Control and function of terminal gap gene activity in the posterior pole region of the Drosophila embryo.

We have studied the genetic requirement for the normal expression of the terminal gap genes huckebein (hkb) and tailless (tll) and their possible function in the posterior pole region of the Drosophila embryo. At the early blastoderm stage, both genes are expressed in largely coextensive expression domains. Our results show that in the posterior region of the embryo both the activation and the control of the spatial limits of tll and hkb expression are critically dependent on torso (tor) activity, which is thought to be a crucial component of a cellular signal transduction pathway provided by the terminal maternal system. Furthermore, the spatial control of hkb and tll expression does not require mutual interactions among each other, nor does it require regulatory input from other gap genes which are essential for the establishment of segmentation in the trunk region of the embryo ("central gap genes"). Therefore, the terminal gap genes have unique regulatory features which are distinct from the central gap genes. In the absence of terminal gap gene activities, as in hkb and tll mutant embryos, the expression domains of the central gap genes expand posteriorly, indicating that the terminal gap gene activities prevent central gap gene expression in the posterior pole region of the wildtype embryo. This, in turn, suggests that the terminal gap gene activities prevent metamerization by repression of central gap genes, thereby distinguishing the segmented trunk from the nonsegmented tail region of the embryo.     

Pole region-dependent repression of the Drosophila gap gene Krüppel by maternal gene products

We examined the protein domain of the gap gene Krüppel (Kr) in mutants that affect the establishment of different regions of the segment pattern along the longitudinal axis of the Drosophila embryo. Our data suggest that Kr provides cues for establishing the "central" pattern elements at the blastoderm stage, and that Kr activity is controlled by maternal effect genes acting at the poles. The formation of the Kr protein domain may involve ubiquitous activation of Kr gene expression which, however, is limited by region-specific repression through the action of the maternal anterior and posterior pattern organizer genes. In addition, the formation of the Kr protein domain depends on the activity of gap genes acting adjacent to the Kr domain, but it is independent of subordinate pair-rule gene activities.     

Cell and cubitus interruptus dominant: two segment polarity genes on the fourth chromosome in Drosophila

The cubitus interruptus Dominant 4-O [ciD] locus is a member of a class of genes required for the proper formation of the repeating segmental pattern of the embryo. We have found a second locus on the fourth chromosome, Cell (Ce), which is also required for proper segmentation. Mutations in Ce cause the elimination of the posterior half and anterior margin of each segment. The complementation pattern of ciD and Ce alleles is complicated; certain alleles complement fully while others do not. We have isolated a series of translocations which have breakpoints within the ciD gene. These mutations also disrupt Ce2 function, indicating that ciD and Ce2 are intimately associated. We have also studied the maternal contribution of the wild-type gene product of the ciD and Ce genes by pole cell transplants. These experiments suggest that there is no maternal contribution of the ciD+ and Ce+ gene products, and that they are not required for oogenesis. Thus, it appears that the ciD and Ce loci may represent a gene complex that is activated during zygotic development and is required for the proper formation of segments.     

The Drosophila gene tailless is expressed at the embryonic termini and is a member of the steroid receptor superfamily

The zygotically active tailless (tll) gene plays a key role in the establishment of nonmetameric domains at the anterior and posterior poles of the Drosophila embryo. We have cloned the tll gene and show that it encodes a protein with striking similarity to steroid hormone receptors in both the DNA binding "finger" and ligand binding domains. tll RNA is initially expressed in embryos in two mirror-image symmetrical domains; this pattern then quickly resolves into a pattern consistent with the mutant phenotype: a posterior cap and an anterior dorsal stripe. That the tll gene may also play a role in the nervous system is suggested by its strong expression in the forming brain and transient expression in the peripheral nervous system.     

Spatial regulation of the gap gene giant during Drosophila development

We describe the regulated expression of the segmentation gene giant (gt) during early embryogenesis. The gt protein is expressed in two broad gradients in precellular embryos, one in anterior regions and the other in posterior regions. Double immunolocalization studies show that the gt patterns overlap with protein gradients specified by the gap genes hunchback (hb) and knirps (kni). Analysis of all known gap mutants, as well as mutations that disrupt each of the maternal organizing centers, indicate that maternal factors are responsible for initiating gt expression, while gap genes participate in the subsequent refinement of the pattern. The maternal morphogen bicoid (bcd) initiates the anterior gt pattern, while nanos (nos) plays a role in the posterior pattern. Gene dosage studies indicate that different thresholds of the bcd gradient might trigger hb and gt expression, resulting in overlapping but noncoincident patterns of expression. We also present evidence that different concentrations of hb protein are instructive in defining the limits of kni and gt expression within the presumptive abdomen. These results suggest that gt is a bona fide gap gene, which acts with hb, Krüppel and kni to initiate striped patterns of gene expression in the early embryo.     

l(1)pole hole is required maternally for pattern formation in the terminal regions of the embryo

Maternal expression of the l(1)pole hole (l(1)ph) gene product is required for the development of the Drosophila embryo. When maternal l(1)ph+ activity is absent, alterations in the embryonic fate map occur as visualized by the expression of segmentation genes fushitarazu and engrailed. If both maternal and zygotic activity is absent, embryos degenerate around 7 h of development. If only maternal activity is missing, embryos complete embryogenesis and show deletions of both anterior and posterior structures. Anteriorly, structures originating from labral and acron head regions are missing. Posteriorly, abdominal segments A8, 9 and 10, the telson and the proctodeum are missing. Similar pattern deletions are observed in embryos derived from the terminal class of female sterile mutations. Thus, the maternal l(1)ph+ gene product is required for the establishment of cell identities at the anterior and posterior poles of the Drosophila embryo.     

Interaction between torso and dorsal, two elements of different transduction pathways in the Drosophila embryo.

Early development of the Drosophila embryo is under the control of some maternal genes responsible for the establishment of its general pattern. Three sets of genes determine the anteroposterior pattern; two distinct systems specify anterior and posterior development and a third one, the terminal system, is responsible for the development of the poles of the embryo. A different set of genes specifies dorsoventral polarity, which is established by the graded activity of the dorsal gene product. Here I analyze the effect of the terminal system on the expression of two zygotic genes involved in dorsoventral pattern, snail and decapentaplegic, and I show that this effect is mediated by a reduction on dorsal activity by the terminal system. Due to the interaction of these two systems, both of which use transmembrane signalling mechanisms, the poles adopt a more dorsalized fate than their counterparts in the middle of the embryo.     

Maternal-effect genes that alter the fate map of the Drosophila blastoderm embryo

The pattern of segmentation in the Drosophila embryo is controlled by at least 25 zygotically active genes and at least 20 maternally active genes. We have examined the pattern of expression of the protein product of the zygotically active segmentation gene fushi tarazu (ftz) at the cellular blastoderm stage in progeny of mutant females homozygous for each of six maternal-effect segmentation genes to observe the early effects of the maternal-effect genes on zygotic gene expression. The genes included exuperantia (a member of the anterior class of maternal-effect segmentation genes); staufen and vasa (members of the posterior class); and torso, trunk, and fs(1)N (members of the terminal class). Mutations in the genes caused a disruption of the normal pattern of ftz stripes in regions of the embryo where gene activity is known to be required. The ftz stripes provide a marker for segmental determination at the cellular blastoderm stage, making it possible to correlate aberrant patterns of ftz protein with defects in cuticle morphology at the end of embryogenesis. ftz protein expression in progeny of females mutant for combinations of the above genes was also examined. The changes in the ftz pattern in progeny of females doubly mutant for genes of the anterior and terminal classes or of the posterior and terminal classes can largely be understood as the result of the additive effects of the single mutations. In contrast, clearly nonadditive effects on the ftz pattern were seen when a mutation in a gene of the anterior class (exuperantia) was combined with mutations in posterior class genes.     

The egg came first, of course! Anterior-posterior pattern formation in Drosophila embryogenesis and oogenesis

The anterior-posterior body pattern of the Drosophila embryo is initiated through the action of maternal gene products. In particular, three groups of maternally acting genes (the anterior, posterior and terminal groups) have been shown to direct the synthesis and spatial restriction of the three major organizing activities in the egg. The initial spatial localizations of the maternal organizing activities are established during oogenesis. After fertilization these activities regulate zygotic gene activity along the anterior-posterior axis of the egg.     

Pattern formation under the control of the terminal system in the Drosophila embryo

The specification of the most anterior and posterior domains of the Drosophila embryo depends on the activity of the torso protein, a putative tyrosine kinase receptor. Localized torso activity at the poles of the embryo generates graded information that specifies distinct portions of the body. The primary response to the terminal signal in the posterior end of the embryo is likely to be the activation of the gap genes huckebein and tailless. Here I address the question of how the graded maternal signal generates different elements of the pattern at the posterior end of the embryo and what role huckebein and tailless activities may play in this process. These experiments show that distinctly localized activities of huckebein and tailless are responsible for the appropriate expression of other genes known to be under the control of the terminal system. Moreover, they suggest that different elements of the terminal pattern can be specified in response to distinct levels of graded tailless activity.     

Localized enhancement and repression of the activity of the TGF-beta family member, decapentaplegic, is necessary for dorsal-ventral pattern formation in the Drosophila embryo.

Seven zygotically active genes are required for normal patterning of the dorsal 40% of the Drosophila embryo. Among these genes, decapentaplegic (dpp) has the strongest mutant phenotype: in the absence of dpp, all cells in the dorsal and dorsolateral regions of the embryo adopt fates characteristic of more ventrally derived cells (Irish and Gelbart (1987) Genes Dev. 1, 868-879). Here we describe the phenotypes caused by alleles of another of this set of genes, tolloid, and show that tolloid is required for dorsal, but not dorsolateral, pattern. Extragenic suppressors of tolloid mutations were isolated that proved to be mutations that elevate dpp activity. We studied the relationship between tolloid and dpp by analyzing the phenotypes of tolloid embryos with elevated numbers of the dpp gene and found that doubling the dpp+ gene dosage completely suppressed weak tolloid mutants and partially suppressed the phenotypes of tolloid null mutants. We conclude that the function of tolloid is to increase dpp activity. We also examined the effect of doubling dpp+ gene dosage on the phenotypes caused by other mutations affecting dorsal development. Like tolloid, the phenotypes of mutant embryos lacking shrew gene function were suppressed by elevated dpp, indicating that shrew also acts upstream of dpp to increase dpp activity. In contrast, increasing the number of copies of the dpp gene enhanced the short gastrulation (sog) mutant phenotype, causing ventrolateral cells to adopt dorsal fates. This indicates that sog gene product normally blocks dpp activity ventrally. We propose that the tolloid, shrew and sog genes are required to generate a gradient of dpp activity, which directly specifies the pattern of the dorsal 40% of the embryo.     

Nanos is the localized posterior determinant in Drosophila

Segmental pattern in the Drosophila embryo is established by two maternal factors localized to the anterior and posterior poles of the egg cell. Here we provide molecular evidence that the localized posterior factor is the RNA of the nanos (nos) gene. nos RNA is localized to the posterior pole of early embryos, and nos protein acts at a distance to direct abdomen formation. Synthetic nos RNA has biological activity identical to that of the posterior pole plasm. Injection of nos RNA rescues the segmentation defect of embryos derived from females mutant for all nine known posterior group genes. Injection of nos RNA into the anterior is able to direct formation of ectopic posterior structures. Our results demonstrate that a localized source of nos RNA is sufficient to specify abdominal segmentation and imply that other posterior group genes are required for localization, stabilization, or distribution of the nos gene product.     

Gene Circuit Analysis of the Terminal Gap Gene huckebein

The early embryo of Drosophila melanogaster provides a powerful model system to study the role of genes in pattern formation. The gap gene network constitutes the first zygotic regulatory tier in the hierarchy of the segmentation genes involved in specifying the position of body segments. Here, we use an integrative, systems-level approach to investigate the regulatory effect of the terminal gap gene huckebein (hkb) on gap gene expression. We present quantitative expression data for the Hkb protein, which enable us to include hkb in gap gene circuit models. Gap gene circuits are mathematical models of gene networks used as computational tools to extract regulatory information from spatial expression data. This is achieved by fitting the model to gap gene expression patterns, in order to obtain estimates for regulatory parameters which predict a specific network topology. We show how considering variability in the data combined with analysis of parameter determinability significantly improves the biological relevance and consistency of the approach. Our models are in agreement with earlier results, which they extend in two important respects: First, we show that Hkb is involved in the regulation of the posterior hunchback (hb) domain, but does not have any other essential function. Specifically, Hkb is required for the anterior shift in the posterior border of this domain, which is now reproduced correctly in our models. Second, gap gene circuits presented here are able to reproduce mutants of terminal gap genes, while previously published models were unable to reproduce any null mutants correctly. As a consequence, our models now capture the expression dynamics of all posterior gap genes and some variational properties of the system correctly. This is an important step towards a better, quantitative understanding of the developmental and evolutionary dynamics of the gap gene network.     

Defects of the body plan of mutant embryos lacking Lim1, Otx2 or Hnf3beta activity

The orientation of the anterior-posterior (A-P) axis was examined in gastrula-stage Hnf3beta, Otx2 and Lim1 null mutant embryos that display defective axis development. In situ hybridization analysis of the expression pattern of genes associated with the posterior germ layer tissues and the primitive streak (T, Wnt3 and Fgf8) and anterior endoderm (Cer1 and Sox17) revealed that the A-P axis of mutant embryos remains aligned with the proximo-distal plane of the gastrula. Further analysis revealed that cells which express Chrd activity are either absent in Hnf3beta mutant embryos or localised in heterotopic sites in Lim1 and Otx2 null mutants. Lim1-expressing cells are present in the Hnf3beta mutant embryo albeit in heterotopic sites. In all three mutants, Gsc-expressing cells are missing from the anterior mesendoderm. These findings suggest that although some cells with organizer activity may be present in the mutant embryo, they are not properly localised and fail to contribute to the axial mesoderm of the head. By contrast, in T/T mutant embryos that display normal head fold development, the expression domains of organizer, primitive streak and anterior endoderm genes are regionalised correctly in the gastrula.     

Diversity in insect axis formation: two orthodenticle genes and hunchback act in anterior patterning and influence dorsoventral organization in the honeybee (Apis mellifera)

Axis formation is a key step in development, but studies indicate that genes involved in insect axis formation are relatively fast evolving. Orthodenticle genes have conserved roles, often with hunchback, in maternal anterior patterning in several insect species. We show that two orthodenticle genes, otd1 and otd2, and hunchback act as maternal anterior patterning genes in the honeybee (Apis mellifera) but, unlike other insects, act to pattern the majority of the anteroposterior axis. These genes regulate the expression domains of anterior, central and posterior gap genes and may directly regulate the anterior gap gene giant. We show otd1 and hunchback also influence dorsoventral patterning by regulating zerknült (zen) as they do in Tribolium, but that zen does not regulate the expression of honeybee gap genes. This suggests that interactions between anteroposterior and dorsal-ventral patterning are ancestral in holometabolous insects. Honeybee axis formation, and the function of the conserved anterior patterning gene orthodenticle, displays unique characters that indicate that, even when conserved genes pattern the axis, their regulatory interactions differ within orders of insects, consistent with relatively fast evolution in axis formation pathways.     

High bicoid levels render the terminal system dispensable for Drosophila head development

In Drosophila, the gradient of the Bicoid (Bcd) morphogen organizes the anteroposterior axis while the ends of the embryo are patterned by the maternal terminal system. At the posterior pole, expression of terminal gap genes is mediated by the local activation of the Torso receptor tyrosine kinase (Tor). At the anterior, terminal gap genes are also activated by the Tor pathway but Bcd contributes to their activation. Here we present evidence that Tor and Bcd act independently on common target genes in an additive manner. Furthermore, we show that the terminal maternal system is not required for proper head development, since high levels of Bcd activity can functionally rescue the lack of terminal system activity at the anterior pole. This observation is consistent with a recent evolution of an anterior morphogenetic center consisting of Bcd and anterior Tor function.