caudal
To infer similarities and differences in terminal pattern formation
in insects, several of the key genes of this process were analyzed in the
beetle Tribolium castaneum. Two
genes of the terminal pattern cascade, namely tailless
(tll) and forkhead (fkh), from Tribolium were cloned and
their expression patterns were studied. In addition, the pattern of MAP
kinase activation was analyzed at blastoderm stage as a possible signature for
torso-dependent signaling. Further, the late
expression of the previously cloned Tribolium caudal
(Tc-cad) gene was examined. Finally, the upstream region of
Tc-tll was used to drive a reporter gene construct in
Drosophila. This construct is activated at
the terminal regions in Drosophila, suggesting that the
torso-dependent pathway is conserved between the
species. Most of the expression patterns of the genes
studied here are similar in Drosophila and Tribolium, suggesting conserved functions (Schroder, 2000).
A caudal homolog of Bombyx mori encodes a protein of 244 amino acids. The homology between Drosophila
and Bombyx homeodomains is 80%. There is a single maternal transcript of 2.3 kb. Bombyx CAD transcripts accumulate first in the nurse cells and are later transferred into the oocyte at a defined time
during oogenesis. The maternal transcripts of Bombyx CAD form a concentration gradient
spanning the anteroposterior axis during the gastrulation stage and are restricted to the anal pad,
the most posterior domain (Xu, 1994).
A homolog to the Drosophila caudal (cad) gene has been isolated from the flour beetle Tribolium
castaneum and its expression pattern has been studied. The Tribolium caudal (Tc-cad) gene arrangement is
unusual in that there is a partial duplication of the gene resulting in alternative transcripts with identical
5'-exons, but different 3'-exons encoding two different homeoboxes. Expression analysis was carried out using
whole-mount in situ hybridization and staining with an antibody raised against the N-terminal part of
the protein that is common to both transcripts. At early stages a homogeneously distributed
maternal mRNA is observed which is initially also translated throughout the embryo. A little later, a posterior to
anterior CAD protein gradient is formed, very similar to that in Drosophila. However, because of the
differences in the fate map between Drosophila and Tribolium (see Tribolium early embryonic development), the CAD protein expression at
blastoderm stage covers the prospective head and thoracic region and not the abdomen as in
Drosophila. Expression of Tc-cad in the prospective abdomen is only seen during further germband
growth where it becomes restricted to the growth zone in which the segments are formed. This
expression is very similar to the growth zone expression in the somitogenic region seen for cad
homologs in vertebrates. After the completion of the segmentation process Tc-cad expression becomes confined to a terminal stripe that resembles a similar stripe at late blastoderm stages in Drosophila (Schulz, 1998).
Insect axis formation is best understood in Drosophila, where rapid anteroposterior patterning of zygotic determinants is directed by maternal gene products. The earliest zygotic control is
by gap genes, which determine regions of several contiguous segments and are largely conserved in insects. Isolation of mutations has been used to approach a genetic question: do early zygotic patterning genes control similar
anteroposterior domains in the parasitoid wasp Nasonia vitripennis as in Drosophila? Nasonia is
advantageous for identifying and studying recessive zygotic lethal mutations because unfertilized eggs develop as males while fertilized eggs develop as females.
On first consideration, the Hymenopteran Nasonia and the
Dipteran Drosophila appear very similar in their embryonic
development, though the Hymenoptera diverged from the
Diptera >200 million years ago. Embryos of both species
produce larvae in about 1 day at 25ƒC. In Nasonia, the fertilized egg
gives rise to an embryo that undergoes syncytial and cellular
blastoderm stages morphologically similar to those of
Drosophila. Both Nasonia and Drosophila undergo the long
germband mode of embryonic development. Despite these
similarities, two observations suggest that the relative
importance of maternal versus zygotic patterning functions
may differ in the two insects. (1) Although postgastrulation
events proceed with very similar timing, the time for early
development differs substantially - at 25ƒC: the events
preceding gastrulation take only about 3 hours in Drosophila
but almost 10 hours in Nasonia. This difference in timing may
allow for greater zygotic control of patterning in Nasonia than
in Drosophila. (2) Among the relatives of Nasonia, a
polyembryonic mode of development has evolved in which a
single fertilized egg gives rise to hundreds or thousands of
progeny. Polyembryonic development
is likely to rely heavily on zygotic control of patterning.
Polyembryony has arisen several times in the Hymenoptera,
and the polyembryonic Copidosoma floridanum is in the same superfamily as Nasonia. These
considerations pose the following question -- is early development
substantially controlled by the zygotic genome in
Hymenopterans?
This question may be approached genetically, by isolating zygotic mutations that disrupt early anteroposterior patterning in Nasonia. Recessive zygotic
mutations have identified three Nasonia genes: head only mutant embryos have posterior defects, resembling loss of both maternal and zygotic Drosophila caudal function; headless mutant embryos
have anterior and posterior gap defects, resembling loss of both maternal and zygotic Drosophila hunchback function, and squiggy mutant embryos develop only four full trunk segments, a phenotype more
severe than those caused by lack of Drosophila maternal or zygotic terminal gene functions. head only mutant embryos lack all segmentation posterior to
the head, in the strongest manifestation of the phenotype, and have only a narrow domain of Ubx-Abd-A expression. head only differs from Drosophila gap genes with respect to the extent of pattern deleted and effects on Ubx-Abd-A. In Drosophila, neither Krüppel nor knirps affects a domain as large as that of head only. Moreover, the wild-type functions of Krüppel and knirps are not required for the positive regulation of Ubx or abd-A in Drosophila (Pultz, 1999).
The characterization is reported of a caudal gene from the rhizocephalan cirripede Sacculina carcini and its embryonic and larval expression patterns. Cirripedes are maxillopodan crustaceans that are devoid of any complete abdominal segment at the adult stage. The genetic basis of this peculiar body plan is being explored. They probably lack the abdominalA gene, while possessing the other Hox genes shared by arthropods. However, at least a part of the genetic program might be conserved, since the engrailed.a and engrailed.b genes are expressed in a posterior region that is interpreted as a relic of an ancestral abdomen. The Sacculina caudal gene is expressed early in embryogenesis, which makes it the earliest genetic marker evidenced in the development of Sacculina and of any other crustacean species. It is expressed later in the embryo in the caudal papilla, a posterior proliferating zone of cells. During the larval stages, the caudal gene is first expressed in the whole thoracic region; then its expression regresses to the posterior end of the larva. Surprisingly, it is never expressed in the vestigial abdomen. This lack of expression of the Sacculina caudal gene in a posterior region, at odds with what is known in all other studied metazoan species, might be correlated with the defective development of the abdomen (Rabet, 2001).
All arthropods share the same basic set of Hox genes, although the
expression of these genes differs among divergent groups. In the brine shrimp
Artemia franciscana, their expression is limited to the head,
thoracic/trunk and genital segments, but is excluded from more posterior parts
of the body, consisting of six post-genital segments and the telson (bearing
the anus). Nothing is currently known about the genes that specify the
identity of these posterior structures. The expression patterns was studied of
four candidate genes, Abdominal-B, caudal/Cdx,
even-skipped/Evx and spalt, the homologs of which are
known to play an important role in the specification of posterior structures
in other animals. Abdominal-B is expressed in the genital segments of
Artemia, but not in the post-genital segments at any developmental
stage. The expression of caudal, even-skipped and
spalt in the larval growth-zone suggests they may play a role in the
generation of body segments (perhaps comparable with the role of gap and
segmentation genes in insects), but not a direct role in defining the identity
of post-genital segments. The expression of caudal at later stages
suggests a role in the specification of anal structures. A PCR screen designed
to isolate Hox genes expressed specifically in the posterior part of the body
failed to identify any new Hox genes. It is concluded that the post-genital
segments of Artemia are not defined by any of the genes known to play
a role in the specification of posterior segments in other arthropods. It is
argued that these segments constitute a unique body region that bears no
obvious homology to previously characterized domains of Hox gene activity (Copf, 2003).
The number of leg-bearing segments in centipedes varies
extensively, between 15 and 191, and yet it is always odd. This
suggests that segment generation in centipedes involves a stage with
double segment periodicity and that evolutionary variation in segment
number reflects the generation of these double segmental units.
However, previous studies have revealed no trace of this. The
expression of two genes, an odd-skipped related gene
(odr1) and a caudal homolog, is described that serve as
markers for early steps of segment formation in the geophilomorph
centipede, Strigamia maritima. Dynamic expression of
odr1 around the proctodaeum resolves into a series of
concentric rings, revealing a pattern of double segment periodicity
in overtly unsegmented tissue. Initially, the expression of the
caudal homolog mirrors this double segment periodicity, but
shortly before engrailed expression and overt segmentation,
the intercalation of additional stripes generates a repeat with
single segment periodicity. These results provide the first clues
about the causality of the unique and fascinating "all-odd" pattern
of variation in centipede segment numbers and have implications for
the evolution of the mechanisms of arthropod segmentation (Chipman,
2004).
These observations invite comparison with the process of segment
generation in Drosophila. There, a pattern of double segment
periodicity is first generated and then subdivided to yield the final
single segment repeat. However, the generation of the 'pair-rule'
pattern in Drosophila shows few if any similarities with the early
stages of segmentation in Strigamia . Drosophila subdivides
the entire body axis into unique domains by activating 'gap genes'
under the influence of maternal gradients and then uses the complex
promoters of the pair-rule genes to compute a repetitive pattern of
gene activity from this underlying aperiodic pattern. The generation
of this pattern is almost static with respect to the forming cells of
the blastoderm. In Strigamia, the initial patterns of
odr1 expression are not static with respect to the
underlying cells. It is suggested that the patterns of odr1
gene expression are oscillations of cell state, coordinated as waves
that move across the population of cells in the blastodisc,
sharpening to encompass fewer cells and stabilizing to double segment
periodicity. Thus, despite the fact that odd-skipped is one
of the genes expressed in a pair-rule pattern during Drosophila
segmentation, it is thought likely that the processes that generate
this pair-rule pattern are different in the two species (Chipman,
2004).
Interestingly, odd-skipped family members are downstream
targets of the Notch signaling pathway during Drosophila limb
segmentation. Recently, it has been shown that the Notch
ligand Delta and its target hairy are expressed in
a striped pattern during early development and segmentation in the
embryo of the spider Cupiennius salei. It has been suggested
that Notch signaling in the spider is generating a reiterated pattern
through a mechanism analogous to that shown for vertebrate
segmentation. These two observations, taken together, suggest the
possibility that the odd-skipped family in
Strigamia, and possibly in other arthropods, is modulated
through a Notch-Delta-mediated oscillator to generate the first
serially repeated pattern that begins the segmentation process
(Chipman, 2004).
A separate and unresolved issue is whether there are similarities
between the process that resolves the pair-rule repeat of Drosophila
into a single segment pattern and the process whereby secondary
caudal stripes intercalate between primary stripes to
generate the single segment repeat in Strigamia. The
possibility that such 'frequency doubling' processes may be
widespread among the arthropods is supported by the observation of
analogous phenomena in chelicerates and short germ insects: in the
mite Tetranychus urticae, expression of the paired
gene in the prosoma is initially at double segment intervals, with
secondary stripes intercalating between them to generate the single
segment repeat. In the growing abdomen of the grasshopper
Schistocerca americana, paired gene expression also
shows a transition from double to single segment periodicity, though
in this case the process is one of stripe splitting rather than
intercalation. However, in other cases, either no such pattern has
been described or the periodicity of gene expression is not yet
clear. Of particular relevance in this context is a recent study of
segmentation gene expression in the lithobiomorph centipede,
Lithobius atkinsoni (only distantly related to
Strigamia). The expression of even-skipped in the
posterior of the Lithobius germ band shows broad rings
around the proctodeum that could reflect dynamic expression,
resolving into stripes. However, there is as yet no evidence of
subsequent frequency doubling (Chipman, 2004).
These results provide a possible explanation for the observation
that, in nature, centipede segment number varies in two-segment
increments. It is proposed that variation in segment number among
centipedes is caused by variation in the number of cycles of a
primary segmentation oscillator, each cycle of which generates two
segments. The anteroposterior range of this process may well extend
beyond the trunk to include the poison claw and parts of the head and
genital regions. Therefore, the occurrence of odd rather than even
numbers of leg-bearing segments is not incompatible with this
explanation (Chipman, 2004).
Although the molecular mechanisms directing anteroposterior patterning of the Drosophila embryo (long-germband mode) are well understood, how these mechanisms were evolved from an ancestral mode of insect embryogenesis remains largely unknown. In order to gain insight into mechanisms of evolution in insect embryogenesis, the expression and function of the orthologue of Drosophila caudal (cad) was examined in the intermediate-germband cricket Gryllus bimaculatus. A posterior (high) to anterior (low) gradient in the levels of Gryllus bimaculatus cad (Gb′ cad) transcript is formed in the early-stage embryo, and then Gb' cad is expressed in the posterior growth zone until the posterior segmentation is completed. Reduction of Gb' cad expression level by RNA interference results in deletion of the gnathum, thorax, and abdomen in embryos, remaining only anterior head. The gnathal and thoracic segments are formed by Gb' cad probably through the transcriptional regulation of gap genes including Gb' hunchback and Gb' Krüppel. Furthermore, Gb'cad is found to be involved in the posterior elongation, acting as a downstream gene in the Wingless/Armadillo signalling pathways. These findings indicate that Gb'cad does not function as it does in Drosophila, suggesting that regulatory and functional changes of cad occurred during insect evolution. The Wg/Cad pathway in the posterior pattern formation may be common in short- and intermediate-germband embryogenesis. During the evolutionary transition from short- or intermediate- to long-germband embryogenesis, an ancestral cell-cell signalling system including Wg/Arm signalling may have been replaced by a diffusion system of transcription factors as found in Drosophila. Since Wnt/Cdx pathways are involved in the posterior patterning of vertebrates, such mechanisms may be conserved in animals that undergo sequential segmentation from the posterior growth zone (Shinmyo, 2005).
One of the earliest steps of embryonic development is the establishment of polarity along the anteroposterior axis. Extensive studies of Drosophila embryonic development have elucidated mechanisms for establishing polarity, while studies with other model systems have found that many of these molecular components are conserved through evolution. One exception is Bicoid, the master organizer of anterior development in Drosophila and higher dipterans, which is not conserved. Thus, the study of anteroposterior patterning in insects that lack Bicoid can provide insight into the evolution of the diversity of body plan patterning networks. To this end, the long germ parasitic wasp Nasonia vitripennis has been establised as a model for comparative studies with Drosophila. In Nasonia, a gradient of localized caudal mRNA directs posterior patterning, whereas, in Drosophila, the gradient of maternal Caudal protein is established through translational repression by Bicoid of homogeneous caudal mRNA. Loss of caudal function in Nasonia results in severe segmentation defects. Nasonia caudal is an activator of gap gene expression that acts far towards the anterior of the embryo, placing it atop a cascade of early patterning. By contrast, activation of gap genes in flies relies on redundant functions of Bicoid and Caudal, leading to a lack of dramatic action on gap gene expression: caudal instead plays a limited role as an activator of pair-rule gene expression. These studies, together with studies in short germ insects, suggest that caudal is an ancestral master organizer of patterning, and that its role has been reduced in higher dipterans such as Drosophila (Olesnicky, 2006).
mRNA localization is a powerful mechanism for targeting factors to different regions of the cell and is used in Drosophila to pattern the early embryo.
The parasitoid wasp Nasonia (Hymenoptera) undergoes long germ development similar to that of Drosophila, yet is evolutionarily very distant from flies (> 200 MY) and lacks bicoid. During oogenesis of Nasonia, mRNA localization is used extensively to replace the function of the bicoid gene for the initiation of patterning along the antero-posterior axis. Nasonia localizes both caudal and nanos to the posterior pole, whereas giant mRNA is localized to the anterior pole of the oocyte; orthodenticle1 (otd1) is localized to both the anterior and posterior poles. The abundance of differentially localized mRNAs during Nasonia oogenesis provided a unique opportunity to study the different mechanisms involved in mRNA localization. Through pharmacological disruption of the microtubule network, it was found that both anterior otd1 and giant, as well as posterior caudal mRNA localization was microtubule-dependent. Conversely, posterior otd1 and nanos mRNA localized correctly to the posterior upon microtubule disruption. However, actin is important in anchoring these two posteriorly localized mRNAs to the oosome, the structure containing the pole plasm. Moreover, knocking down the functions of the genes tudor and Bicaudal-D mimics disruption of microtubules, suggesting that tudor’s function in Nasonia is different from flies, where it is involved in formation of the pole plasm (Olesnicky, 2007).
Both the Drosophila and Nasonia ovariole are meroistic, meaning that the nurse cells and oocyte are both of germ cell descent and originate from the same primordium, but differentiate during subsequent cell divisions. As each ovarian follicle develops and is positioned more distally along the ovariole, the nurse cells remain attached to one another and to the oocyte through ring canals, which arise from incomplete cleavage during cell division. The 16 sister cells that make up each germline cyst result from four of these incomplete divisions. An egg chamber forms comprising of 15 nurse cells and the oocyte, surrounded by the somatic follicle cells, which form an epithelial layer around the oocyte. Nurse cells produce metabolites and other factors that transit through the ring canals to accumulate in the oocyte (Olesnicky, 2007).
The Drosophila oocyte is specified early during oogenesis as a result of the asymmetric segregation of the fusome, an organelle that connects the 16 cells. Once the oocyte has been specified, the polarity of the oocyte microtubule network becomes extremely dynamic and undergoes a major reorganization resulting from communication between the oocyte and follicle cells. This reorganization is essential to localize maternal mRNAs that will generate the axes of the embryo. At first, microtubule minus ends extend from the nurse cells into the oocyte toward a microtubule organizing center (MTOC) localized at the posterior pole of the oocyte, near its nucleus. Later, however, the posterior MTOC disassembles while multiple MTOCs form toward the anterior of the growing oocyte. At this stage, the microtubules are therefore pointing from the plus end at the posterior of the oocyte to the minus end at the anterior. mRNAs and the oocyte nucleus utilize the polarity of the microtubules to localize to the anterior or posterior pole (Olesnicky, 2007).
Nasonia oogenesis presents striking similarities to that of Drosophila. It is divided into five morphologically distinct stages. In stage 1, the nurse cells and oocyte are indistinguishable until they begin to segregate, with the oocyte lying towards the posterior of the follicle. By stage 2, the nurse cells and a smaller oocyte are clearly distinguishable, as a constriction forms between the oocyte and its supporting nurse cells. At this stage, the oocyte nucleus is positioned in the center of the cell. The oocyte continues to grow throughout stage 3 until it becomes larger than its accompanying nurse cells. Concomitantly, the oocyte nucleus migrates to the dorsal anterior cortex of the developing oocyte, as in Drosophila. Later, during stage 4, the nurse cells begin to degenerate as they empty all their contents into the oocyte. In the final stage (5), a vitelline membrane is constructed around the embryo (Olesnicky, 2007).
This study shows that the localization of four maternal mRNAs is achieved using at least 2 distinct mechanisms. It is shown that, during Nasonia oogenesis, microtubules play a major role in oocyte polarity and in the control of anterior localization of otd1 and gt mRNA and the posterior localization of cad mRNA. In contrast, the actin cytoskeleton is important for anchoring the oosome and is therefore essential for the localization of nanos and otd1 mRNA to the posterior pole of the oocyte (Olesnicky, 2007).
It is proposed that Nasonia utilizes two basic mechanisms for the localization of mRNA, a microtubule-dependent mechanism and an actin-dependent, microtubule-independent one. Anterior localization of gt and otd1 mRNA, as well as posterior localization of cad mRNA, all rely on a similar microtubule-dependent mechanism while posterior localization of otd1 and nos mRNAs relies on actin. In wild-type follicles, cad and gt mRNAs are initially localized, while later in oogenesis this localization is relaxed to achieve a more graded mRNA distribution. otd1 anterior mRNA, although not graded, is also localized loosely in wild-type follicles. nos mRNA localization and posteriorly localized otd1 mRNA, however, are tightly localized to the posterior in a microtubule-independent manner. Interestingly, in freshly laid embryos both posterior otd1 mRNA and nos mRNA are localized to the oosome. Maintaining localization of these two posteriorly localized mRNAs relies on the actin cytoskeleton. Additionally, actin might be required to anchor the oosome to the posterior pole of the oocyte, as well as to trap mRNA to the oosome. It is therefore likely that both mRNAs are localized to structures within the germ plasm, resulting in a tight localization that is maintained throughout oogenesis and early embryogenesis and does not rely extensively on microtubules (Olesnicky, 2007).
In the development of most arthropods, the caudal region of the elongating
germ band (the growth zone) sequentially produces new segments. Previous work
with the spider Cupiennius salei suggested involvement of Delta-Notch
signaling in segmentation. This study reports that, in the spider Achaearanea
tepidariorum, the same signaling pathway exerts a different function in
the presumptive caudal region before initiation of segmentation. In the
developing spider embryo, the growth zone becomes morphologically apparent as
a caudal lobe around the closed blastopore. Preceding caudal
lobe formation, transcripts of a Delta homolog, At-Delta,
are expressed in evenly spaced cells in a small area covering the closing
blastopore and then in a progressively wider area of the germ disc epithelium.
Cells with high At-Delta expression are likely to be prospective
mesoderm cells, which later express a twist homolog,
At-twist, and individually internalize. Cells remaining at the
surface begin to express a caudal homolog, At-caudal, to
differentiate as caudal ectoderm. Knockdown of At-Delta by parental
RNA interference results in overproduction of At-twist-expressing
mesoderm cells at the expense of At-caudal-expressing ectoderm cells.
This condition gives rise to a disorganized caudal region that fails to
pattern the opisthosoma. In addition, knockdown of Notch and
Suppressor of Hairless homologs produces similar phenotypes. It is
suggested that, in the spider, progressive activation of Delta-Notch signaling from around the blastopore leads to stochastic cell fate decisions between mesoderm and caudal ectoderm through a process of lateral inhibition to set up a functional caudal lobe (Oda, 2007).
The early asymmetric cleavages of C. elegans embryos produce
blastomeres with distinct developmental potentials. The caudal-like
homeodomain protein PAL-1 is required to specify the somatic identity of one
posterior blastomere in the 4 cell embryo. pal-1 activity is sequentially
restricted to this blastomere. Initially, at the 4 cell stage, it is translated only in the two
posterior blastomeres. Its function is then restricted to one of these blastomeres. This
second targeting step is dependent on the activities of the posteriorly localized SKN-1
and asymmetrically segregated PIE-1 proteins. It is proposed that the segregation of
PIE-1, combined with the temporal decay of SKN-1, targets pal-1 activity to this
posterior lineage, thus coupling the regulation of this conserved posterior patterning
gene to asymmetric cell cleavages (Hunter, 1997).
In Caenorhabditis elegans males, a row of epidermal precursor cells called seam cells generates a
pattern of cuticular alae in anterior body regions and neural sensilla (called rays) in the posterior. The
Abdominal-B homolog mab-5 is required for two posterior seam cells, V5 and V6, to generate rays. The V5 lineage generates one ray and the V6 lineage generates five rays. In mab-5
mutant males, V5 and V6 do not generate sensory ray lineages but instead generate lineages that lead
to alae, cuticular ridges that extend along the two sides of the animal. Alae are normally generated by the V1-V4 cells only. Two independent regulatory pathways can activate mab-5 expression in the
V cells. (1) The caudal homolog pal-1 turns on mab5 in V6 during embryogenesis. (2) A Wnt
signaling pathway is capable of activating mab-5 in the V cells during postembryonic development,
however, during normal development Wnt signaling is inhibited by signals from neighboring V cells. The
inhibition of this Wnt signaling pathway by lateral signals between the V cells limits the number of rays
in the animal and also determines the position of the boundary between alae and rays (Hunter, 1999).
The Caenorhabditis elegans gene pal-1, a homolog of Drosophila caudal, is required maternally for blastomere specification in the early embryo and postembryonically for tail development in males. Both pal-1 in C. elegans and caudal in
Drosophila are transcribed maternally as well as in the
embryo. However, whereas either transcription mode alone
is sufficient for normal embryonic development in Drosophila, both modes of expression appear to be required in C.
elegans. Zygotic transcription of pal-1 is required for posterior patterning during later embryogenesis.
Embryos homozygous for strong loss-of-function mutations arrest as nonviable L1 larvae with gross posterior defects. PAL-1
protein produced from zygotic transcripts is expressed dynamically during gastrulation and morphogenesis in specific cells
of all major lineages except the germ line. Most expressing cells are undergoing cell movements or forming midline
structures or both. Mutant embryos exhibit defects involving most of the expressing cells. Aberrant early cell positions are
observed in posterior hypodermis, both in the C-lineage cells that express pal-1 and in the neighboring hypodermal seam
cell precursors, which do not, as well as in posterior muscle derived from the C and D lineages. Defects in late gastrulation,
ventral hypodermal enclosure, and formation of the rectum result from failures of cell movements of ABp and MS descendants. Limited mosaic analysis supports the view that most of the required pal-1 functions are cell autonomous (Edgar, 2001).
Although PAL-1 seems likely to have several regulatory
targets, there is evidence at present for only two. One is the
Hox gene mab-5, which is activated in the ABa-derived V6
cells during midembryogenesis. This activation requires
pal-1 function, and recent evidence
indicates that pal-1 may directly activate mab-5
transcription in the V6 cells. mab-5
could also require pal-1 function for activation in the sex
myoblasts descended from the M cell. Regulation by pal-1 of another Hox gene, the posterior-group gene egl-5, would be consistent with the expression
patterns of pal-1 and egl-5 in the late embryo, but at present there is
no direct evidence for such regulation. Another target for which there is evidence is vab-7, a homolog of the Drosophila pair-rule gene even-skipped. Lack of zygotic vab-7 function causes embryonic defects in embryonic C-lineage hypodermal and muscle patterning, but the embryos are viable. vab-7 is expressed in descendants of the four Cpxx cells, which generate the
posterior set of right and left C-derived hypodermal and
muscle cells. This expression has been
shown to require maternal but not zygotic pal-1 expression. However, the capacity to respond to ectopically expressed pal-1 extends until morphogenesis, and in cosmid-rescued lines, in which
maternal pal-1 expression is not detected, zygotically
expressed pal-1 may activate vab-7. Moreover, pal-1 mutations
and vab-7 mutations cause very similar C-lineage
phenotypes, suggesting that pal-1 may function to maintain
vab-7 expression in the late embryo (Edgar, 2001 and references therein).
Translational control is an essential mechanism of gene control utilized throughout development, yet the molecular mechanisms underlying translational activation and repression are poorly understood. The translational control of the C. elegans caudal homolog, pal-1, has been investigated and it has been found that GLD-1, a member of the evolutionarily conserved STAR/Maxi-KH domain family, acts through a minimal pal-1 3' UTR element to repress pal-1 translation in the distal germline. Data is provided suggesting that GLD-1 may repress pal-1 translation after initiation. Finally, GLD-1 is shown to repress the distal germline expression of the KH domain protein MEX-3, which was previously shown to repress PAL-1 expression in the proximal germline and which appears specialized to control PAL-1 expression patterns in the embryo. Hence, GLD-1 mediates a developmental switch in the control of PAL-1 repression, allowing MEX-3 to accumulate and take over the task of PAL-1 repression in the proximal germline, where GLD-1 protein levels decline (Mootz, 2004).
GLD-1 is homologous to a sub-family of KH domain proteins known as the GSG or STAR domain family, whose members include the evolutionarily conserved Quaking protein, mammalian Sam68 and SF1 and Drosophila How. The ~200 amino acid STAR domain consists of an enlarged KH RNA-binding domain (maxi-KH domain) flanked by conserved residues on both sides. While the functions of these family members are not well understood, they have been implicated in various aspects of RNA metabolism, including mRNA splicing, nuclear export and translation. Other than GLD-1, only one family member, the mouse Quaking I isoform 6, has thus far been implicated as a translational regulator, and this is based on its ability to repress tra-2 expression when expressed in C. elegans (Mootz, 2004 and references therein).
Maternal and zygotic activities of the homeodomain protein PAL-1 specify the identity and maintain the development of the multipotent C blastomere lineage in the C. elegans embryo. To identify PAL-1 regulatory target genes, microarrays were used to compare transcript abundance in wild-type embryos with mutant embryos lacking a C blastomere and to mutant embryos with extra C blastomeres. pal-1-dependent C-lineage expression was verified for select candidate target genes by reporter gene analysis, though many of the target genes are expressed in additional lineages as well. The set of validated target genes includes 12 transcription factors, an uncharacterized wingless ligand and five uncharacterized genes. Phenotypic analysis demonstrates that the identified PAL-1 target genes affect specification, differentiation and morphogenesis of C-lineage cells. In particular, cell fate-specific genes (or tissue identity genes) and a posterior HOX gene are activated in lineage-specific fashion. Transcription of targets is initiated in four temporal phases, which together with their spatial expression patterns lead to a model of the regulatory network specified by PAL-1 (Baugh, 2005).
A model is proposed for the structure of the network specified by PAL-1 based only on temporal and spatial expression patterns in wild type. The rationale behind the model is simple: genes activated in one cell cycle affect the expression of genes expressed in the next cell cycle. This premise is supported by both functional analysis of the endodermal network and global analysis of expression dynamics. Furthermore, whereas zygotic pal-1 transcripts are first detected at the 2C-cell stage in Ca and Cp (phase I), loss of zygotic pal-1 function results in a detectable mutant phenotype in their daughters at the 4C-cell stag. In addition, protein for the phase II gene elt-1 is first detected at the end of the 4C-cell stage, and transcription of its confirmed phase III target elt-3 begins in the 8C-cell stage (Baugh, 2005).
That ectopic PAL-1 activity in early blastomeres is sufficient to cause complete transformation of one lineage into another indicates that the regulatory network specified by PAL-1 is modular or self-contained. After maternal PAL-1 specifies the C lineage, embryonically expressed PAL-1 is required for C-lineage development. It is therefore hypothesized that PAL-1 continuously regulates target genes during patterning of the C lineage, as opposed to simply initiating a transcriptional cascade. Although it is not known how far into development PAL-1 function is required, phenotypically mutant pal-1 mosaic animals were recovered corresponding to loss of pal-1 in one Cxx cell at the 4C-cell stage and PAL-1 expression is detectable in the C lineage until the 16C-cell stage, leaving open the possibility that PAL-1 directly activates each of the target genes identified in this study. Combinatorial control of gene expression, where early targets regulate late targets in combination with PAL-1, offers one possible mechanism for the timing of gene expression within this modular network (Baugh, 2005). There must be additional regulation not predicted by this model. Genes that are not PAL-1 targets are likely to participate in transcriptional regulation and patterning of the C lineage. For example, the Homothorax ortholog unc-62 and the Extradenticle homologs ceh-20 and ceh-40 have superficially similar phenotypes to nob-1 and pal-1, suggesting that these co-factor homeodomain proteins interact with and modify the function of PAL-1 and NOB-1 (Baugh, 2005).
Likewise, the Tcf/Lef factor pop-1 is thought to mediate cell-fate decisions associated with every cell division on the AP axis of the early embryo, and, as has been shown for development of the E lineage, POP-1 is expected to contribute to patterning of PAL-1 target expression, in particular where targets are expressed in only the anterior or posterior daughters following a round of C cell divisions (e.g., hlh-1, elt-1 and vab-7). Repression is completely ignored in the current model, but is probably crucial for patterning, as indicated by the fact that very few targets are expressed in all PAL-1-expressing cells. So there may also be genes repressed by PAL-1. The model does not allow for genes of the same temporal phase to regulate each other, though it is likely that there is mutual repression between genes specifying muscle and epidermis, leading to insulation of the two states. In addition, genes of the same temporal phase expressed in the same cells may activate the expression of one another, and multiple auto-regulatory positive feedbacks are expected in addition to the one demonstrated for pal-1. It will be interesting to compare the structures of different developmental regulatory networks in an effort to understand better how different topological motifs contribute to the functional properties of the regulatory network and ultimately how network structure relates to body plan (Baugh, 2005).
Genes of the Hox cluster are restricted to the animal kingdom and play a central role in axial patterning in divergent animal
phyla. Despite its evolutionary and developmental significance, the origin of the Hox gene cluster is obscure. The
consensus is that a primordial Hox cluster arose by tandem gene duplication close to animal origins. Several homeobox
genes with high sequence identity to Hox genes are found outside the Hox cluster and are known as 'dispersed' Hox-like
genes; these genes may have been transposed away from an expanding cluster. Three of these dispersed
homeobox genes form a novel gene cluster in the cephalochordate amphioxus.
AmphiCdx, AmphiXlox and AmphiGsx are in respectively the Caudal, Zerknullt, and Gsh (related to Labial and Proboscipedia) families of homeodomain proteins.
The finding that amphioxus Gsx, Xlox and Cdx class genes form a novel homeobox cluster challenges the idea that these homeobox gene classes are 'dispersed' Hox genes.
Instead it is argued that this 'ParaHox' gene cluster is
an ancient paralogue (evolutionary sister) of the Hox gene cluster. ParaHox and Hox gene clusters arose by duplication of a ProtoHox
gene cluster containing Gsx, Xlox and Cdx genes representing an anterior, 'group three' and posterior subfamily. Amphioxus ParaHox genes have co-linear developmental expression patterns in
anterior, middle and posterior tissues. It is proposed that the origin of distinct Hox and ParaHox genes by gene-cluster
duplication facilitated an increase in body complexity during the Cambrian explosion (Brooke, 1998).
Although the tail is one of the major characteristics of animals of the phylum Chordata, evolutionary aspects of the molecular mechanisms involved in its formation are
not clear. To obtain insights into these issues, the caudal gene of an ascidian, one of the lower animal groups among chordates, has been isolated and investigated.
Ascidian caudal is expressed from the midgastrula stage onward in the lateral walls of the posterior neural tube cell lineage and also in the posterior epidermal cells
from the neurula stage. Thus, ascidian caudal expression is restricted to the ectoderm of a tail-forming region throughout embryogenesis. Vertebrate caudal genes are expressed in all three germ layers. Suppression of caudal
function by an antisense oligonucleotide or a dominant negative construct causes inhibition of the cell movement required for tail formation. Overexpression of
wild-type caudal mRNA in an ascidian animal cap, an animal half explant prepared at the eight-cell stage, causes elongation of the cap. Furthermore, Xenopus
embryos injected with dominant negative ascidian caudal exhibit defects in elongation, suggesting a conserved caudal function among chordates. In ascidian embryos, intercalating cell movement along the posterior dorsal midline occurs within neuroectoderm and notochord upon tail elongation. Furthermore, it has been reported that the change of cell shape from columnar to wedge-shaped is first observed in the cells at the lateral borders of the neural plate. The initial expression of ascidian caudal coincides with this cell shape change, which may be responsible for the subsequent dorsal convergence of the ectodermal cell sheet. These results
indicate that caudal function is required for chordate tail formation and may play a key role in its evolution (Katsuyama, 1999).
Interaction of Wnt and caudal-related genes in zebrafish posterior body formation
Although Wnt signaling plays an important role in body patterning during early vertebrate embryogenesis, the mechanisms by which Wnts control the individual processes of body patterning are largely unknown. In zebrafish, wnt3a and wnt8 are expressed in overlapping domains in the blastoderm margin and later in the tailbud. The combined inhibition of Wnt3a and Wnt8 by antisense morpholino oligonucleotides leads to anteriorization of the neuroectoderm, expansion of the dorsal organizer, and loss of the posterior body structure -- a more severe phenotype than with inhibition of each Wnt alone -- indicating a redundant role for Wnt3a and Wnt8. The ventrally expressed homeobox genes vox, vent, and ved mediate Wnt3a/Wnt8 signaling to restrict the organizer domain. Of posterior body-formation genes, expression of the caudal-related cdx1a and cdx4/kugelig, but not Bmps or Cyclops, is strongly reduced in the wnt3a/wnt8 morphant embryos. Like the wnt3a/wnt8 morphant embryos, cdx1a/cdx4 morphant embryos display complete loss of the tail structure, suggesting that Cdx1a and Cdx4 mediate Wnt-dependent posterior body formation. cdx1a and cdx4 expression is dependent on Fgf signaling. hoxa9a and hoxb7a expression is down-regulated in the wnt3a/wnt8 and cdx1a/cdx4 morphant embryos, and in embryos with defects in Fgf signaling. Fgf signaling is required for Cdx-mediated hoxa9a expression. Both the wnt3a/wnt8 and cdx1a/cdx4 morphant embryos failed to promote somitogenesis during mid-segmentation. These data indicate that the cdx genes mediate Wnt signaling and play essential roles in the morphogenesis of the posterior body in zebrafish (Shimizu, 2004).
Fibroblast growth factor (Fgf) and retinoic acid (RA) signals control the
formation and anteroposterior patterning of posterior hindbrain. They are also
involved in development processes in other regions of the embryo. Therefore,
responsiveness to Fgf and RA signals must be controlled in a context-dependent
manner. Inhibiting the caudal-related genes cdx1a and
cdx4 in zebrafish embryos caused ectopic expression of genes that are
normally expressed in the posterior hindbrain and anterior spinal cord, and
ectopic formation of the hindbrain motor and commissure neurons in the
posteriormost neural tissue. Combinational marker analyses suggest
mirror-image duplication in the Cdx1a/4-defective embryos, and cell
transplantation analysis further revealed that Cdx1a and Cdx4 repress a
posterior hindbrain-specific gene expression cell-autonomously in the
posterior neural tissue. Expression of fgfs and retinaldehyde
dehydrogenase 2 suggested that in the Cdx1a/4-defective embryos, the Fgf
and RA signaling activities overlap in the posterior body and display opposing
gradients, compared with those in the hindbrain region. Fgf and
RA signals were required for ectopic expression. Expression of the posterior
hox genes hoxb7a, hoxa9a or hoxb9a, which function
downstream of Cdx1a/4, or activator fusion genes of hoxa9a or
hoxb9a (VP16-hoxa9a, VP16-hoxb9a) suppressed this
loss-of-function phenotype. These data suggest that Cdx suppresses the
posterior hindbrain fate through regulation of the posterior hox
genes; the posterior Hox proteins function as transcriptional activators and
indirectly repress the ectopic expression of the posterior hindbrain genes in
the posterior neural tissue. These results indicate that the Cdx-Hox code
modifies tissue competence to respond to Fgfs and RA in neural tissue (Shumizu, 2006).
In vertebrates, the caudal genes begin their
expression during gastrulation and they take up a posterior position. By injecting sense and antisense RNA of the Xenopus caudal gene Xcad-2, a number of regulatory interactions were studied among homeobox genes along the anterior-posterior axis.
Initially, the Xcad-2 and Otx-2 genes are mutually repressed; by late gastrulation,
they mark the posterior- or anterior-most domains of the embryo, respectively. During
late gastrulation and neurulation, Xcad-2 plays an additional regulatory function in
relation to the Hox genes. Hox genes normally expressed anteriorly are repressed by
Xcad-2 overexpression, while those normally expressed posteriorly exhibit more
anterior expression. The results show that the caudal genes are part of a posterior
determining network that during early gastrulation functions in the subdivision of the
embryo into anterior head and trunk domains. Later in gastrulation and neurulation
these genes play a role in the patterning of the trunk region (Epstein, 1997).
Recently, a model to explain the mechanism of Xenopus tail bud formation has been proposed. The NMC model proposes that three regions around the late blastopore lip are required to initiate tail formation. These are the posterior-most neural plate, fated to form tail somites (M), the neural plate (N), immediately anterior to M, and the underlying caudal notochord (C). To initiate tail formation, C must underlie (and presumably signal to) the junction of N and M, which subsequently forms the tip of the tail. During normal development, the NMC interaction leading to specification of the tail bud occurs at the end of gastrulation. Outgrowth of the tail bud commences much later, becoming clearly visible by stage 30 (Beck, 1998 and references).
Several domains of the Xenopus tail bud are defined by two phases of gene expression. The first group of genes are already expressed in the tail bud region before its determination at stage 13 and are subsequently restricted in the extending tail bud by stage 30. This group, the early genes, includes the Notch ligand X-delta-1, the lim domain homeobox factor Xlim1, the T-box factor Xbra, and the homeobox factor Xnot2 and Xcad2, a member of the caudal family. X-delta-1 is expressed specifically in the posterior wall of the neuroenteric canal but is excluded from the chordoneural hinge at stage 30, thus maintaining its earlier expression in the lateral and ventral blastopore lips. Xim1 is expressed in the notochord and dorsal blastopore lip at the end of gastrulation, and is maintained in the chordoneural hinge and posterior tip of the differentiated notochord in later stages. Xnot2 is expressed in the ventral neural tube and chordoneural hinge, but not in the posterior notochord. The posterior notochord therefore represents a novel tail bud region by stage 30, marked by Xlim but not Xnot transcripts, whereas the posterior ventral neural tube is marked by Xnot but not Xbra or Xlim1. Xbra is expressed in the chordoneural hinge and posterior wall. Xcad3 expression in the posterior neural plate is later maintained in the posterior wall and posterior dorsal neural tube. Xpo is expressed in all tissues of the tail bud with the exception of the chordoneural hinge, and is expressed in the fin and epidermis (Beck 1998).
Unlike the early genes, the regional expression of the second group of genes in the extended tail bud can not be traced back to the stage of tail bud initiation. These genes have a late onset of localized expression in the tail bud, corresponding to the beginning of tail outgrowth, although they may be expressed elsewhere in the embryo at stage 13. The dorsal roof domain of the tail bud is marked by expression of Xwnt3a and lunatic fringe. Xwnt5a expression is restricted to the tail bud roof. The distal tip of the tail, which comprises part of the posterior wall, is marked by expression of Xhox3, which marks the distal cells of the tail bud. Xhox3 is a vertebrate homolog of Drosophila evenskipped. Other late genes include BMP-4, X-serrate-1 and BMP-2 (Beck, 1998).
The existence of distinct domains in the positions predicted for C and M is proposed. The restriction of Xcad3 and Xlim1 transcripts to the posterior of the notochord in the early neurula demonstrates that the posterior part of the notochord differs from the crest, corresponding to the C region. Novel domains of the tail bud are proposed to express different combinations of genes. These domains include the dorsal roof of the tail bud, the distal tip of the tail, marked by Xhox3, the chordoneural hinge, the posterior tip of the chordoneural hinge, the posterior wall domain, the tip of the posterior wall, the posterior notochord, the posterior wall of the neuroenteric canal and the ventral neural tube (Beck, 1998).
Expression of the Xenopus Xcad-1 and Xcad-2 genes initiates during early gastrulation, exhibiting a
dorsoventral asymmetry in their transcription domains. At mid-gastrulation the ventral preference
becomes stronger and the expression of caudal genes takes up a posterior localization, which
will be maintained until their downregulation along the dorsal midline. Comparison of the three Xenopus
caudal genes reveals a temporal and spatial nested set of expression patterns. The transcription of the
caudal genes is sequentially downregulated with the one expressed most caudally (Xcad-2) being shut
down first, this sequence is most evident along the dorsal midline. This pattern of expression suggests a
role for the caudal genes as posterior determinants along the anteroposterior axis (Pillemer, 1998a).
Patterning of the marginal zone in the Xenopus embryo has been attributed to interactions between
dorsal genes expressed in the organizer and ventral-specific genes. In this antagonistic interplay of
activities, BMP-4, a gene that is not expressed in the organizer, provides a strong ventralizing signal.
The Xenopus caudal type homeobox gene, Xcad-2, which is expressed around the blastopore with the exception of a
gap over the dorsal lip, was analyzed as part of the ventral signal. Xcad-2 efficiently
represses during early gastrula stages the dorsal genes gsc, Xnot-2, Otx-2, XFKH1 and Xlim-1, while
it positively regulates the ventral genes Xvent-1 and Xvent-2, with Xpo exhibiting a strong positive
response to Xcad-2 overexpression. Xcad-2 is also capable of inducing BMP-4 expression in the
organizer region. Support for a ventralizing role for Xcad-2 was obtained from
co-injection experiments with the dominant negative BMP receptor, which was used to block BMP-4
signaling. Under lack-of-BMP-signaling conditions Xcad-2 can still regulate dorsal and ventral gene
expression and restore normal development, suggesting that it can act either downstream of BMP-4
signaling or independent of it. Xcad-2 can also inhibit secondary axis formation and dorsalization
induced by the dominant negative BMP receptor. Xcad-2 efficiently reverses the
dorsalizing effects of LiCl. These results place Xcad-2 as part of the ventralizing gene program, which acts during early gastrula stages, and demonstrate that Xcad-2 can execute its ventralizing function in the absence of BMP signaling (Pillemer, 1998b).
A dorsal-ventral difference in the specification of mesoderm in vivo has been discovered by examining the effect of
the dominant-negative FGF receptor on a new member of the Xenopus caudal gene family, Xcad-3. Xcad-3 is
expressed throughout the marginal zone during the gastrula stages and serves as a useful marker for events
occurring within the mesoderm. Disruption of the FGF signaling pathway by the dominant-negative FGF receptor,
disrupts the Xcad-3 expression pattern, eliminating expression preferentially from the dorsal regions of the
embryo. The expression of the Xenopus brachyury homolog, Xbra, is more readily eliminated
from the dorsal than the ventral region of the embryo by the dominant-negative FGF receptor, indicating that the
observed dorsal-ventral differences are not unique to Xcad-3. These results demonstrate the importance of
regional effects on FGF-mediated induction in vivo and suggest that FGF-dependent expression of mesodermal
genes depends upon the localization of other factors which establish dorsal-ventral differences within the
embryo (Northrop, 1994).
Recent studies on Xenopus development have revealed an increasingly complex array of
inductive, prepatterning, and competence signals that are necessary for proper mesoderm
formation. Fibroblast growth factor (FGF) signals through
mitogen-activated protein kinase kinase (MAPKK) to induce mesodermal gene expression. A partially activated form of MAPKK restores expression of the mesodermal
genes Xcad-3 and Xbra, eliminated by the dominant-negative FGF receptor (delta FGFR). Expression of a dominant-negative form of
MAPKK (MAPKKD) preferentially eliminates the dorsal expression of Xcad-3 and Xbra. Does the regional localization of bone morphogenetic protein-4 (BMP-4) explain
why both MAPKKD and delta FGFR eliminate the dorsal but not the ventral expression of
Xcad-3 and Xbra? Ectopic expression of BMP-4 is sufficient to maintain the dorsal
expression of Xcad-3 and Xbra in embryos containing delta FGFR, and expression of a
dominant-negative BMP receptor reduces the dorsal-ventral differences in delta FGFR embryos.
These results indicate that regional localization of BMP-4 is responsible for the dorsal-ventral
asymmetry in FGF/MAPKK-mediated mesoderm induction (Northrop, 1995).
The biological activities of the Xenopus caudal (Cdx) family member Xcad3 have been examined. A series of
domain-swapping experiments demonstrate that the N-terminus of Xcad3 is necessary for it to activate Hox gene expression
and that this function can be replaced by the activation domain from the viral protein VP16. Injection of 50 pg or more of
Xcad3 mRNA leads to activation of HoxC6 and HoxA7, which are normally expressed in both
the mesoderm and neuroectoderm, and HoxB7 and HoxB9, which are expressed predominantly in
the neuroectoderm. Xcad3 does not upregulate the expression of the general mesodermal
marker Xbra, indicating that it does not induce the formation of ectopic mesoderm. Experiments using an
Xcad3 repressor mutant (XcadEn-R), which potently blocks the activity of wild-type Xcad3, are reported. Overexpression of
XcadEn-R in embryos inhibits the activation of the same subset of Hox genes that are activated by wild-type Xcad3 and leads
to a dramatic disruption of posterior development. Xcad3 is shown to be an immediate early target of the FGF signaling
pathway: Xcad3 and FGF both posteriorize anterior neural tissue in similar ways. Xcad3 is also required for the
activation of Hox genes by FGFs. These data provide strong evidence that Xcad3 is required for normal posterior development
and that it regulates the expression of the Hox genes downstream of FGF signaling (Isaacs,1998).
Is there any evidence that the role for caudal-related genes in regulating Hox genes is conserved
outside the vertebrates? In Drosophila, homologs of the vertebrate Hox genes (Hom-C complex)
are considered to be largely epistatic to caudal, but there is now evidence suggesting that some aspects
of expression from the HOM-C complex member Abdominal-B are regulated by caudal. Interestingly, ectopic anterior expression of caudal results in a disruption of head
development, part of which appears to be due to the suppression of expression by the deformed
gene, which is also a member of the Drosophila Hom-C complex.
Certain parallels can be seen with the posterior-promoting/anterior-suppressing activity of the
Xenopus Xcad proteins. With regard to the role of caudal-related genes in other invertebrates, it
has been suggested that pal-1 is involved in regulation of the C.elegans Abd-B homolog mab-5 (Isaacs, 1998).
The KH domain protein MEX-3 is central to the temporal and spatial control of PAL-1 expression in the C. elegans early embryo. PAL-1 is a Caudal-like homeodomain protein that is required to specify the fate of posterior blastomeres. While pal-1 mRNA is present throughout the oocyte and early embryo, PAL-1 protein is expressed only in posterior blastomeres, starting at the four-cell stage. To better understand how PAL-1 expression is regulated temporally and spatially, MEX-3 interacting proteins (MIPs) have been identified and two that are required for the patterning of PAL-1 expression are described in detail. RNA interference of MEX-6, a CCCH zinc-finger protein, or SPN-4, an RNA recognition motif protein, causes PAL-1 to be expressed in all four blastomeres starting at the four-cell stage. Genetic analysis of the interactions between these mip
genes and the par genes, which provide polarity information in the early embryo, defines convergent genetic pathways that regulate MEX-3 stability and activity to control the spatial pattern of PAL-1 expression. These experiments suggest that par-1 and par-4 affect distinct processes. par-1 is
required for many aspects of embryonic polarity, including the restriction of MEX-3 and MEX-6 activity to the anterior blastomeres.
PAL-1 is not expressed in par-1 mutants, because MEX-3 and MEX-6 remain active in the posterior blastomeres. The role of par-4 is less well
understood. This analysis suggests that par-4 is required to inactivate MEX-3 at the four-cell stage. Thus, PAL-1 is not expressed in par-4 mutants because MEX-3 remains active in all blastomeres. It is proposed that MEX-6 and SPN-4 act with MEX-3 to translate the temporal and spatial
information provided by the early acting par genes into the asymmetric expression of the cell fate determinant PAL-1 (Huang, 2002).
Anterior-posterior patterning of the embryo requires the activity of multiple homeobox genes, among them Hox, caudal (Cdx, Xcad) and Otx2. During early gastrulation, Otx2 and Xcad2 establish a cross-regulatory network, which is an early event in the anterior-posterior patterning of the embryo. As gastrulation proceeds and the embryo elongates, a new domain forms, which expresses neither Otx2 nor Xcad2 genes. Early transcription of the Xenopus Gbx2 homolog, Xgbx2a, is spatially restricted between Otx2 and Xcad2. When overexpressed, Otx2 and Xcad2 repress Xgbx2a transcription, suggesting their role in setting the early Xgbx2a expression domain. Homeobox genes have been shown to play crucial roles in the specification of the vertebrate brain. The border between the transcription domains of Otx2 and Gbx2 is the earliest known marker of the region where the midbrain/hindbrain boundary (MHB) organizer will develop. Xgbx2a is a negative regulator of Otx2 and a weak positive regulator of Xcad2. Using obligatory activator and repressor versions of Xgbx2a, it has been demonstrated that during early embryogenesis, Xgbx2a acts as a transcriptional repressor. In addition, taking advantage of hormone-inducible versions of Xgbx2a and its antimorph, it has been shown that the ability of Xgbx2a to induce head malformations is restricted to gastrula stages and correlates with its ability to repress Otx2 during the same developmental stages. It is therefore suggested that the earliest known step of the MHB formation, the establishment of Otx2/Gbx2 boundary, takes place via mutual inhibitory interactions between these two genes and this process begins as early as midgastrulation (Tour, 2002).
The organizer in vertebrate embryos is responsible for the formation of the primary body axis. In amphibian embryos, the organizer forms in the dorsal marginal zone (prospective dorsal mesoderm) at a location determined by the point of sperm entry. Using inducible versions of axis-inducing proteins, it has been shown that, irrespective of the mode of secondary axis induction, organizer formation in the ventral marginal zone is temporally restricted from the midblastula transition to the onset of gastrulation. The competence of marginal zone cells to respond to organizer-inducing signals is under temporal control, one of the regulators being the homeobox transcription factor Xcad2. Overexpression of Xcad2 restricts the temporal competence for axis induction, whereas partial loss of function expands this competence, supporting the suggestion. It is proposed Xcad2 competes with putative axis-inducing signals within the marginal zone to prevent expression of organizer-specific genes. Elimination of endogenous Xcad2 allows for the activation of organizer genes beyond the normal competence window during early/mid-gastrulation. It is concluded that Xcad2, through its early expression in the ventrolateral marginal zone, terminates the competence of this embryonic region to respond to organizer-inducing signals by preventing the activation of organizer-specific genes (Levy, 2002).
Continued: Evolutionary Homologs part 2/2
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