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Mutual regulation of the Drosophila disconnected (disco) and Distal-less (Dll) genes contributes to proximal-distal patterning of antenna and leg

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Abstract

The Drosophila disconnected (disco) gene encodes a C2H2-type zinc finger transcription factor required for the development of the central and peripheral nervous systems. We report that disco participates in a positive feedback loop with the Dll gene, a master regulator of ventral appendage development. Dll function is not only required for proper disco expression in antenna and leg discs, but is also sufficient for ectopic expression of disco in the developing retina and wing imaginal discs. Conversely, disco gene function is required for the maintenance of Dll expression. We show that Dll phenotypes are partially rescued by the up-regulation of disco expression in the Dll domain. Reduction in disco gene function disrupts antenna and leg development, and the phenotypes closely resemble that produced by Dll alleles. These observations demonstrate that disco plays a fundamental role in the Dll-dependent patterning of antenna and leg, perhaps as a regulator of Dll gene expression.

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Acknowledgements

We thank Drs. Gerard Campbell, Grace Boekhoff-Falk, Ian Duncan, Isabel Guerrero, Frank Laski, University of Iowa Developmental Studies Hybridoma Bank, and Bloomington Drosophila Stock Center for kindly providing the antibodies and fly lines used in this study. We are also grateful to Dr. Richard Carthew for the pWIZ RNAi vector and to Drs. Andre Bedard, Roger Jacobs and Anindya Dutta for critical reading of the manuscript.

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Correspondence to Ana Regina Campos.

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This work is supported by a grant from the Natural Sciences and Engineering Research Council of Canada (NSERC) to A.R. Campos.

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Suppl. Fig. 1

a Endogenous disco mRNA is gradually down-regulated when UAS-disco RNAi is driven by HSP70-GAL4. Embryos of genotype HSP70-GAL4; UAS-disco RNAi, aged 0–14 h, were heat-shocked for 1 h at 37°C and allowed to recover at 25°C for 1, 2, or 3 h. Whole-mount in situ hybridization was carried out (see: Tautz D, Pfeifle C [1989] A non-radioactive in situ hybridization method for the localization of specific RNAs in Drosophila embryos reveals translational control of the segmentation gene hunchback.Chromosoma 98:81–85) with hybridization probes generated from a discocDNA cloned into Bluescript vector by using the DIG RNA Labeling Kit of Roche AppliedSciences (Top a–c stage-matched non-heat-shocked controls, Bottom d–f heat-shocked embryos). b Endogenous disco level of heat-shocked HSP70-GAL4; UAS-disco RNAi embryos was measured by real-time PCR. Embryos carrying HSP70-GAL4; UAS-disco RNAi elements and stage-matched OR control embryos, aged 0–14 h, were heat-shocked for 1 h at 37°C and allowed to recover at 25°C for 1, 2, or 3 h. RNA was isolated and real-time PCR was carried out with DNA Master SYBR green-1. An approximately two-, four-, and nine-fold reduction of disco RNA level was observed 1, 2, and 3 h, respectively, after heat-shock-induced expression of UAS-disco RNAi transgenes compared with that of wild type (OR) embryos. (DOC 1000 kb)

Suppl. Fig. 2

Expression of UAS-disco RNAi suppresses the phenotype of GMR-GAL4; UAS-disco. Environmental scanning electron micrographs of Drosophila compound eyes (a–d are shown at higher magnification in e–h). a, e Wild type. b, f GMR-GAL4; UAS-disco. Ectopic expression of UAS-disco posterior to the morphogenetic furrow by using the GMR-GAL4 driver causes small rough eyes lacking eye pigments. The lenses of the cone cells appear fused, the interommatidial sensory bristles are displaced and/or missing, and many tiny bristles of unknown origin are present. c, g GMR-GAL4; UAS-disco/UAS-disco RNAi. This phenotype is completely rescued by co-expressing UAS-disco RNAi. d, h GMR-GAL4; UAS-disco RNAi shows a normal eye. (DOC 3330 kb)

Suppl. Fig. 3

Function of disco is not required for proper expression of DAC in leg discs. a Wild type. b Dll-GAL4/CyO. c disco 1/disco 1. d Dll-GAL4; UAS-disco RNAi. (DOC 433 kb)

Suppl. Table 1

Dll alleles increase the lethality of disco 1. Dll 5, Dll9, and Dll 3 in combination with disco 1 hemizygous flies exhibit 100%, 75%, and 52.38% lethality, respectively, compared with disco 1 hemizygous alone, whereas the hth 0475 allele has no impact on lethality. (DOC 33 kb)

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Dey, B.K., Zhao, XL., Popo-Ola, E. et al. Mutual regulation of the Drosophila disconnected (disco) and Distal-less (Dll) genes contributes to proximal-distal patterning of antenna and leg. Cell Tissue Res 338, 227–240 (2009). https://doi.org/10.1007/s00441-009-0865-z

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