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Movement and fusion of separate cell populations are critical for several developmental processes, such as neural tube closure in vertebrates or embryonic dorsal closure and pupal thorax closure in Drosophila. Fusion failure results in an opening or groove on the body surface. Drosophila pupal thorax closure is an established model to investigate the mechanism of tissue closure. Here, we report the identification of T-box transcription factor genes Dorsocross (Doc) as Decapentaplegic (Dpp) targets in the leading edge cells of the notum in the late third instar larval and early pupal stages. Reduction of Doc in the notum region results in a thorax closure defect, similar to that in dpp loss-of-function flies. Nine genes are identified as potential downstream targets of Doc in regulating thorax closure by molecular and genetic screens. Our results reveal a novel function of Doc in Drosophila development. The candidate target genes provide new clues for unravelling the mechanism of collective cell movement. [ABSTRACT FROM AUTHOR]

The closely related T-box transcription factors TBX2 and TBX3 are frequently overexpressed in melanoma and various types of human cancers, in particular, breast cancer. The overexpression of TBX2 and TBX3 can have several cellular effects, among them suppression of senescence, promotion of epithelial-mesenchymal transition, and invasive cell motility. In contrast, loss of function of TBX3 and most other human T-box genes causes developmental haploinsufficiency syndromes. Stephens and colleagues (1), by exome sequencing of breast tumor samples, identified five different mutations in TBX3, all affecting the DNA-binding T-domain. One in-frame deletion of a single amino acid, p.N212delN, was observed twice. Due to the clustering of these mutations to the T-domain and for statistical reasons, TBX3 was inferred to be a driver gene in breast cancer. Since mutations in the T-domain generally cause loss of function and because the tumorigenic action of TBX3 has generally been attributed to overexpression, we determined whether the putative driver mutations had loss- or gain-of-function properties. We tested two in-frame deletions, one missense, and one frameshift mutant protein for DNA-binding in vitro, and for target gene repression in cell culture. In addition, we performed an in silico analysis of somatic TBX mutations in breast cancer, collected in The Cancer Genome Atlas (TCGA). Both the experimental and the in silico analysis indicate that the observed mutations predominantly cause loss of TBX3 function. [ABSTRACT FROM AUTHOR]

Organ formation requires a delicate balance of positive and negative regulators. In Drosophila eye development, wingless (wg) is expressed at the lateral margins of the eye disc and serves to block retinal development. The T-box gene optomotor-blind (omb) is expressed in a similar pattern and is regulated by Wg. Omb mediates part of Wg activity in blocking eye development. Omb exerts its function primarily by blocking cell proliferation. These effects occur predominantly in the ventral margin. Our results suggest that the primary effect of Omb is the blocking of Jak/STAT signaling by repressing transcription of upd which encodes the Jak receptor ligand Unpaired. [ABSTRACT FROM AUTHOR]

The horizontal system and vertical system cells of the dipteran optic lobes are well understood regarding their physiology and role in visually guided behavior. Little is known, however, about their development. Drosophila optomotor-blind ( omb) is required for the development of the HS/VS cells which are lacking in the adult brain of the In(1)omb[H31] regulatory mutant. We have analyzed the omb regulatory region, required for HS/VS development, for enhancers active in the central nervous system. A 1-kb fragment, ombJb, was identified 114 kb downstream of the omb transcription start site, that could drive expression in much of the presumptive embryonic optic lobe anlage. Expression in these cells is lost in In(1)omb[H31] suggesting that they contain the HS/VS precursor cell(s). We used Laser ablation in the embryonic CNS in order to localize the position of the HS/VS precursor cell(s) in this tissue. An omb-Gal4 enhancer trap line, which showed activity in the optic lobe anlage in a pattern similar to ombJb enhancer, was used to drive GFP expression, thus allowing to focus the Laser beam to the relevant area. We identified a small region in the embryonic brain from which the HS/VS cells are likely to develop. Omb encodes a transcription factor of the T-box family. Since loss of omb disrupts HS/VS cell development, we assume that HS/VS ontogeny is controlled by Omb target genes. As a first step toward their identification, we characterized the Omb DNA-binding specificity. [ABSTRACT FROM AUTHOR]

The Drosophila wing imaginal disc is subdivided into notum, hinge and blade territories during the third larval instar by formation of several deep apical folds. The molecular mechanisms of these subdivisions and the subsequent initiation of morphogenic processes during metamorphosis are poorly understood. Here, we demonstrate that the Dorsocross (Doc) T-box genes promote the progression of epithelial folds that not only separate the hinge and blade regions of the wing disc but also contribute to metamorphic development by changing cell shapes and bending the wing disc. We found that Doc expression was restricted by two inhibitors, Vestigial and Homothorax, leading to two narrow Doc stripes where the folds separating hinge and blade are forming. Doc mutant clones prevented the lateral extension and deepening of these folds at the larval stage and delayed wing disc bending in the early pupal stage. Ectopic Doc expression was sufficient to generate deep apical folds by causing a basolateral redistribution of the apical microtubule web and a shortening of cells. Cells of both the endogenous blade/hinge folds and of folds elicited by ectopic Doc expression expressed Matrix metalloproteinase 2 (Mmp2). In these folds, integrins and extracellular matrix proteins were depleted. Overexpression of Doc along the blade/hinge folds caused precocious wing disc bending, which could be suppressed by co-expressing MMP2RNAi. [ABSTRACT FROM AUTHOR]

The T-box transcription factors TBX2 and TBX3 are overexpressed in many human cancers raising the need for a thorough understanding of the cellular function of these proteins. In Drosophila, there is one corresponding ortholog, Optomotor-blind (Omb). Currently, only two missense mutations are known for the two human proteins. Making use of the developmental defects caused by inactivation of omb, we have isolated and molecularly characterized four new omb mutations, three of them are missense mutations of amino acids fully conserved in all Tbx proteins. We interpret the functional defects in the framework of the known structure of the human TBX3 protein and provide evidence for loss of Omb DNA-binding activity in all three newly identified missense mutations. [ABSTRACT FROM AUTHOR]

Background: Decapentaplegic (Dpp) is one of the best characterized morphogens, required for dorso-ventral patterning of the Drosophila embryo and for anterior-posterior (A/P) patterning of the wing imaginal disc. In the larval wing pouch, the Dpp target gene optomotor-blind (omb) is generally assumed to be expressed in a step function above a certain threshold of Dpp signaling activity. Results: We show that the transcription factor Omb forms, in fact, a symmetrical gradient on both sides of the A/P compartment boundary. Disruptions of the Omb gradient lead to a re-organization of the epithelial cytoskeleton and to a retraction of cells toward the basal membrane suggesting that the Omb gradient is required for correct epithelial morphology. Moreover, by analysing the shape of omb gain- and loss-of-function clones, we find that Omb promotes cell sorting along the A/P axis in a concentration-dependent manner. Conclusions: Our findings show that Omb distribution in the wing imaginal disc is described by a gradient rather than a step function. Graded Omb expression is necessary for normal cell morphogenesis and cell affinity and sharp spatial discontinuities must be avoided to allow normal wing development. [ABSTRACT FROM AUTHOR]

The “cysteine string protein” (CSP) genes of higher eukaryotes code for a novel family of proteins characterized by a “J” domain and an unusual cysteine-rich region. Previous studies had localized the proteins in neuropil and synaptic terminals of larval and adult Drosophila and linked the temperature-sensitive paralysis of the mutants described here to conditional failure of synaptic transmission. We now use the null mutants as negative controls in order to reliably detect even low concentrations of CSPs by immunohistochemistry, employing three monoclonal antibodies. In wild-type flies high levels of cysteine string proteins are found not only in apparently all synaptic terminals of the embryonic, larval, and adult nervous systems, but also in the “tall cells” of the cardia, in the follicle cells of the ovary, in specific structures of the female spermatheca, and in the male testis and ejaculatory bulb. In addition, low levels of CSPs appear to be present in all tissues examined, including neuronal perikarya, axons, muscles, Malpighian tubules, and salivary glands. Western blots of isolated tissues demonstrate that of the four isoforms expressed in heads only the largest is found in non-neural organs. The wide expression of CSPs suggests that at least some of the various phenotypes of the null mutants observed at permissive temperatures, such as delayed development, short adult lifespan, modified electroretinogram, and optomotor behavior, may be caused by the lack of CSPs outside synaptic terminals. [ABSTRACT FROM AUTHOR]

Compartment boundaries prevent cell populations of different lineage from intermingling. In many cases, compartment boundaries are associated with morphological folds. However, in the Drosophila wing imaginal disc, fold formation at the anterior/posterior (A/P) compartment boundary is suppressed, probably as a prerequisite for the formation of a flat wing surface. Fold suppression depends on optomotor-blind (omb). Omb mutant animals develop a deep apical fold at the A/P boundary of the larval wing disc and an A/P cleft in the adult wing. A/P fold formation is controlled by different signaling pathways. Jun N-terminal kinase (JNK) and Yorkie (Yki) signaling are activated in cells along the fold and are necessary for the A/P fold to develop. While JNK promotes cell shape changes and cell death, Yki target genes are required to antagonize apoptosis, explaining why both pathways need to be active for the formation of a stable fold. [ABSTRACT FROM AUTHOR]