INTRODUCTION Embryonic segmentation and formation of the adult body are under the control of homeotic genes and signalling cascades. In Drosophila, the zinc-finger protein Teashirt (Tsh) is expressed from early embryogenesis to adulthood in specific domains where it acts both with Hox proteins and the Wingless signalling pathway for patterning (de Zulueta et al., 1994; Gallet et al., 1999; Gallet et al., 1998; Röder, 1992). Tsh is involved in the specification of the embryonic trunk (Fasano et al., 1991), parts of the intestine (Mathies et al., 1994) and the proximal part of the adult appendages (Erkner et al., 1999; Wu and Cohen, 1999). Expression pattern analysis of two putative murine Tsh orthologues suggest that Tsh function may have been conserved for patterning (Caubit et al., 2000). During the first stages of embryonic development, the tsh expression pattern in the ectoderm is very dynamic and at gastrulation tsh mRNAs are homogeneously distributed in the presumptive trunk region. Genetic evidence suggest that tsh is activated and restricted in the trunk of early embryos by a combination of maternal and segmentation genes (Röder, 1992). Maternal and segmentation genes act either as repressors or activators of tsh transcription, in order to delimit the boundaries of tsh expression domains. The pair-rule gene fushi tarazu (ftz) activates tsh expression directly in even- numbered parasegments in the embryonic ectoderm (Coré et al., 1997). Later during embryogenesis, Tsh expression is maintained by homeotic genes (Röder, 1992) and autoregulation (Coré et al., 1997). Gallet et al. (Gallet et al., 1998) have shown that Wg signalling is necessary to accumulate a high amount of Tsh protein in the nucleus in order to give a trunk specific output for Wg signalling. During Drosophila embryogenesis, a group of epithelial cells from each thoracic hemisegment will invaginate to form the primordia of the adult legs. These epithelia proliferate during larval life to give rise to the imaginal discs, which undergo morphogenesis and differentiation during metamorphosis (Bryant, 1978; Cohen, 1993). In the leg discs, hedgehog (hh) is transcribed in the posterior compartment and its protein is secreted to the anterior part to induce wingless (wg) and decapentaplegic (dpp) transcription in ventral and dorsal domains, respectively. Wg and Dpp proteins, which are homologous to Wnt1 and TGFβ in vertebrates, specify the ventral and dorsal cell fates, respectively, and via mutual repression establish the dorsoventral and the proximodistal axes of the leg. These signalling proteins impose progressively restricted patterning decisions on neighbouring cell groups, via independent transduction pathways, to give largely invariant appendages whether in Drosophila or vertebrates (Basler and Struhl, 1994; Brook and Cohen, 1996; Diaz-Benjumea and Cohen, 1994; Ingham and Fietz, 1995; Jiang and Struhl, 1996; Klingensmith et al., 1994; Lecuit and Cohen, 1997; Massague, 1998; Penton and Hoffmann, 1996; Wodarz and Nusse, 1998; Wolpert, 1969; Yang and Niswander, 1995). Several genes have been isolated that exhibit differential, proximodistal patterns of expression in the imaginal discs 1119 Development 129, 1119-1129 (2002) Printed in Great Britain © The Company of Biologists Limited 2002 DEV2830 We have carried out a genetic screen designed to isolate regulators of teashirt expression. One of these regulators is the Grunge gene, which encodes a protein with motifs found in human arginine-glutamic acid dipeptide repeat, Metastasis-associated-like and Atrophin-1 proteins. Grunge is the only Atrophin-like protein in Drosophila, whereas several exist in humans. We provide evidence that Grunge is required for the proper regulation of teashirt but also has multiple activities in fly development. First, Grunge is crucial for correct segmentation during embryogenesis via a failure in the repression of at least four segmentation genes known to regulate teashirt. Second, Grunge acts positively to regulate teashirt expression in proximoventral parts of the leg. Grunge has other regulatory functions in the leg, including the patterning of ventral parts along the entire proximodistal axis and the proper spacing of bristles in all regions. Key words: Drosophila, Grunge, Teashirt, Legs, Atrophin-1-like proteins, Metastasis-associated proteins, Segmentation SUMMARY Grunge, related to human Atrophin-like proteins, has multiple functions in Drosophila development Alfrun Erkner*, Agnès Roure, Bernard Charroux, Michèle Delaage, Nicolas Holway, Nathalie Coré, Christine Vola, Corinne Angelats, Françoise Pagès, Laurent Fasano and Stephen Kerridge † Laboratoire de Génétique et Physiologie du Développement, UMR 9943 C.N.R.S.-Université, I.B.D.M. CNRS-INSERM-Université de la Méditerranée, Campus de Luminy Case 907, F-13288 Marseille, Cedex 09, France *Present address: Institut für Molekularbiologie, Universität Zürich, Winterthurerstrasse 190, 8057 Zürich, Switzerland † Author for correspondence (e-mail: kerridge@ibdm.univ-mrs.fr) Accepted 14 December 2001