DELTA

Spatial and Temporal Dynamics of the Notch Ligand Delta During Development in Drosophila Melagonaster

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 Obiettivo del progetto (Objective)

'The Notch (N) pathway is a highly conserved and ubiquitous signalling system involved in determining cell fate. There is a medical relevance to understand the molecular mechanisms involved in N signalling as N dysfunction is linked to cancer, stroke and Alzheimer’s disease. N is activated by its ligands (Delta/Serrate/LAG2;DSL) expressed on adjacent cells and there is a clear correlation between ubiquitination and endocytosis for effective ligand signalling. Although several models for DSL ligand endocytosis have been identified, the behaviour of DSL ligands during these events remains unclear. Attempts to address this issue have lacked the ability to follow spatial and temporal movement of DSL ligands accurately. The current project will therefore address these issues by using live fluorescent imaging techniques to follow movement of Delta (Dl) ligand, fused to a fluorescent reporter, in the wing imaginal disc of Drosophila Melagonaster. This will be achieved by examining Dl molecules present in large intracellular vesicles throughout the wing disc where the ubiquitin ligases Neuralized and Mindbomb1 are expressed. To establish whether Dl activity is dynamic in these particles, we will utilise the FRAP imaging technique. We will follow the movement of Dl molecules from these particles, using a photoswitchable reporter to address the model of “recycling”, where DSL ligands are processed into an active state within these vesicles before being trafficked back to the cell surface to activate N. The effects of impairing endocytosis upon Dl movement from these particles will also be addressed. The project will also address the importance of endocytosis/recycling for effective Dl/N signalling via utilisation of a fluorescent N reporter. These studies will advance our understanding about the dynamic nature of Dl molecules and further assist in elucidating the mechanisms involved in endocytic regulation of N signalling during development.'

Introduzione (Teaser)

European researchers worked to elucidate the mechanisms involved in the regulation of Notch signalling during development. By focusing on the internalisation of the Notch ligand Delta, they provided important information on the dynamics of signalling and its potential link with disease.

Descrizione progetto (Article)

During development, one of the central signal transduction pathways implicated in the determination of cell fate is the Notch pathway. Signalling through the Notch receptors has been highly conserved through evolution. It mediates cell communication and essentially drives cell differentiation early in life.

Given its involvement in many diseases including cancer, stroke and Alzheimer's disease, understanding how Notch signalling works is of high medical relevance. Scientists of the EU-funded DELTA project used the fruit fly Drosophila melanogaster to study the mechanism of action of central players in this pathway, like Delta.

The Notch ligand Delta is also a transmembrane protein which requires specific modifications to accommodate signal transduction.

Experimental evidence surprisingly suggests that Delta proteins need to be cleared from the membrane to exert a mechanical force on the Notch receptor in the adjacent cell.

To investigate the intracellular fate of the Delta proteins after internalisation, researchers fused it with a fluorescent red protein. Through time-lapse confocal microscopy, they were able to detect Delta within different cellular compartments at different stages of larva development.

Genetic studies revealed the modification enzymes essential for Delta endocytosis and recycling, and the stage during internal trafficking where Delta exerts its function on the Notch receptor.

Taken together, the findings of the DELTA study provide important insight on the Notch signalling pathway. Besides advancing our understanding on the dynamic nature of Delta molecules, this information could form the basis to study the implication of Delta in Notch-related pathologies.