not involved in the maintenance of segmental identity in the mesoderm or the epidermis.
They may have a role in very early patterning of the embryo
in all insects, but this role appears to be different in different
insects. In Schistocerca and Triboliuffi, whatever constraints
maintain the integrity of the YPWM motif and its flanking
sequences are retained, but the function of ftz tn Drosophila
appears no longer to impose the same sequence requirement.
in the lineage
leading to Drosophila, have
escaped from the conservative selection that characterises
homeotic genes. We have recently isolated from Schistocerca
a cDNA that lends some support to this hypothesis. It encodes
a homeodomain protein that shows some similarities to the
proboscipedia and Zen genes, but has no close homologue in
Drosophila. It resembles most closely Hox class 3 vertebrate
sequences (F. F., unpublished results). As yet we know nothing
of the function or expression of this gene tn Schistocerca, but
we guess that it may derive from an ancestral sequence that
gave rise, after considerable divergence, to the zen and/or
bicoid genes of Drosophila.
Work from the authors laboratory was supported by the Wellcome
Trust. We thank Nick Butterfield, Dieter Maier, Sue Brown and Rob
Denell for communicating results prior to publication; Geoffrey Fryer
and Hans-Georg Frohnhofer for their advice at the early stages of our
Crustacean work, Peter Holland and David Stern for valuable
comments on the manuscript.
REFERENCES
Aisemberg, G. O. and Macagno, E. R. (1994). Loxl, an Antennapedia-Class
homeobox gene is expressed during leech gangliogenesis in both transient
and stable central neurons. Dev. Biol. 161,455-465.
Akam, M., Dawson,I. and Tear, G. (1988). Homeotic genes and the control of
segment identity. Development 104 Supplement , 123-133.
Averof, M. (1994). HOM/Hox Genes of a Crustacean; Evolutionary
Implications. Ph.D. thesis, University of Cambridge.
Averof, M. and Akam, M. ( 1993). HOM/Hox genes of Artemia: rmplications
for the origin of insect and crustacean body plans. Current Biology 3,7378.
Ballard, J. W. O., Olse[, G. J., Faith, D. P., Odgers, \ry. A., Rowell, D. M.
and Atkinson, P. \ry. 0992). Evidence from 12S ribosomal RNA sequences
that onychophorans are modified arthropods. Science 258, 1345-1348.
Beeman, R. \ry., Stuart, J. J., Brown, S. J. and Denell, R. E. (1993). Structure
and function of the homeotic gene complex (HOM-C) in the beetle Tribolium
c as tane um. B io E s s ay s 15, 439 -444.
Beverley, S. M. and Wilson, A. C. (1984). Molecular evolution in Drosophila
and the higher Diptera.z. A time scale for fly evolution. J. Mol. Evol.2l, l13.
Booker, R. B. and Truman, J. \ry. (1989). Octopod, a homeotic mutation of
the moth Manduca sexta, influences the fate of identifiable pattern elements
within the CNS . Development 105, 621-629.
Boulet, A. M., Lloyd, A. and Sakonju, S. (1991). Molecular definition of the
morphogenetic and regulatory functions and the cis-regulatory elements of
the Drosophila Abd-B homeotic gene. Development 11.L, 393-406.
Briggs, D. E. G., Fortey, R. A. and Wills, M. A. (1992). Morphological
disparity in the Cambrian. Science 256, 1670-1673.
Hox genes in arthropods 215
Brown, S. J., Hilgenfeld, R. B. and Denell, R. E. (1994). The beetle Tribolium
castaneum has a fushi tarazu homolog expressed in stripes during
segmentation. Proc. Nat. Acad. Sci. USA (in press).
Brusca, R. C. and Brusca, G. J. (1990). Invertebrates. Massachusetts:
Sinauer.
Biirglin, T. R. (1994). A comprehensive classification of homeobox
genes. In
Guidebook to the Homeobox Genes (ed. D. Duboule), pp. 25-71. Oxford:
Oxford University Press.
Busturifl, A., Vernosr I., Macias, A., Casanovfl, J. A. and Morata, G. (1990).
Different forms of Ultrabithorax proteins generated by alternative splicing
are functionally equivalent. EMBO J. 9,3551-3555.
Butterfield, N. J. (1994). Burgess Shale-type fossils from an early Cambrian
shallow-shelf sequence, northwestern Canada. Nature 369, 477 -479
Carroll, S. B. (1994). Developmental regulatory mechanisms in the evolution
of insect diversity . Development 1994 Supplement,
Cartwright, P., Dick, M. and Buss, L.'W. (1993). Hom/Hox type homeoboxes
in the Chelicerate Limulus polyphemus. Mol. Phylogenet. Evol. 2,185-192.
Cisne, J. L. (1974). Evolution of the world fauna of aquatic free-living
arthropo ds . Evolution 28, 337 -366
Dawes, R., Dawson, I., Falciani, F., Tear, G. and Akam, M. ( 1994). Dax, a
Locust Hox gene related tofushi-tarazu but showing no pair-rule expression.
.
.
Development 120, I 56 I -157 2.
Doe, C. Q., Hiromi, Y., Gehring, W. J. and Goodman, C. S. (1988).
Expression and function of the segmentation gene fushi tarazu during
Drosophila neurogenesis. Science 239, 17 0-17 5 .
Driever, W. and Niisslein-Volhard, C. (1988). The bicoid protein determines
position in the Drosophila embryo in a concentration-dependent manner.
Cell 54,95-104.
Egglestor, P. (1992).Identification of the abdominal-Ahomologue from Aedes
aegypti and structural comparisons among related genes. Nucleic Acids
Re s e arch 20, 4095 -4096.
Eldridge, N. (1977). Trilobites and evolutionary patterns. In Patterns of
evolution, as illustrated by the fossil record (ed. A. Hallam), pp. 305-332.
Amsterdam: Elsevier.
Fergusor, E. L. and Andersor, K. Y. ( 199 I ). Dorsal-ventral pattern formation
in the Drosophila embryo - the role of zygotically active genes. Current
Topics Dev. Biol. 25, 17 -43.
Fleig, R., Walldorf, U., Gehring, \ry. J. and Sander, K. (1983). In situ
localisation of the transcripts of a homeobox gene in the honeybee Apis
mellifura L. (Hymenoptera). Roux's Arch. Dev. Biol. 197,269-276.
Garcia-Fernandez, J. and Holland, P. 'W. H. (1994). Archetypal organization
of the Amphioxus Hox gene cluster. Nature 370,563-566.
Gonzalez-Reyes, A., Urquia, N., Gehring, W., Struhl, G. and Morata, G.
(
1990).
Are cross regulatory
interactions between homeotic genes
functionally significant? Nature 344, 78-80.
Jeram, A. J., Selden, P. A. and Edwards, D. (1990). Land animals in the
Silurian: arachnids and myriapods from Shropshire, England. Science 658669.
Karch, F., Bender, W. and Weiffenbach, B. (1990). abdominal-A expression
rn Drosophila embryos. Genes Dev. 4, 1573-1588.
Kaufmar, T. C., Seeger, M. A. and Olsen, G. (1990). Molecular and genetic
organisation of the Antennapedia complex of Drosophila melanogaster.In
Genetic Regulatory Hierarchies in Development (ed. T. R. F. Wright), pp.
Academic Press.
Kelsh, R., Dawson, I. A. and Akam, M. (1993). An analysis of Abdominal-B
expression in the locust Schistocerca gregaria. Development ll7, 293305.
Kelsh, R., Weinzierl, R. O. J., White, R. A. H. and Akam, M. (1994).
Homeotic gene expression in the Locust Schi stocerca: an antibody that
detects conserved epitopes in Ultrabithorax and abdominal-A proteins.
Developmental Genetics 15, 19-31.
Kristensen, N. P. (1991). Phylogeny of extant hexapods. In The Insects of
Australia (ed. CSIRO. Div. of Entomology), pp. 125-140. Melbourne:
Melbourne University Press.
J. (1991). Fossil history and the evolution of hexapod
structures. In The Insects of Australia (ed. CSIRO Div. of Entomology), pp.
l4l-179. Melbourne: Melbourne Univ. Press.
Lawrence, P. A. and Johnstor, P. (1989). Pattern formation in the Drosophila
embryo: allocation of cells to parasegments by even-skipped and fushi
tarazu. Development 105, 7 6l-7 69 .
Kukalova-Peck,
Lewis, E. B. (1978). A gene complex controlling segmentation rn Drosophila.
Nature 276,565-570.
L. and Zimm, G. G. (1992). The genome of Drosophila
melanogaster. San Diego: Academic Press.
Macias, A., Casanova, J. and Morata, G. (1990). Expression and regulation
of the abd-A gene of Drosophila. Development 110, ll97 -1207 .
Mann, R. S. and Hogness, D. S. (1990). Functional dissection of Ultrabithorax
proteins tn D . melano gaster. Cell 60, 597 -6 10.
Lindsley, D.
Nagy,
L. M., Booker, R. and Riddiford, L. M.
(1991). Isolation and
embryonic expression of an abdominal-A-llke gene from the lepidopteran,
Manduca sexta. Development ll2, lI9-I31.
Nardelli-Haefliger, D. and Shankland, M. (1992). Lox2, a putative leech
segment identity gene, is expressed in the same segmental domain in
different stem cell linea ges. Development 116, 697 -7 10.
A. and Kaesler, R. L. (1987). Phylum Arthropoda. In Fossil
Invertebrates (ed. R. S. Boardman, A. H. Cheetham and A. J. Rowell), pp.
205-269. Palo Alto, CA: Blackwell Scientific.
Robison, R.
S6nchez-Herrero,8., Vern6s, f., Marco, R. and Morata, G.
Genetic organisation
(1985).
313,
of the Drosophila bithorax complex . Nature
108-l 13.
Schram, F. R. (1986). Crustacea. Oxford: University Press.
Schriider, R. and Sander, K. (1993). A comparison of transplantable bicoid
activity and partial bicoid homeobox sequences in several Drosophila and
blowfly species (Calliphoridae). Roux's Arch. Dev. 9ioL203,34-43.
Schubert, F. R., Nieselt-Struwe, K. and Gruss, P. (1993). The Antennapediatype homeobox genes have evolved from three precursors separated early in
metazoan evolution. PNAS 90, 143-147 .
Scott, M. P. (1992). Vertebrate Homeobox gene nomenclature. Cell7l,55l-553.
Scott, M. P., Tamkun, J. W. and Hartzell, G.'W. I. (1989). The structure and
function of the homeodomain. Biochim. Biophys. Acta 989, 25-48.
Shankland, M., Martindale, M. Q., Nardelli-Haefliger, D., Baxter, E. and
Price, D. J. (1991). Origin of segmental identity in the leech nervous system.
D ev e lo p me nt Supplem ent 2,, 29 -38 .
Snodgrass, R. E. (1935). Principles of Insect Morphology. London and New
York: McGraw-Hill.
Sommer, R. and Tautz,
D. (1991). Segmentation gene expression
housefly Musca domestica. Development 113, 419-30.
in
the
Stuart, J. J., Brown, S. J., Beeman, R. \ry. and Denell, R. E. (1991). A
deficiency of the homeotic complex of the beetle Tribolium. Nature 350,7274.
Stuart, J. J., Brown, S. J., Beeman, R. \ry. and Denell, R. B. (1993). The
Tribolium homeotic gene Abdominal rs homologous to abdominal-A of the
e I o p me nt ll7, 233 -243
Tazima, Y. (1964). The Genetics of the Silkworm. London: Logos Press.
D r o s o p hil a bithorax complex. D ev
.
Tear, G., Akam, M. and Martinez-Arias, A. (1990). Isolation of
an
abdominal-A gene from the locust Schistocerca gregaria and its expression
during early embryogenesis. Development 110, 915-926.
Turbeville, J. M., Pfeifer, D. M., Field, K. G. and Raff, R. A. (1991). The
phylogenetic status of arthropods as inferred from 18S rRNA sequences.
Mol. Biol. Evol.8, 669-686.
Ueno, K., Hui, C.-C., Fukuta, M. and Suzuki, Y. (1992). Molecular analysis
of the deletion mutants in the E homeotic complex of the silkw orm Bombyx
mori. D ev elopment ll4, 555-563.
Vachon, G., Cohen,8., Pfeifl€, C., McGuffin, M. E., Botas, J. and Cohen, S.
M.
(1992). Homeotic genes
of the bithorax
complex repress limb
development in the abdomen of the Drosophila embryo through target gene
Distal-less. Cell Tl, 437 -450.
Walldorf, U., Fleig, R. and Gehring, W. J. (1989). Comparison of homeoboxcontaining genes of the honeybee and Drosophila. Proc. Natl. Acad. Sci.
usA 86,9971-997 5.
Whiting, M. F. and Wheeler, W. C. (1994). Insect homeotic transformation.
Nature 368, 696.
Whitington, P. M., Meier,
T. and King, P. (1991).
Segmentation,
neurogenesis and formation of early axonal pathways in the centipede,
Ethmostigmus rubripes (Brandt) . Roux's Arch Dev Biol 199,349-363.
Williams, J. A. and Carroll, S. B. (1993). The origin, patterning and evolution
of insect appenda ges. BioEssays l5, 567 -577 .
Wysocka-Diller, J. W.o Aisemb€rg, G. O., Baumgarten, M., Levine, M. and
Macagno, E. R. (1989). Characterrzatron of a homologue of bithorax
complex genes in the leech Hirudo medicinalis. Nature 341,760-763.