Seminar

A new tagging strategy to identify ubiquitin conjugates from Drosophila neurons

Dr. Ugo Mayor

Ubiquitination, a covalent modification mediating protein degradation and transport, appears to be implicated in many neuronal functions, including plasticity. I have developed a method to isolate and identify, in a tissue specific manner, the neuronal targets of the ubiquitination machinery in Drosophila, an accessible model organism that still contains the myriad of components required to build a complex nervous system. This new technique involves the expression of tagged ubiquitin, and its biotinylation in vivo, which occurs very efficiently. We are isolating ubiquitin conjugates from embryonic neuronal extracts as well as from adult fly brains, and the proteins are being identified by standard mass spectroscopy techniques.

In addition to elucidating which proteins are ubiquitinated in a wild type background, I am currently setting up the experiments to compare these extracts with those of E3 ubiquitin ligase mutants. Identifying the proteins ubiquitinated by specific E3 ligases should allow us to dissect the molecular pathways of interaction, which could then be further investigated using standard genetic techniques, readily available when working with flies.