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. 2000 Sep 1;14(17):2140–2145. doi: 10.1101/gad.180900

Single mesodermal cells guide outgrowth of ectodermal tubular structures in Drosophila

Christian Wolf 1, Reinhard Schuh 1,1
PMCID: PMC316895  PMID: 10970878

Abstract

The Drosophila tracheal system, a tubular network, is formed from isolated ectodermal metameres by guided branch outgrowth and branch fusion. Branch outgrowth is triggered by the localized and transient activity of Branchless (Bnl/dFGF). Here, we report the discovery of a mesodermal cell that links the leading cells of outgrowing main branches 2.5 hr before they fuse. This bridge-cell serves as an essential guidance post and needs Hunchback (Hb) activity to exert its function. The bridge-cell provides cues acting in concert with Bnl/dFGF signaling to mediate directed branch outgrowth that ultimately leads to position-specific branch fusion.

Keywords: hunchback, FGF, tracheal system, Drosophila, guidance

Formation of three-dimensional tubular structures, such as the insect tracheal system (Manning and Krasnow 1993; Samakovlis et al. 1996), the vertebrate vascular system (Risau 1997), and the lung (Hogan et al. 1997), involves the guided outgrowth of epithelial cells. In Drosophila, the tracheal system is generated from 10 isolated lateral cell clusters on each side of the embryo (Fig. 1A). These cell clusters, which are each composed of about 80 ectodermal cells, invaginate in a strictly coordinated manner into the underlying mesoderm, where they establish a pattern of six primary tubular branches (Fig. 1B). Some of these branches grow along the dorsoventral body axis to form the dorsal, the lateral, and the ganglionic branches. Additional primary branches extend along the anteroposterior axis to generate the visceral and dorsal trunk anterior and posterior branches. The individual tracheal cell clusters connect by fusion of the dorsal trunk and the lateral trunk branches (Fig. 1C). The two halves of the network interconnect by anastomosis formation, and the three-dimensional system starts with the transport of gases during larval development (for details, see Manning and Krasnow 1993; Samakovlis et al. 1996).

Figure 1.

Figure 1

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A non-tracheal mesodermal cell connects outgrowing tracheal dorsal trunk branches 2.5 hr prior to their fusion. (A–C) Whole-mount in situ hybridization of 1-eve-1 embryos at stage 11 (A), stage 12 (B), and stage 13 (C) with a lacZ antisense RNA probe. 1-eve-1 embryos reveal lacZ marker gene expression in the tracheal cells. (D–H) Whole-mount in situ double hybridization of 1-eve-1 embryos at stage 11 (D), early stage 12 (E), late stage 12 (F), stage 13 (G), and stage 14 (H) with lacZ (red) and hb (blue) antisense RNA probes; lateral view focusing on tracheal metameres 2 and 3. Arrowhead in E points to a bridge-cell before and arrows in E and F point to bridge-cells after connection to dorsal trunk anterior branches. (A) Dorsal trunk anterior branch; (P) dorsal trunk posterior branch. (I,J) Whole-mount antibody double staining of 1-eve-1 embryos at stage 11 (I) and stage 12 (J) with anti-β-galactosidase antibodies (red) and anti-Hb antibodies (green). The nuclear stainings with anti-Hb antibodies reveal a single nucleus at stage 11 (arrows in I) and two nuclei at stage 12 (arrows in J) posterior to the adjacent tracheal metameres. (K) Whole-mount in situ double hybridization of 1-eve-1 embryos at stage 11 with lacZ (red) and string (blue) antisense RNA probes. string (stg) expression (arrows), a marker for cell proliferation (Edgar and O'Farrell 1990), corresponds to the localization of the Hb-expressing nuclei (see I). (L) Whole-mount antibody double staining of stage 13 embryos bearing esg–lacZ with anti-β-galactosidase antibodies (red) and anti-Hb antibodies (green). The esg–lacZ enhancer trap line was used to mark the dorsal trunk fusion cell nuclei by β-galactosidase expression (Whiteley et al. 1992). Merged images of red and green pattern reveal no co-expression of β-galactosidase and Hb. (M) Scheme of tracheal branch formation (grey) and the localization of dorsal trunk fusion cell nuclei (red) and Hb-expressing nuclei (green) at stage 13. (N,O) Whole-mount antibody double staining of stage 12 embryos bearing UAS–GFPNlacZ and btl–Gal4 with anti-β-galactosidase antibodies (red) and anti-Hb antibodies (green). Merged images (N) of β-galactosidase expression in the tracheal cell nuclei (red) and nuclear Hb expression (green) as well as single-channel image for β-galactosidase expression (O; asterisks indicate Hb expression) reveals no co-expression of β-galactosidase and Hb. (P,Q) Antibody staining of a stage 11 wild-type (P) and a trh mutant (Q) embryo with anti-Hb antibodies. (Arrows) Bridge-cell precursors that correspond to tracheal metameres 1 and 4. (R,S) Whole-mount antibody double staining of stage 11 embryos bearing UAS–GFPNlacZ and twi–Gal4 with anti-β-galactosidase antibodies (red) and anti-Hb antibodies (green). (R) Merged images of nuclear β-galactosidase (red) and Hb expression (green) reveal co-expression of Hb and β-galactosidase (yellow) showing Hb expression in mesodermal cells. (S) Single channel image for twi-driven β-galactosidase expression in mesodermal cell nuclei (red); broken lines indicate tracheal placodes.

Tubular branch outgrowth is guided by the local and complex expression pattern of a Drosophila FGF homolog, Branchless (Bnl/dFGF), emanating from cell clusters surrounding each tracheal metamere (Sutherland et al. 1996; Metzger and Krasnow 1999). However, although mutant analysis shows that Bnl/dFGF is necessary for primary branch outgrowth, the restricted Bnl/dFGF expression seems not to be essential for the directed outgrowth of all primary branches. This conclusion is based on the observation that the constitutive activation of Bnl/dFGF signaling in bnl mutant embryos partially restores outgrowth of the main tracheal tube, the dorsal trunk, whereas the other primary branches are not generated. Thus, it was proposed that additional guidance cues might be necessary for the outgrowth of dorsal trunk branches (Sutherland et al. 1996).

Results and Discussion

We noted a single cell that is marked by expression of the gene hunchback (hb; Lehmann 1985; Tautz et al. 1987; Hülskamp 1991) at the posterior lateral margin of each tracheal metamere (Fig. 1D,I). This cell gives rise to daughter cells that maintain hb expression (Fig. 1E,J,K). The more ventrally located daughter cell maintains a round morphology and remains in position, whereas the dorsal daughter cell connects to the posterior bud of the tracheal metamere, termed the dorsal trunk posterior branch (Fig. 1E). Subsequently, the dorsal daughter cell elongates and extends posteriorly and thereby contacts to the anterior bud, termed the dorsal trunk anterior branch, of the adjacent posterior tracheal metamere (Fig. 1E,F). In this way, the dorsal daughter cell bridges the leading cells of the dorsal trunk anterior and posterior branches of two adjacent metameres (Fig. 1F), which then fuse about 2.5 hr later to form the continuous dorsal trunk. Thus, we refer to the dorsal daughter cell as the bridge-cell. The cell remains at this position until fusion between the dorsal trunk anterior and posterior branches occurs (Fig. 1G). During this fusion process, the bridge-cell becomes displaced and hb expression starts to fade (Fig. 1H).

To trace the origin of the bridge-cell, we performed double-staining experiments with tracheal-specific markers and hb. β-Galactosidase expression in nuclei of dorsal trunk fusion cells and in nuclei of tracheal cells revealed a lack of colocalization with bridge-cell hb expression (Fig. 1L–1O). Furthermore, trachealess (trh; Isaac and Andrew 1996; Wilk et al. 1996) mutant embryos, which lack tracheal cell identity, show hb-expressing bridge-cells as found in wild-type embryos (Fig. 1P,Q). Thus, these results indicate that the bridge-cell is of nontracheal origin. Finally, double-staining of hb and a mesodermal marker (Greig and Akam 1993) revealed coexpression of hb and the marker in bridge-cell precursors (Fig. 1R,S). Therefore, the bridge-cell is a nontracheal cell and of mesodermal origin.

To understand the function of bridge-cells in dorsal trunk formation, we first asked whether bridge-cell development is affected in hb mutant embryos. Homozygous hbFB mutant embryos, which express a nonfunctional Hb protein because of a premature stop codon mutation (Hülskamp 1991), express the hb transcript only transiently in bridge-cell precursors (not shown), raising the possibility that these cells may die. In fact, TUNEL staining suggests cell death is occurring at positions that correspond to those of bridge-cell precursors in hbFB mutants but not in wild-type embryos (Fig. 2A–D). This finding implies that the lack of hb activity causes bridge-cell precursors to undergo apoptosis. To show apoptosis as the underlying event of transient hb expression in bridge-cells more directly, we ubiquitously expressed in hbFB mutant embryos the baculovirus P35 protein, a suppresser of apoptosis in Drosophila (Hay et al. 1994). In contrast with hbFB mutants, which lack hb expression in the bridge-cells at stage 12 (Fig. 2E), hbFB embryos expressing P35 protein maintain hb expression in bridge-cells (Fig. 2F) as is found in wild-type embryos (Fig. 2G). Thus, expression of hb serves as a marker for bridge-cells, whereas its product, a transcription factor (Tautz et al. 1987; Hoch et al. 1991), is essential for bridge-cells viability. Therefore, analysis of tracheal development in hbFB mutant embryos would allow us to study bridge-cell function in dorsal trunk formation directly.

Figure 2.

Figure 2

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hb mutant embryos reveal apoptosis of the bridge-cell. (A–D) Whole-mount staining with anti-β-galactosidase antibody (brown) and TUNEL (blue) analysis of late stage 11 (A,C) and early stage 12 (B,D) hbFB mutant (A,B) and wild-type (C,D) embryos bearing the 1-eve-1 chromosome. Apoptotic nuclei are detectable in hbFB mutant embryos at posterior lateral positions of the tracheal placodes (arrows in A and B) but not in wild-type embryos (C,D). (E) Whole-mount in situ double hybridization of a stage 12 hbFB mutant embryo bearing the 1-eve-1 chromosome with lacZ (red) and hb (blue) antisense RNA probes. No hb expression is detectable at the expected positions of the bridge-cells (arrows in E). (F) Whole-mount in situ hybridization of a stage 12 hbFB mutant embryo bearing actin–Gal4 and UAS–P35 with a hb antisense RNA probe. hb expression is detectable in corresponding cells (arrows in F) after suppression of apoptosis by ubiquitous expression of P35 protein (Hay et al. 1994). (G) Whole-mount in situ double hybridization of a stage 12 wild-type embryo bearing the 1-eve-1 chromosome with lacZ (red) and hb (blue) antisense RNA probes. hb expression is detectable in the bridge-cells (arrows in G).

In hbFB mutant embryos initial tracheal development, including primary branch outgrowth, appears normal up to the end of stage 12 (Fig. 3A–D). Subsequently, the dorsal trunk branches become stalled and misrouted, whereas the other primary branches are formed as in wild-type embryos (Fig. 3E–H). Despite the strong dorsal trunk phenotype, the dorsal trunk branches occasionally fuse in hbFB mutant embryos and form dorsal trunk rudiments (Fig. 3F,H). This observation and normal expression of the escargot (esg) fusion cell marker (Whiteley et al. 1992) in hbFB mutant embryos (Fig. 3I) suggest that the fusion process, required for dorsal trunk formation, is not impaired in hbFB mutant embryos. These results indicate that hb is not necessary for the initial outgrowth but for the subsequent outgrowth of dorsal trunk branches. Thus, the results also suggest that the hb-dependent bridge-cells are involved in the outgrowth of dorsal trunk branches toward their fusion partners.

Figure 3.

Figure 3

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hb expression in the bridge-cell is essential for directed outgrowth of dorsal trunk branches and does not interfere with dFGF signaling. Whole-mount in situ hybridization with a lacZ antisense RNA probe of wild-type (A,E) and hbFB mutant (B,F) embryos bearing the 1-eve-1 chromosome at stage 12 (A,B) and stage 15 (E,F). (C,D) Whole-mount antibody staining of a stage 12 wild-type (C) and a hbFB mutant (D) embryo with anti-Crumbs antibodies. (G,H) Whole-mount antibody staining of a stage 15 wild-type (G) and a hbFB mutant (H) embryo with antibody 2A12. This antibody specifically stains the tracheal lumen and reveals a normal lateral trunk but a lack of dorsal trunk formation in the hbFB mutant (H). Note that the hb9Q mutant embryos reveal an identical tracheal phenotype to that found in hbFB mutant embryos. (I) Whole-mount antibody double staining of a stage 15 hbFB mutant embryo bearing the G6 chromosome with 2A12 (brown) and anti-β-galactosidase (blue) antibodies reveals esg-driven β-galactosidase expression in the dorsal trunk fusion cells of hbFB mutant embryos. (J,K) Whole-mount in situ hybridization of a stage 12 wild-type (J) and a hbFB mutant (K) embryo using bnl antisense RNA. The dynamic bnl expression pattern surrounding a single tracheal metamere (broken lines indicate bnl expression that guides dorsal trunk outgrowth) in either a wild-type (J) or hbFB mutant (K) embryo is identical. (L) Whole-mount antibody double staining of a stage 12 btlH82Δ3 mutant embryo with anti-β-galactosidase (brown) and anti-Hb (blue) antibodies. The btlH82Δ3 mutant embryo reveals β-galactosidase expression in the tracheal nuclei (Reichman-Fried et al. 1994) and Hb expression in the bridge-cells (arrows).

Recent studies have shown that Bnl/dFGF is necessary for the primary tracheal branching, including the formation of the dorsal trunk (Sutherland et al. 1996; Metzger and Krasnow 1999). Therefore, we asked whether the absence of bridge-cells might interfere with bnl expression. We found that the expression pattern of bnl was unaffected in hb mutant embryos (Fig. 3J,K). Also, hb expression in the bridge-cells was not affected in bnl mutant embryos and in embryos that lack the activity of breathless (btl), which codes for the Bnl/dFGF receptor (Fig. 3L; data not shown). Thus, bridge-cells do not interfere with the proper expression of Bnl/dFGF around the developing tracheal branches, and hb-expression in the bridge-cells is independent of Bnl/dFGF signaling.

Because localized Bnl/dFGF signaling is not necessary for dorsal trunk formation (Reichman-Fried et al. 1994; Lee et al. 1996; Sutherland et al. 1996), we asked whether the bridge-cell mediates the proposed additional guidance mechanism for dorsal trunk branch outgrowth (Sutherland et al. 1996). By use of the Gal4/UAS-system (Brand and Perrimon 1993), we expressed Bnl/dFGF ectopically in tracheal cells to impede the spatial cues that are normally derived from the local arrangement of cell clusters expressing Bnl/dFGF. In contrast with wild-type embryos (Fig. 4A), embryos with ectopic expression of Bnl/dFGF develop complete dorsal trunk structures but lack the other primary branches (Fig. 4B). However, hbFB mutant embryos that express Bnl/dFGF ectopically had no signs of dorsal trunk branch outgrowth at all (Fig. 4C). These results indicate that the bridge-cell is necessary and essential for dorsal trunk formation, suggesting that this cell provides guidance cues specifically during the anterior–posterior dorsal trunk branch outgrowth. Thus, the bridge-cell, in combination with Bnl/dFGF signaling, directs outgrowth of the main tracheal tube and may mediate the proposed additional guidance mechanism.

Figure 4.

Figure 4

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The bridge-cell is necessary for dorsal trunk branch outgrowth. (A–C) Whole-mount antibody 2A12 staining of a stage 15 wild-type embryo (A), an embryo bearing UAS–bnl and btl–Gal4 (B), and a hbFB mutant embryo bearing UAS–bnl and btl–Gal4 (C). Note that the tracheal phenotype of amorphic bnlP1 mutant embryos bearing UAS–bnl and btl–Gal4 is indistinguishable from the phenotype shown in B. The lack of tracheal metameres in hbFB mutant embryos is caused by the lack of segmental anlagen in such embryos (Lehmann 1985; Tautz et al. 1987). (D) Whole-mount in situ double-hybridization of a stage 12 embryo bearing UAS–hb, PO163–Gal4 and the 1-eve-1 chromosome with lacZ (red) and hb (blue) antisense RNA probes. PO163–Gal4 (Janning 1997) drives hb expression in SOP cells. (Arrow) Outgrowing dorsal trunk anterior branch that attaches to a hb expressing cell but lacks contact with the bridge-cell. (E) Whole-mount antibody staining of a stage 15 embryo bearing UAS–hb and PO163–Gal4 with antibody 2A12. Ectopic hb expression causes bottleneck-like lumen formation (arrowheads) and a lack of dorsal trunk interconnection (arrow). (F,G) Whole-mount in situ double hybridization of a stage 12 embryo bearing the 1-eve-1 chromosome with lacZ (red) and hb (blue) antisense RNA probes. The same image is shown focused on the bridge-cell (F) and the tracheal cell extensions (G), respectively.

To test the above inference, we expressed hb ectopically via the Gal4/UAS-system (Brand and Perrimon 1993) in sensory organ precursor (SOP) cells in positions close to the bridge-cells. The outgrowing dorsal trunk anterior branches were seen in contact with the cells that ectopically express hb, even in the presence of the normal bridge-cells (Fig. 4D). As a consequence of the ectopic hb expression, the dorsal trunk of the embryos show interruptions and abnormal bottleneck-like fusion points (Fig. 4E). Thus, hb expression in ectopic cells close to bridge-cells triggers a differentiation program that interferes with the directed outgrowth of the dorsal trunk branches suggesting that hb activity is required not only for the viability but also for the identity of the bridge-cell. Whether the differentiation program involves local and short-range signals and/or provides a migration matrix by cell adhesion is unknown. However, we prefer the hypothesis that the bridge-cell serves as an adhesion-dependent guiding post, as we observed tracheal cell extensions along the bridge-cell directly after the initial contact (Fig. 4F,G).

Our discovery of the bridge-cell and previous studies on Bnl/dFGF signaling provide a coherent model of how dorsal trunk formation may occur. After invagination of the tracheal placodes, budding of the tracheal metameres is triggered by localized Bnl/dFGF activity (Sutherland et al. 1996). This signal apparently does not always have the necessary precision on its own to guide the leading cells. The bridge-cell provides this precision by serving as a guidance post to properly position the budding dorsal trunk branches. The results also demonstrate an interplay of cells deriving from two different germ layers, mesoderm and ectoderm, which is necessary to establish the interconnected tubular tracheal network during embryogenesis. The identification of a key player in bridge-cell differentiation, namely the transcription factor Hb, provides an entry point to unravel the molecular targets of hb. Their analysis may also contribute to gaining further insights into the function of the bridge-cells during tubular network formation, possibly in organisms other than Drosophila.

Materials and methods

Materials

We used the following antibodies: monoclonal antibody 2A12 to stain tracheal lumen (DSHB, Iowa); anti-β-galactosidase antibody (Promega); anti-Hunchback antibody (gift from A. La Rosée, MPI, Göttingen); anti-Crumbs antibody (Tepass and Knust 1993); Alexa 488 or Alexa 546 goat antirabbit IgG or goat antimouse IgG (Molecular Probes); alkaline phosphatase-conjugated or biotinylated antirabbit IgG or antimouse IgG; biotinylated antimouse IgM (Vector Laboratories); anti-digoxigenin- and anti-fluorescein-AP, Fab fragments (Roche).

We used a number of alleles and fly strains: UAS-hb flies (Wimmer et al. 2000); hbFB, hb9Q, dfrE82, trh5D55, and btlH82Δ3 were obtained from the Tübingen Stock Center. btl-Gal4 drives Gal4 expression ubiquitously in the tracheal system from stage 10 onward (Shiga et al. 1996). UAS-GFPNlacZ was used to detect nuclear β-galactosidase expression (Shiga et al. 1996). The lacZ enhancer trap line 1-eve-1 reveals P-element integration in the trh gene and was used to mark tracheal cells by cytoplasmic β-galactosidase (Perrimon et al. 1991). PO163 drives Gal4 in a subset of peripheral nervous system precursor cells (Janning 1997). UAS-P35 was provided by H. Steller (MIT, Cambridge). The P-element of the lacZ enhancer trap line G6 is integrated in the esg gene and marks dorsal trunk homotip cell nuclei (Whiteley et al. 1992). We also used actin–Gal4 and twi–Gal4 flies (Greig and Akam 1993).

TUNEL assay

TUNEL analysis of embryos was done by RNA in situ hybridization with the following modifications: After the second fixation step, the embryos were washed with TUNEL reaction mixture (In Situ Cell Death Detection Kit, AP; Roche) and incubated with 100 μl of TUNEL reaction mixture for 120 min at 37°C; the embryos were washed with PBT and incubated with anti-fluorescein antibody (Roche) for 12 hr at 4°C.

Immunostainings and in situ hybridizations

RNA in situ hybridizations and immunostainings to whole-mount embryos were performed as described (Goldstein and Fryberg 1994). The RNA probes used in our experiments were derived from bnl (Sutherland et al. 1996), stg (Edgar and O'Farrell 1990), lacZ, and sal (Kühnlein et al. 1994). Immunostained embryos were viewed with a Zeiss Axiophot microscope. Embryos stained with fluorescent antibodies were analyzed by laser scanning microscopy as described (Kühnlein and Schuh 1996).

Acknowledgments

We are grateful to M. González-Gaitán for initiation of the project. Special thanks go to H. Jäckle for providing a stimulating environment and critical reading of the manuscript. We thank M.A. Krasnow, A. La Rosée, C. Samakovlis, H. Steller, and E. Wimmer for antibodies, cDNAs and fly stocks. We also thank M. Affolter, R.P. Kühnlein, C. Krause and E. Wimmer for critical comments on the manuscript. This work was supported by the Max-Planck-Society (MPIbpc Abt. 170) and the Deutsche Forschungsgemeinschaft (SFB 271).

The publication costs of this article were defrayed in part by payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 USC section 1734 solely to indicate this fact.

Footnotes

E-MAIL rschuh@gwdg.de; FAX 49-551-201-1755.

Article and publication are at www.genesdev.org/cgi/doi/10.1101/gad.180900.

References

  1. Brand AH, Perrimon N. Targeted gene expression as a means of altering cell fates and generating dominant phenotypes. Development. 1993;118:401–415. doi: 10.1242/dev.118.2.401. [DOI] [PubMed] [Google Scholar]
  2. Edgar BA, O'Farrell PH. The three postblastodermal cell cycles of Drosophila embryogenesis are regulated in G2 by String. Cell. 1990;62:469–480. doi: 10.1016/0092-8674(90)90012-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Goldstein LSB, Fryberg EA. Drosophila melanogaster: Practical uses in cell and molecular biology. Methods Cell Biol. 1994;44:446–485. [Google Scholar]
  4. Greig S, Akam M. Homeotic genes autonomously specify one aspect of pattern in the Drosophila mesoderm. Nature. 1993;362:630–632. doi: 10.1038/362630a0. [DOI] [PubMed] [Google Scholar]
  5. Hay BA, Wolff T, Rubin GM. Expression of baculovirus P35 prevents cell death in Drosophila. Development. 1994;120:2121–2129. doi: 10.1242/dev.120.8.2121. [DOI] [PubMed] [Google Scholar]
  6. Hoch M, Seifert E, Jäckle H. Gene expression mediated by cis-acting sequences of the Krüppel gene in response to the Drosophila morphogens bicoid and hunchback. EMBO J. 1991;10:2267–2278. doi: 10.1002/j.1460-2075.1991.tb07763.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hogan BLM, Grindley J, Bellusci S, Dunn NR, Emoto H, Itoh N. Branching morphogenesis of the lung: New models for a classical problem. Cold Spring Harbor Symp Quant Biol. 1997;62:249–256. [PubMed] [Google Scholar]
  8. Hülskamp M. Funktionelle Analyse des Segmentierungsgens hunchback von Drosophila melanogaster. Thesis. Germany: University of Tübingen; 1991. [Google Scholar]
  9. Isaac DD, Andrew DJ. Tubulogenesis in Drosophila: A requirement for the trachealess gene product. Genes & Dev. 1996;10:103–117. doi: 10.1101/gad.10.1.103. [DOI] [PubMed] [Google Scholar]
  10. Janning W. Fly View, a Drosophila image database, and other Drosophila databases. Cell Dev Biol. 1997;8:469–475. doi: 10.1006/scdb.1997.0172. [DOI] [PubMed] [Google Scholar]
  11. Kühnlein RP, Schuh R. Dual function of the region specific homeotic gene spalt during Drosophila tracheal system development. Development. 1996;122:2215–2223. doi: 10.1242/dev.122.7.2215. [DOI] [PubMed] [Google Scholar]
  12. Kühnlein RP, Frommer G, Friedrich M, Gonzalez-Gaitan M, Weber A, Wagner-Bernholz JF, Gehring WJ, Jäckle H, Schuh R. spalt encodes an evolutionarily conserved zinc finger protein of novel structure which provides homeotic gene function in the head and tail region of the Drosophila embryo. EMBO J. 1994;13:168–179. doi: 10.1002/j.1460-2075.1994.tb06246.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Lee T, Hacohen N, Krasnow M, Montell DJ. Regulated Breathless receptor tyrosine kinase activity required to pattern cell migration and branching in the Drosophila tracheal system. Genes & Dev. 1996;10:2912–2921. doi: 10.1101/gad.10.22.2912. [DOI] [PubMed] [Google Scholar]
  14. Lehmann R. Regionsspezifische Segmentier-ungsmutanten bei Drosophila melanogaster Meigen. Thesis. Germany: University of Tübingen; 1985. [Google Scholar]
  15. Manning G, Krasnow MA. Development of the Drosophila tracheal system. In: Bate M, Martinez Arias A, editors. The development of Drosophila melanogaster. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press; 1993. pp. 609–685. [Google Scholar]
  16. Metzger RJ, Krasnow MA. Genetic control of branching morphogenesis. Science. 1999;284:1635–1639. doi: 10.1126/science.284.5420.1635. [DOI] [PubMed] [Google Scholar]
  17. Perrimon N, Noll E, McCall K, Brand A. Generating lineage-specific markers to study Drosophila development. Dev Genet. 1991;12:238–252. doi: 10.1002/dvg.1020120309. [DOI] [PubMed] [Google Scholar]
  18. Reichman-Fried M, Dickson B, Hafen E, Shilo B-Z. Elucidation of the role of breathless, a Drosophila FGF receptor homolog, in tracheal cell migration. Genes & Dev. 1994;8:428–439. doi: 10.1101/gad.8.4.428. [DOI] [PubMed] [Google Scholar]
  19. Risau W. Mechanism of angiogenesis. Nature. 1997;386:671–674. doi: 10.1038/386671a0. [DOI] [PubMed] [Google Scholar]
  20. Samakovlis C, Hacohen N, Manning G, Sutherland DC, Guillemin K, Krasnow MA. Development of the Drosophila tracheal system occurs by a series of morphologically distinct but genetically coupled branching events. Development. 1996;122:1395–1407. doi: 10.1242/dev.122.5.1395. [DOI] [PubMed] [Google Scholar]
  21. Shiga Y, Tanaka-Matakatsu M, Hayashi S. A nuclear GFP/β-galactosidase fusion protein as a marker for morphogenesis in living Drosophila. Dev Growth Differ. 1996;38:99–106. [Google Scholar]
  22. Sutherland D, Samakovlis C, Krasnow MA. branchless encodes a Drosophila FGF homolog that controls tracheal cell migration and the pattern of branching. Cell. 1996;87:1091–1101. doi: 10.1016/s0092-8674(00)81803-6. [DOI] [PubMed] [Google Scholar]
  23. Tautz D, Lehmann R, Schnürch H, Schuh R, Seifert E, Kienlin A, Jones K, Jäckle H. Finger protein of novel structure encoded by hunchback, a second member of the gap class of Drosophila segmentation genes. Nature. 1987;327:383–389. [Google Scholar]
  24. Tepass U, Knust E. Crumbs and stardust act in a genetic pathway that controls the organization of epithelia in Drosophila melanogaster. Dev Biol. 1993;159:311–326. doi: 10.1006/dbio.1993.1243. [DOI] [PubMed] [Google Scholar]
  25. Whiteley M, Noguchi PD, Sensabaugh SM, Odenwald WF, Kassis JA. The Drosophila gene escargot encodes a zinc finger motif found in snail-related genes. Mech Dev. 1992;36:117–127. doi: 10.1016/0925-4773(92)90063-p. [DOI] [PubMed] [Google Scholar]
  26. Wilk R, Weizman I, Shilo B-Z. trachealess encodes a bHLH-PAS protein that is an inducer of tracheal cell fates in Drosophila. Genes & Dev. 1996;10:93–102. doi: 10.1101/gad.10.1.93. [DOI] [PubMed] [Google Scholar]
  27. Wimmer EA, Carleton A, Harjes P, Turner T, Desplan C. bicoid-independent formation of thoracic segments in Drosophila. Science. 2000;278:2476–2479. doi: 10.1126/science.287.5462.2476. [DOI] [PubMed] [Google Scholar]

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