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Open Access Research article

Notch signaling during development requires the function of awd, the Drosophila homolog of human metastasis suppressor gene Nm23

Marilena Ignesti1, Marilena Barraco15, Gouthami Nallamothu2, Julie A Woolworth3, Serena Duchi16, Giuseppe Gargiulo1, Valeria Cavaliere1* and Tien Hsu24*

Author Affiliations

1 Dipartimento di Farmacia e Biotecnologie, Alma Mater Studiorum Università di Bologna, Via Selmi, 3, Bologna 40126, Italy

2 Department of Medicine, Boston University School of Medicine, Boston, Massachusetts 02118, USA

3 Department of Pathology and Laboratory Medicine, Medical University of South Carolina, Charleston, South Carolina 29425, USA

4 Graduate Institute of Systems Biology and Bioinformatics, National Central University, Jhongli, Taiwan

5 Present address: Institute of Hematology “L. e A. Seràgnoli”, University of Bologna, Bologna, Italy

6 Present address: Bone Regeneration Laboratory, Research Institute Codivilla-Putti, Rizzoli Orthopaedic Institute, Bologna, Italy

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BMC Biology 2014, 12:12  doi:10.1186/1741-7007-12-12

Published: 14 February 2014

Abstract

Background

The Drosophila abnormal wing discs (awd) belongs to a highly conserved family of genes implicated in metastasis suppression, metabolic homeostasis and epithelial morphogenesis. The cellular function of the mammalian members of this family, the Nm23 proteins, has not yet been clearly defined. Previous awd genetic analyses unraveled its endocytic role that is required for proper internalization of receptors controlling different signaling pathways. In this study, we analyzed the role of Awd in controlling Notch signaling during development.

Results

To study the awd gene function we used genetic mosaic approaches to obtain cells homozygous for a loss of function allele. In awd mutant follicle cells and wing disc cells, Notch accumulates in enlarged early endosomes, resulting in defective Notch signaling. Our results demonstrate that awd function is required before γ-secretase mediated cleavage since over-expression of the constitutively active form of the Notch receptor in awd mutant follicle cells allows rescue of the signaling. By using markers of different endosomal compartments we show that Notch receptor accumulates in early endosomes in awd mutant follicle cells. A trafficking assay in living wing discs also shows that Notch accumulates in early endosomes. Importantly, constitutively active Rab5 cannot rescue the awd phenotype, suggesting that awd is required for Rab5 function in early endosome maturation.

Conclusions

In this report we demonstrate that awd is essential for Notch signaling via its endocytic role. In addition, we identify the endocytic step at which Awd function is required for Notch signaling and we obtain evidence indicating that Awd is necessary for Rab5 function. These findings provide new insights into the developmental and pathophysiological function of this important gene family.

Keywords:
Awd; Notch signaling; Endocytosis