branchless
FGF, cell migration, cell maturation and organization of the vertebrate a/p axis FGF-8 has attracted particular attention because of its importance for limb development in the chick
and mouse, although it also has a number of earlier expression domains in these species. An FGF-8 homolog has been cloned from Xenopus, a species in which it is relatively easy to do functional studies on early
development. There is no maternal expression, while zygotic expression is highest in the gastrula and
neurula stages. XFGF-8 is expressed as a ring around the blastopore and subsequently in the tail bud.
There are several domains in the head including the hatching gland, the branchial clefts, and the
midbrain-hindbrain border. At later stages there is a prominent band of expression in the limb bud
epidermis. Although there is no morphological apical ridge, this band of expression suggests that the
Xenopus limb bud contains a cryptic region with a similar ability to stimulate mesenchymal outgrowth.
The mesoderm-inducing activity of XFGF-8 is somewhat lower than that of other FGFs, while the
posteriorizing activity is similar. These differences are probably due to different receptor
specificities. The posterior expression and high posteriorizing activity suggest that XFGF-8 contributes to
the patterning of the anterior-posterior axis by FGF family members during gastrulation. In contrast to
the amniotes, Xenopus limb buds can regenerate following damage. Regeneration is
correlated with the reexpression of XFGF-8 in the distal epidermis, suggesting that this ability is critical
for successful limb regeneration (Christen, 1997).
In ascidian embryos (phylum Urochordata), inductive interactions are necessary for the fate specification of notochord cells. Previous studies have shown that notochord induction occurs at the 32-cell stage and that basic fibroblast growth factor (bFGF) has notochord-inducing activity in ascidian embryos. In vertebrates, it is known that bFGF receptors have tyrosine kinase domain; the signaling pathway is mediated by the
small-GTP binding protein, Ras. To study the role of Ras in ascidian embryos, dominant
negative Ras (RasN17) was injected into fertilized eggs. RasN17 inhibits the formation of notochord, suggesting
that the Ras signaling pathway is involved in signal transduction in the induction of notochord cells.
When the presumptive-notochord (A6.2) blastomere is co-isolated with the inducer (A6.1)
blastomere and then RasN17 is injected into the A6.2 blastomere, notochord differentiation is
suppressed. The presumptive-notochord blastomeres injected with RasN17 were treated with bFGF.
Many of them fail to develop notochord-specific features. Next to be examined was the effect of injecting
constitutively active Ras (RasV12) into the A6.2 blastomeres. However, microinjection of RasV12 into
these cells does not bypass notochord induction. These results suggest that the Ras signaling pathway is
essential for the formation of notochord and that another signaling pathway must also be activated
simultaneously in notochord formation during ascidian embryogenesis (Nakatani, 1997).
Hepatocyte Growth Factor/Scatter factor is implicated in branching morphogenesis in a variety of mammalian organs. SF/HGF is a protein with structural similarities to plasminogen. SF/HGF contains four characteristic kringle domains at the N-terminus and a C-terminal, serine protease-like domain. However, amino acids in the active site that are essential for catalytic activites of serine proteases are altered in SF/HGF. Besides being able to induce cellular growth and movement, SF/HGF increases invasiveness of epithelial cells and can induce epithelial cells to make tubular structures. The receptor for SF/HGF, cMet, is a receptor tyrosine kinase. c-met is expressed in epithelial cells and SF/HGF is expressed in mesenchymal cells in close vicinity. SF/HGF expression is rapidly induced when beads soaked in FGF are implantated into the lateral plate mesoderm of the interlimb region of chicken embryos. The initial induction of SF/HGF by FGF does not require limb formation. Expression of SF/HGF during early limb bud stages is found in the entire developing bud and the adjacent lateral plate mesoderm with direct contacts to the lateral edge of the dermomyotome. Later, the SF/HGF expression domain retracts to a distal region below the apical ectodermal ridge. Beads soaked in SF/HGF causes delamination of migratory cells from the dermomyotomal epithelium but no chemotactic attraction of migrating cells toward the SF/HGF source can be detected. It is concluded that SF/HGF is sufficient for delamination of dermomyotomal cells from the epithelium but not for targeted movement toward the SF/HGF source (Heymann, 1996 and references).
FGFs are likely to act to delay differentiation of myoblasts in the early development of limbs. Differentiation of muscle and cartilage within developing vertebrate limbs occurs in a
proximodistal progression. To investigate the cues responsible for regulating muscle
pattern, mouse myoblasts were implanted into early chick wings prior to endogenous chick
muscle differentiation. Fetal myogenic cells originating from transgenic mice carrying a lacZ
reporter are readily detected in vivo after implantation and their state of differentiation
determined with species-specific antibodies to MyoD (Drosophila homolog: Nautilus) and myosin heavy chain. When mouse
myogenic cells are implanted at the growing tip of early stage 21 limbs MyoD expression is
suppressed and little differentiation of the mouse cells is detected initially. At later stages
ectopically implanted mouse cells come to lie within muscle masses, re-express MyoD and
differentiate in parallel with differentiating chick myoblasts. However, if mouse cells are
implanted either proximally at stage 21 or into the limb tip at stage 24, situations in which
mouse cells encounter endogenous differentiating chick myoblasts earlier, MyoD
suppression is not detected and a higher proportion of mouse cells differentiate. Mouse
cells that remain distal to endogenous differentiating myogenic cells are more likely to
remain undifferentiated than myoblasts that lie within differentiated chick muscle.
Undifferentiated distal mouse cells are still capable of differentiating if explanted in vitro,
suggesting that myoblast differentiation is inhibited in vivo. In vitro, MyoD is suppressed in
primary mouse myoblasts by the addition of FGF2 and FGF4 to the culture media. Taken
together, these data suggest that the inhibition of myogenic differentiation in the distal limb
involves MyoD suppression in myoblasts, possibly through an FGF-like activity (Robson, 1996).
Classical embryological experiments suggest that a posterior signal is required for
patterning the developing anteroposterior axis. In this paper, a potential role in Xenopus is investigated
for FGF signaling during this process. During normal development, embryonic fibroblast growth
factor (eFGF) is expressed in the dorsal mesoderm, specifically, in the notochord and in the posterior mesoderm around the closing blastopore. Overexpression of eFGF from the start of gastrulation results in a posteriorised
phenotype of reduced head and enlarged proctodeum. The overexpression of eFGF causes the up-regulation of a
number of posteriorly expressed genes, and prominent among these are Xcad3, a caudal
homolog, and the Hox genes, in particular HoxA7. There is both an increase of expression
within the normal domains and an extension of expression towards the anterior. Application
of eFGF-loaded beads to specific regions of gastrulae reveals that anterior truncations arise
from an effect on the developing dorsal axis. Similar anterior truncations are caused by the
dorsal overexpression of Xcad3 or HoxA7. This suggests that this aspect of the eFGF
overexpression phenotype is caused by the ectopic activation of posterior genes in anterior
regions. Further results using the dominant negative FGF receptor show that the normal expression
of posterior Hox genes is dependent on FGF signaling and that this regulation is likely
mediated by the activation of Xcad3. It has been demonstrated that the eFGF regulates the transcription of Xbra (Drosophila homolog: T-related gene) and that Xbra can in turn activate eFGF expression. Xbra does not directly activate Hox gene expression. However, at the very least, Xbra clearly plays an indirect role in anteroposterior specification through its regulation of eFGF expression in the notochord and the posterior of the embryo. The biological activity of eFGF, together with its
expression in the posterior of the embryo, make it a good candidate to fulfil the role of the
'transforming' activity proposed by Nieuwkoop in his 'activation and transformation' model
for neural patterning (Pownall, 1996).
As with the existence of a notochord, the involvement of an FGF in anterior posterior patterning in vertebrates has no precedence in Drosophila. FGF in Drosophila, and also in vertebrate development, is involved in mesoderm fate. It is likely that the development of the notochord and FGF involvement in anteroposterior axis formation in the vertebrate evolutionary lineage go hand in hand. Involvement of HoxA7, an Antennapedia class homeodomain protein, as well as a Caudal homolog, a protein expressed in the posterior of the fly, as FGF targets suggests that anteroposterior axis development in mammals required the evolutionary development of new regulatory circuits, combining those already utilized by invertebrates into new associations.
Anteroposterior patterning of neural tissue is thought to be directed by the axial mesoderm, which is functionally divided into head (or precordal) and trunk organizer (notochord). In Xenopus the homeobox genes goosecoid (Drosophila homolog: Goosecoid) and Otx2 (Drosophila homolog: Orthodenticle) are expressed in the procordal mesoderm; the LIM class homeobox gene Xlim-1 (Drosophila homolog: Apterous) is expressed in the entire axial mesoderm, whereas the distinct Brachyury related transcription factor Xbra (Drosophila homolog: T-related gene) is expressed in the notochord but not in the procordal mesoderm. Messenger RNA injection experiments show that Xenopus animal pole explants (caps) expressing an activated form of Xlim-1 (a LIM domain mutant named 3m) induce anterior neural markers, whereas caps coexpressing Xlim-1/3m and Xbra induce posterior neural markers. These data indicate that in terms of neural inducing ability, Xlim-1/3m-expressing caps correspond to the head organizer and Xlim-1/3m plus Xbra-coexpressing caps to the trunk organizer. Thus the expression domains of Xlim-1 and Xbra correlate with, and possibly define, the functional domains of the organizer. In animal caps Xlim-1/3m initiates expression of a neuralizing factor chordin (Drosophila homolog: Short gastrulation, which counteracts the antineurogenic effects of Decapentaplegic), whereas Xbra activates embryonic fibroblast growth factor (eFGF expression); these factors could mediate the neural inducing and patterning effects that are observed. A dominant-negative FGF receptor (XFD) inhibits posteriorization by Xbra in a dose-dependent manner, supporting the suggestion that eFGF or a related factor has posteriorizing influence. Retinoic acid, postulated to be a posteriorizing factor based on the observations that RA treatment of embryos leads to truncation of anterior structures in Xenopus, can posteriorize neural tissue generated by Xlim-1. RA strongly inhibits Otx2 expression and induces Krox-20 and beta2-tubulin expression, indicating that RA can act as a posteriorizing factor for neural tissue in the absence of mesoderm (Taira, 1997).
It is generally held that vertebrate muscle precursors depend totally on environmental cues for their development. Instead, it is shown that somites are predisposed toward a particular myogenic program. This predisposition depends on the somite's axial identity: when flank somites are transformed into limb-level somites, either by shifting somitic boundaries with FGF8 or by overexpressing posterior Hox genes, they readily activate the program typical for migratory limb muscle precursors. The intrinsic control over myogenic programs can only be overridden by FGF4 signals provided by the apical ectodermal ridge of a developing limb (L. Alvares, 2003).
Since the competence to start a particular program of hypaxial myogenesis is linked to the position of the somites along the anteroposterior body axis, it was reasoned that this competence may be a consequence of the somite's axial identity. Indeed, when the ability was exploited of FGF8 to move the somitic boundaries, thereby changing the axial identity of somites in the flank into the identity of hindlimb-level somites, these somites gained the ability to produce Lbx1-expressing migratory muscle precursors (MMPs). This suggests that the positional values intrinsic to the somites determine their competence to generate either migratory or nonmigratory hypaxial muscle precursors (L. Alvares, 2003).
It is established that the axial identity of somites is controlled by the overlapping expression of Hox/HOM genes. In particular, a code of Hox gene expression is conserved for crucial anatomical landmarks such as the neck-thorax transition. Significantly, conserved Hox gene expression boundaries also demarcate the transition of limb-flank somites. Moreover, Hox gene expression patterns are maintained when somites are heterotopically grafted, as is the ability to express the MMP marker Lbx1. It was therefore reasoned that Hox genes may control the somitic competence to initiate a particular myogenic program. To test this possibility, expression constructs were engineered for HoxD9 and HoxA10, both normally present in hindlimb-level somites but not in the flank. Upon electroporation of either of the constructs into flank somites, these somites readily expressed Lbx1, while control constructs were unable to evoke expression of the Lbx1 gene. Thus, Lbx1 expression—directly or indirectly—is under the control of Hox genes (L. Alvares, 2003).
Differentiation onset in the vertebrate body axis is controlled by a conserved switch from fibroblast growth factor (FGF) to retinoid signalling, which is also apparent in the extending limb and aberrant in many cancer cell lines. FGF protects tail-end stem zone cells from precocious differentiation by inhibiting retinoid synthesis, whereas later-produced retinoic acid (RA) attenuates FGF signalling and drives differentiation. The timing of RA production is therefore crucial for the preservation of stem zone cells and the continued extension of the body axis. Canonical Wnt signalling mediates the transition from FGF to retinoid signalling in the newly generated chick body axis. FGF promotes Wnt8c expression, which persists in the neuroepithelium as FGF signalling declines. Wnt signals then act here to repress neuronal differentiation. Furthermore, although FGF inhibition of neuronal differentiation involves repression of the RA-responsive gene, retinoic acid receptor β (RARβ), Wnt signals are weaker repressors of neuron production and do not interfere with RA signal transduction. Strikingly, as FGF signals decline in the extending axis, Wnt signals now elicit RA synthesis in neighbouring presomitic mesoderm. This study identifies a directional signalling relay that leads from FGF to retinoid signalling and demonstrates that Wnt signals serve, as cells leave the stem zone, to permit and promote RA activity, providing a mechanism to control the timing of the FGF-RA differentiation switch (Olivera-Martinez, 2007).
The ectoderm gives rise to both neural tissue and epidermis. In vertebrates, specification of the neural plate requires repression of bone morphogenetic protein (BMP) signaling in the dorsal ectoderm. The extracellular BMP antagonist Chordin and other signals from the dorsal mesoderm play important roles in this process. Zebrafish mutant combinations that disrupt Chordin and mesoderm formation were used to reveal additional signals that contribute to the establishment of the neural domain. Fibroblast growth factor (FGF) signaling accounts for the additional activity in neural specification. Impeding FGF signaling results in a shift of ectodermal markers from neural to epidermal. However, following inhibition of FGF signaling, expression of anterior neural markers recovers in a Nodal-dependent fashion. Simultaneously blocking, Chordin, mesoderm formation, and FGF signaling together eliminates neural marker expression during gastrula stages. FGF signaling is required for chordin expression but it also acts via other mechanisms to repress BMP transcription during late blastula stages. Activation of FGF signaling is also able to repress BMP transcription in the absence of protein synthesis. These results support a model in which specification of anterior neural tissue requires early FGF-mediated repression of BMP transcript levels and later activities of Chordin and mesodermal factors (Londin, 2005).
The sensory nervous system in the vertebrate head arises from two different cell populations: neural crest and placodal cells. By contrast, in the trunk it originates from neural crest only. How do placode precursors become restricted exclusively to the head and how do multipotent ectodermal cells make the decision to become placodes or neural crest? At neural plate stages, future placode cells are confined to a narrow band in the head ectoderm, the pre-placodal region (PPR). The head mesoderm is identified as the source of PPR inducing signals, reinforced by factors from the neural plate. Several independent signals are needed: attenuation of BMP and WNT is required for PPR formation. Together with activation of the FGF pathway, BMP and WNT antagonists can induce the PPR in naive ectoderm. WNT
signalling plays a crucial role in restricting placode formation to the head.
Finally, the decision of multipotent cells to become placode or neural crest precursors is demonstrated to be mediated by WNT proteins: activation of the WNT pathway promotes the generation of neural crest at the expense of placodes. This mechanism explains how the placode territory becomes confined to the head, and how neural crest and placode fates diversify (Litsiou, 2005).
This study finds that FGF signalling cooperates with WNT and BMP antagonists to
impart generic placode character to uncommitted ectoderm. In the chick,
activation of the FGF pathway in naive ectoderm leads to rapid expression
of pre-neural markers such as Sox3 and Erni, both of which
are later co-expressed at the border of the neural plate. However, activation
of the FGF pathway is not sufficient to specify cells (neural crest and
placode precursors) that arise from this
border. The observation that continued FGF signalling is
not required for pre-placodal Six4 expression, but can directly
induce Eya2, suggests that FGFs may play a dual role. Early FGF
signalling may confer 'border character' to
ectodermal cells to make them responsive to PPR and crest inducing signals.
The finding that
ectopic PPR induction occurs only in the presence of active FGF signalling
supports this notion. Later, FGFs from the head mesoderm, probably FGF4,
initiate the expression of Eya2 in the placode territory as a crucial
step to activate downstream target genes (Litsiou, 2005).
Simultaneously, the head mesoderm provides both BMP and WNT antagonists,
most likely DAN and Cerberus, to counteract the inhibitory effect of both factors
on the generation of placode precursors. The results show that
attenuation of either the BMP or WNT pathway leads to an expansion of the PPR
into the adjacent ectoderm. However, while the expansion in response to BMP
inhibition is limited to the head ectoderm, WNT antagonism also results in the
expression of PPR specific genes in the trunk. This is in agreement with
recent findings in Xenopus reporting that simultaneous overexpression
of BMP and WNT antagonist expands Six1 expression posteriorly along
the induced secondary axis. In the chick, Wnt8c is expressed in trunk
mesoderm and the mesoderm lateral to the heart primordium, whereas Wnt6
is found in trunk ectoderm. It is
proposed that WNT activity from surrounding tissues is essential to restrict
the placode territory to the head ectoderm next to the neural plate and thus
ensure that sensory placodes are confined to the head. To allow placode
formation, WNT antagonists in cooperation with FGF and anti-BMPs from the head
mesoderm protect placode precursors from this inhibitory influence (Litsiou, 2005).
FGF and the boundaries of the neural plate An investigation was carried out of the cell interactions and signaling molecules involved in setting up and maintaining the border between
the neural plate and the adjacent non-neural ectoderm in the chick embryo at primitive streak stages. msx-1, a target of BMP
signaling, is expressed in this border at a very early stage. It is induced by FGF and by signals from the organizer, Hensen's
node. The node also induces a ring of BMP-4, some distance away. By the early neurula stage, the edge of the neural plate is
the only major site of BMP-4 and msx-1 expression, and is also the only site that responds to BMP inhibition or overexpression.
At this time, the neural plate appears to have a low level of BMP antagonist activity. Using in vivo grafts and in vitro assays, it has been
shown that the position of the border is further maintained by interactions between non-neural and neural ectoderm. It is
concluded that the border develops by integration of signals from the organizer, the developing neural plate, the paraxial
mesoderm and the non-neural epiblast, involving FGFs, BMPs and their inhibitors. It is suggested that BMPs act in an autocrine
way to maintain the border state (Streit, 1999).
The definitive retinal progenitors of the eye field are specified by transcription factors that both promote a retinal fate and control cell movements that are critical for eye field formation. However, the molecular signaling pathways that regulate these movements are largely undefined. Both the FGF and ephrin pathways impact eye field formation. Activating the FGF pathway before gastrulation represses cellular movements in the presumptive anterior neural plate and prevents cells from expressing a retinal fate, independent of mesoderm induction or anterior-posterior patterning. Inhibiting the FGF pathway promotes cell dispersal and significantly increases eye field contribution. EphrinB1 reverse signaling is required to promote cellular movements into the eye field, and can rescue the FGF receptor-induced repression of retinal fate. These results indicate that FGF modulation of ephrin signaling regulates the positioning of retinal progenitor cells within the definitive eye field (Moore, 2004).
Retinal development consists of a series of steps that progressively restrict the available cell fates. First, a subset of embryonic cells are prevented from expressing a retinal fate by inherited maternal factors, whereas others become biased toward retinal fates due to their position within the neural inductive field of the animal hemisphere. As the CNS is regionalized, part of the anterior neural plate is specified as the eye field. Potential retinal progenitors need to be positioned within the eye field to receive the local environmental signals that will direct their ultimate fates. Only after these steps are accomplished do the steps of eye organogenesis, cellular lamination, and phenotype specification occur. Although there has been great progress in understanding how retinal cell type specification occurs, the molecular mechanisms that control which embryonic cells become specified as the definitive retinal progenitors in the eye field remain largely undefined (Moore, 2004).
An accepted hypothesis of how the eye field forms is that signals from surrounding anterior structures regionalize the anterior neural plate. The presumptive eye field then expresses several transcription factors that initiate the retina developmental program, e.g., rx1, pax6, and six3. However, cellular movements during gastrulation and neurulation, directed in part by eye field transcription factors, also are critical, and the signaling factors involved in these early steps of eye field formation have not been identified (Moore, 2004).
Several FGF family members have been implicated in affecting cell movements during gastrulation, and the anterior expression patterns of some FGFs and their receptors are consistent with a role in the morphogenetic movements of eye field cells. Therefore, whether FGF signaling prior to gastrulation plays a role in determining which embryonic cells form the eye field was investigated. Using a constitutively active FGF receptor, enhanced FGF signaling was demonstrated to prevent the normal retinal progenitors from populating the presumptive eye field, suggesting that low levels of FGF signaling are normally required for cells to adopt a retinal fate. This was confirmed by demonstrating that reduced FGF signaling, accomplished by expression of a dominant-negative receptor, enhances the number of cells that become retinal progenitors. It is further reported that ephrinB1 signaling during gastrulation is required for retinal progenitors to move into the eye field, and that this movement can be modified by activating the FGF pathway. These results demonstrate that FGF modulation of ephrin signaling is important for establishing the bona fide retinal progenitors in the anterior neural plate (Moore, 2004).
FGF and left-right asymmetry Left-right asymmetry in vertebrate embryos is first recognisable using molecular markers that encode secreted proteins or
transcription factors. The asymmetry becomes morphologically obvious in the turning of the embryo and in the
development of the heart, the gut and other visceral organs. In the chick embryo, a signaling pathway for the specification
of the left body side has been demonstrated. Sonic hedgehog (Shh) protein is the first asymmetric signal identified in the
node. Further downstream in this pathway are the left-specific genes nodal, lefty-1, lefty-2 and Pitx2. On the right body side, a function of the activin pathway is indicated by the right-sided expression of cActRIIa.
Another key molecule in vertebrate development, fibroblast growth factor 8 (FGF8), is expressed
asymmetrically on the right side of the posterior node. Transcription of FGF8 is induced by activin
and the FGF8 protein inhibits the expression of nodal and Pitx2 and leads to expression of the chicken snail related gene
(cSnR). Left-sided application of FGF8 randomises the direction of heart looping (Boettger, 1999).
FGF and segmentation The postgastrulation functions of FGFs in Xenopus development have been studied by the
implantation in various positions and at various stages of heparin beads soaked in FGF2.
Anterior implantations show different effects depending on whether they are made to
early neurulae or to later stages. At stage 13-14 there is a total or partial suppression of
anterior structures including the forebrain, eyes, and midbrain. From stage 15 onward
there is no loss of anterior parts but there is a change in the structure of the eye such that
the neural retina remains continuous with the wall of the diencephalon and the territories
normally forming the optic stalk and pigment epithelium instead become neural retina.
Posterior implantations cause a disruption of somite segmentation without affecting the
differentiation of muscle cells. This is associated with a prolongation of the uniform
expression of X-Delta-2 during the phase of segmental determination. There is also an
induction of ectopic otocysts, which can lie either ipsilateral or contralateral to the
FGF-bead. The results are discussed in terms of the known late expression domains of
the various Xenopus FGFs, and of the late functions of FGFs in higher vertebrates. They
provide new evidence for a role of endogenous FGFs in the development of the eye,
somites, and otocysts (Lombardo, 1998).
The temporal and spatial regulation of somitogenesis requires a molecular oscillator, the segmentation clock. Through Notch signaling, the oscillation in cells is coordinated and translated into a cyclic wave of expression of hairy-related and other genes. The wave sweeps caudorostrally through the presomitic mesoderm (PSM) and finally arrests at the future segmentation point in the anterior PSM. By experimental manipulation and analyses in zebrafish somitogenesis mutants, a novel component involved in this process has been found. The level of Fgf/MAPK activation (highest in the posterior PSM) serves as a positional cue within the PSM that regulates progression of the cyclic wave and thereby governs the positions of somite boundary formation (Sawada, 2001).
Modulating Fgf signaling resulted in alterations in somite size. Detailed analyses of gene expressions in manipulated wild-type and mutant embryos reveal a novel function of Fgf/MAPK signaling in the PSM: the maintenance of cells in an immature state that allows the her1 wave to sweep through the PSM. Suppression of Fgf signaling posteriorizes the domain shift of her1 expression, as well as the expression of other segmentation genes such as mesp, a bHLH transcription factor crucial for segmentation initiation, and paraxial protocadherin. This leads to a posterior shift in segment border formation and larger somites. These results are complementary to those obtained with transplantation of Fgf beads, strengthening the idea that an Fgf signal determines the position of segment border formation by negatively regulating the maturation of the PSM. Since Fgf signal is known to have profound effects on many developmental processes such as cell growth and maintenance of progenitor cells, it is possible that manipulation of an Fgf signal locally changes the cell number in the PSM by regulating cell proliferation and/or cell migration within the mesoderm (axial, paraxial and lateral plate mesoderm). This could cause alterations in somite size. However, no such effect was observed in manipulated PSM, indicating that an Fgf signal in the PSM simply regulates the maturation status of cells without affecting cell proliferation or migration (Sawada, 2001).
The data are largely consistent with the 'clock-and-wavefront' model in which a cyclic wave operates in conjunction with a maturation wavefront that gradually moves posteriorly, resulting in arrest of the cyclic wave and initiation of segment furrow formation. Fgf/MAPK signaling negatively regulates the wavefront activity and restricts it to the anterior PSM that is devoid of MAPK activation. In zebrafish, the essential components of a conserved somite-making mechanism, the segmentation clock and wavefront are Notch- and fused somites-dependent, respectively. Zebrafish after eight/deltaD mutation desychronizes the oscillation wave, while, in the absence of Fused somites, the anterior PSM fails to acquire the wavefront activity. How could Fgf/MAPK signal interact with these components? In fact, it has been reported that the Ras/MAPK pathway interacts with the Notch pathway in C. elegans vulval development and malignant transformation of cultured cells. However, no interaction between Fgf/MAPK and Notch or Fss pathways could be demonstrated in this study: modulating Fgf signaling exerts identical effects on wild-type and after eight/DeltaD or fused somites mutants in terms of gene expression. Furthermore, the patterns of ERK activation and fgf8 expression in the PSM is not affected by after eight/DeltaD or fused somites mutations. Thus, it is concluded that the activation and action of Fgf/MAPK signaling in the PSM are not mediated by Notch or Fss pathway (Sawada, 2001).
The fact that four to five somites are normally formed after SU5402 treatment indicates that the positioning of furrow formation is already specified or Fgf insensitive at least at the position -IV to -V in the PSM. The result also indicates that ERK activation in segmented somites is not involved in segment border formation. Interestingly, the Fgf-sensitive region corresponds approximately to the heat-shock sensitive zone in zebrafish; that is, the initial defects in the segmental pattern of somite boundaries are observed five somites caudal to the forming somite at the time of heat shock. These data suggest that position -IV to -V represents a position at which the level of Fgf/MAPK activation drops below a threshold, rendering the cells competent to maturation signals. In support of this, transplanted Fgf8 beads exert their effects only when they are located in the Fgf-negative anterior PSM. Importantly, the relative position of MAPK activation domain to the newly formed somite is kept constant in the PSM as the embryos extend. These observations are consistent with the idea that the level of Fgf/MAPK activation serves as a positional cue within the PSM (Sawada, 2001).
Fibroblast growth factor (FGF) signaling plays a crucial role in vertebrate segmentation. The FGF pathway establishes a posterior-to-anterior signaling gradient in the presomitic mesoderm (PSM), which controls cell maturation and is involved in the positioning of segmental boundaries. In addition, FGF signaling was shown to be rhythmically activated in the PSM in response to the segmentation clock. This study shows that conditional deletion of the FGF receptor gene Fgfr1 abolishes FGF signaling in the mouse PSM, resulting in an arrest of the dynamic cyclic gene expression and ultimately leading to an arrest of segmentation. Pharmacological treatments disrupting FGF signaling in the PSM result in an immediate arrest of periodic WNT activation, whereas Notch-dependent oscillations stop only during the next oscillatory cycle. Together, these experiments provide genetic evidence for the role of FGF signaling in segmentation, and identify a signaling hierarchy controlling clock oscillations downstream of FGF signaling in the mouse (Wahl, 2007).
Periodic formation of somites is controlled by the segmentation clock, where the oscillator Hes7 regulates cyclic expression of the Notch modulator Lunatic fringe. This study shows that Hes7 also regulates cyclic expression of the Fgf signaling inhibitor Dusp4/MKP2 (MAP kinase phosphatase 2) and links Notch and Fgf oscillations in phase. Strikingly, inactivation of Notch signaling abolishes the propagation but allows the initiation of Hes7 oscillation. By contrast, transient inactivation of Fgf signaling abolishes the initiation, whereas sustained inactivation abolishes both the initiation and propagation of Hes7 oscillation. It is thus proposed that Hes7 oscillation is initiated by Fgf signaling and propagated/maintained anteriorly by Notch signaling (Niwa, 2007).
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