Requirement of teashirt (tsh) function during cell fate specification in developing head structures in Drosophila

 The homeotic gene teashirt (tsh) is known to regulate segmental identity of the trunk region of the Drosophila embryo. Here we report a requirement for tsh function in the development of adult head structures. Animals homozygous for a viable tsh allele or heterozygous for various embryonic recessive lethal alleles displayed miniaturized maxillary palps, a phenotype characteristically induced by dominant gain-of-function mutations of Antennapedia (Antp) homeotic gene. Animals transheterozygous for tsh and Antp mutations displayed an enhanced antenna-to-leg and a striking reduced-eye phenotype suggesting aggravated ANTP misexpression in eye-antennal discs of these animals. In agreement with this, in the developing eye-antennal discs of the tsh mutant animals a significant amount of ANTP protein was detected overlapping the domains where tsh is normally expressed. These results suggest that tsh specifies adult head segments by repressing Antp expression. Received: 7 December 1996 / Accepted: 8 April 1997     

Three putative murine Teashirt orthologues specify trunk structures in Drosophila in the same way as the Drosophila teashirt gene

Drosophila teashirt (tsh) functions as a region-specific homeotic gene that specifies trunk identity during embryogenesis. Based on sequence homology, three tsh-like (Tsh) genes have been identified in the mouse. Their expression patterns in specific regions of the trunk, limbs and gut raise the possibility that they may play similar roles to tsh in flies. By expressing the putative mouse Tsh genes in flies, we provide evidence that they behave in a very similar way to the fly tsh gene. First, ectopic expression of any of the three mouse Tsh genes, like that of tsh, induces head to trunk homeotic transformation. Second, mouse Tsh proteins can rescue both the homeotic and the segment polarity phenotypes of a tsh null mutant. Third, following ectopic expression, the three mouse Tsh genes affect the expression of the same target genes as tsh in the Drosophila embryo. Fourth, mouse Tsh genes, like tsh, are able to induce ectopic eyes in adult flies. Finally, all Tsh proteins contain a motif that recruits the C-terminal binding protein and contributes to their repression function. As no other vertebrate or fly protein has been shown to induce such effects upon ectopic expression, these results are consistent with the idea that the three mouse Tsh genes are functionally equivalent to the Drosophila tsh gene when expressed in developing Drosophila embryos.     

A gene that regulates the bithorax complex differentially in larval and adult cells of Drosophila

Loss-of-function mutations of a new homeotic gene, sxc, in Drosophila cause transformations of body segments, suggesting inappropriate expression of BX-C and ANT-C genes. I present evidence that sxc+ is required during embryogenesis for the selective repression of the BX-C in different larval segments and show that this requirement may be met entirely by maternally derived gene product. sxc+ is also required later in development to ensure the appropriate expression of ANT-C and BX-C genes in adult thoracic and abdominal segments. Absence of sxc+ in the mesothorax apparently results in the ectopic expression of the bx+ (or Ubx+) function in both the anterior and posterior compartments; this suggests that pbx mutations may define a regulatory rather than a structural function.     

Mouse Hox-2.2 specifies thoracic segmental identity in Drosophila embryos and larvae

The mouse genome has a number of homeobox genes that are structurally similar to the Drosophila Antenapedia (Antp) gene. We find that one of the mouse Antp-like genes, Hox-2.2, when expressed in developing Drosophila cells under control of a heat shock promoter, can induce homeotic transformations that are nearly identical to those caused by ectopic expression of Antp. In larvae, the Hox-2.2-induced transformations include thoracic denticle belts in place of head structures; in adults, the Hox-2.2 transformations include thoracic legs in place of antennae. The phenotypic effects of Hox-2.2 do not depend on the endogenous Antp gene, whose spatial limits of expression are unaffected by Hox-2.2 expression. Thus, in the Drosophila embryo, Hox-2.2 can substitute for some of the segmental identity functions of Antp, presumably by regulating the same set of downstream genes.     

Homeotic genes have specific functional roles in the establishment of the Drosophila embryonic peripheral nervous system.

The Drosophila embryonic peripheral nervous system (PNS) contains segment-specific spatial patterns of sensory organs which derive from the ectoderm. Many studies have established that the homeotic genes of Drosophila control segment specific characteristics of the epidermis, and more recently these genes have also been shown to control gut morphogenesis through their expression in the visceral mesoderm (Tremml, G. and Bienz, M. (1989), EMBO J. 8, 2677-2685). We report here the roles of homeotic genes in establishing the spatial patterns of sensory organs in the embryonic PNS. The PNS was examined in embryos homozygous for mutations in the homeotic genes Sex combs reduced (Scr), Antennapedia (Antp), Ultrabithorax (Ubx), abdominal-A (abd-A) and Abdominal-B (Abd-B) with antibodies that label specific subsets of sensory organs. Our results suggest that the homeotic genes have specific roles in establishing the correct spatial patterns of sensory organs in their normal domains of expression. In addition, we also report the effects of ectopic expression of the homeotic genes labial (lab), Deformed (Dfd), Scr, Antp or Ubx on the normal development of sensory organs in the embryonic PNS. Interestingly, while previous studies have concluded that ectopic expression of the homeotic genes Dfd, Scr and Antp has no effect on the segmental identity of the abdominal segments, our results demonstrate that this is not true. We show that ectopic expression of these genes does result in the disruption of the developing PNS in the abdomen. Our results are suggestive of a role for the homeotic gene products in regulating genes which are necessary for generating sensory progenitor cells in the developing PNS.     

Gap genes define the limits of antennapedia and bithorax gene expression during early development in Drosophila.

The maintenance of selective patterns of homeotic gene expression within the Drosophila CNS involves cross-regulatory interactions among the genes of the antennapedia and bithorax complexes (ANT-C and BX-C). Such a mechanism does not appear to be responsible for the establishment of these selective expression patterns during early development. Here we show that mutations in several of the gap genes strongly alter the early patterns of Antp and Abd-B expression. The altered patterns that are observed do not always correlate with simple expectations based on cuticular pattern defects observed in advanced-stage mutants. It appears that the initial patterns of Antp and Abd-B expression involve their differential regulation by a common set of gap genes. We propose that the gap genes are largely responsible for integrating the processes of segmentation and homeosis.     

Ectopic Expression of the Drosophila Homeotic Gene Proboscipedia under Antennapedia P1 Control Causes Dominant Thoracic Defects

A deletion mutation in the Antennapedia Complex of Drosophila melanogaster, Df(3R)SCBXL2, induces both dominant and recessive loss-of-function phenotypes. The deletion is associated with diminished function of proboscipedia (pb), a homeotic gene required for mouthparts formation. Df(3R)SCBXL2 also has associated dominant thoracic defects related to diminished expression of the homeotic Antennapedia (Antp) gene copy on the homologous chromosome. This is shown to be a consequence of ectopic pb expression in the thorax. Newly juxtaposed Antp sequences provide the pb gene on the deletion bearing chromosome with a second promoter, Antp P1, in addition to its own. Ectopic pb protein expression occurs under Antp P1 control, by alternate splicing, and results in diminished accumulation of Antp protein in the imaginal disc cells where Antp P1 is normally expressed. The analysis of this mutant chromosome thus demonstrates that pb protein is capable of participating in the negative regulation of a more posteriorly expressed homeotic gene, as well as serving a homeotic "selector" function in the head.     

The localization and regulation of Antennapedia protein expression in Drosophila embryos

The homeotic Antennapedia (Antp) gene of Drosophila is required for the normal differentiation of the thoracic segments during embryonic development and metamorphosis. Antibodies to a recombinant Antp protein were used to localize the protein in whole mount embryos. Antp is expressed in the nuclei of cells of the thoracic embryonic epidermis and several segments of the ventral and peripheral nervous systems. Analysis of Antp expression in mutant embryos revealed three levels of Antp regulation by genes of the bithorax complex, pleiotropic homeotic loci, and Antp itself. The distributions of the Antp and the Ultrabithorax (Ubx) proteins in doubly-labeled embryos suggest that the Ubx protein may be one direct negative regulator of Antp gene expression.     

Ectopic expression of homeotic genes caused by the elimination of the Polycomb gene in Drosophila imaginal epidermis

The morphological patterns in the adult cuticle of Drosophila are determined principally by the homeotic genes of the bithorax and Antennapedia complexes. We find that many of these genes become indiscriminately active in the adult epidermis when the Pc gene is eliminated. By using the Pc3 mutation and various BX-C mutant combinations, we have generated clones of imaginal cells possessing different combinations of active homeotic genes. We find that, in the absence of BX-C genes, Pc- clones develop prothoracic patterns; this is probably due to the activity of Sex combs reduced which overrules Antennapedia. Adding contributions of Ultrabithorax, abdominal-A and Abdominal-B results in thoracic or abdominal patterns. We have established a hierarchical order among these genes: Antp less than Scr less than Ubx less than abd-A less than Abd-B. In addition, we show that the engrailed gene is ectopically active in Pc- imaginal cells.     

Rescue and regulation of proboscipedia: a homeotic gene of the Antennapedia Complex.

The extraordinary positional conservation of the homeotic genes within the Antennapedia and the Bithorax Complexes (ANT-C and BX-C) in Drosophila melanogaster and the murine Hox and human HOX clusters of genes can be interpreted as a reflection of functional necessity. The homeotic gene proboscipedia (pb) resides within the ANT-C, and its sequence is related to that of Hox-1.5. We show that two independent pb minigene P-element insertion lines completely rescue the labial palp-to-first leg homeotic transformation caused by pb null mutations; thus, a homeotic gene of the ANT-C can properly carry out its homeotic function outside of the complex. Despite the complete rescue of the null, the minigene expresses pb protein in only a subset of pb's normal domains of expression. Therefore, the biological significance of the excluded expression pattern elements remains unclear except to note they appear unnecessary for specifying normal labial identity. Additionally, by using reporter gene constructs inserted into the Drosophila genome and by comparing pb-associated genomic sequences from two divergent species, we have located cis-acting regulatory elements required for pb expression in embryos and larvae.     

Head and tail development of the Drosophila embryo involves spalt, a novel homeotic gene

Mutations in spalt (sal), a novel homeotic gene on the second chromosome of Drosophila, cause opposite transformations in two subterminal regions of the embryo: posterior head segments are transformed into anterior thoracic structures and anterior tail segments are transformed into posterior abdominal structures. The embryonic phenotypes of double mutants for sal and various Antennapedia (ANT-C) or bithorax (BX-C) genes indicate that sal acts independently of the hierarchical order of the latter gene complexes. Trans-regulatory gene mutations causing ectopic expression of ANT-C and BX-C genes do not change the realms of sal action. It is proposed that the region-specific action of the sal gene primarily promotes head as opposed to trunk development, while the BX-C gene AbdB distinguishes tail from head.     

Persistent ectopic expression of Drosophila homeotic genes resulting from maternal deficiency of the extra sex combs gene product

Like other members of the Polycomb group, the extra sex combs gene (esc) is required for the correct repression of loci in the major homeotic gene complexes. We show here that embryos lacking both maternal and zygotic esc+ function display transient, general derepression of both the Ultrabithorax (Ubx) and Antennapedia (Antp) genes during germ band shortening, but Sex combs reduced (Scr) expression is almost normal in the epidermis and lacking in the central nervous system (CNS). In addition, embryos that are maternally esc- but receive two paternal copies of esc+ often are characterized by ectopic expression of the three homeotic genes, especially Ubx and Antp in the CNS. Imaginal discs from these paternally rescued embryos may show discrete patches of expression of Ubx and Scr in inappropriate locations. Thus, lack of esc+ function during a brief period in early embryogenesis results in a heritable change in determined state, even in a genetically wild type animal. Within these ectopic patches, homeotic gene expression may be regulated by the disc positional fields and by cross-regulatory interactions between homeotic genes.     

Establishment of imaginal discs and histoblast nests in Drosophila

In Drosophila the homeotic genes of the bithorax-complex (BX-C) and Antennapedia-complex (ANT-C) specify the identity of segments. Adult segment primordia are established in the embryo as the histoblast nests of the abdomen and the imaginal discs of the head, thorax and terminalia. We have used a molecular probe for the limb primordia and in vivo culture to describe the nature of the adult primordia in mutants in which the pattern of homeotic gene expression was altered. The results suggest that the histoblast or disc 'mode' of development is initiated by the extended germ band stage through activity of the BX-C and ANT-C and is relatively inflexible thereafter [corrected].     

Direct control of antennal identity by the spineless-aristapedia gene of Drosophila

Loss-of-function mutations in the spineless-aristapedia gene of Drosophila (ssa mutants) cause transformations of the distal antenna to distal second leg, deletions or fusions of the tarsi from all three legs, a general reduction in bristle size, and sterility. Because ssa mutants are pleiotropic, it has been suggested that ss+ has some rather general function and that the ssa antennal transformation is an indirect consequence of perturbations in the expression of other genes that more directly control antennal or second leg identity. Here we test whether the ssa transformation results from aberrant expression of Antennapedia (Antp), a homeotic gene thought to specify directly the identity of the second thoracic segment. We find that Antp-ssa mitotic recombination clones in the distal antenna behave identically to Antp+ ssa clones, and are transformed to second leg. This demonstrates that the ssa antennal transformation is independent of Antp+, and suggests that ss+ may itself directly define distal antennal identity. The results also reveal that Antp+ is not required for the development of distal second leg structures, as these develop apparently normally in Antp- ssa antennal clones. Because Antp- mutations cause deletions or transformations that are restricted to proximal structures, whereas ssa alleles cause similar defects that are distally restricted, we suggest that ss+ and Antp+ may play similar, but complementary, roles in the distal and proximal portions of appendages, respectively.