WORKPACKAGE 3

Study of the ciliary protein interaction network to establish a hierarchy of temporal and spatial interactions of BBS, OFD1, and NPHP proteins

Alberto Luini from the Consorzio Mario Negri Sud (Abruzzo, Italy) is the WPL.

This package aims in general to establish the interaction network within which the BBS/OFD1/NPHP proteins operate and to use this information to construct models of molecular pathways that specify the temporal and spatial involvement of these proteins in cilia structure and function.

1. To define the physical interactions of these proteins we will a) perform yeast twohybrid screens using these proteins as baits and b) use the cellular model(s) developed in WP2, or renal tissue, to isolate and identify by immunoprecipitation/mass spectrometry the proteins complexed with the BBS/OFD1/NPHP proteins.
2. To define the functional interactions of these proteins we will a) use yeast as a platform to screen for functional interactions of the BBS/OFD1/NPHP and b) enhance or reduce the levels of the BBS/OFD1/NPHP proteins, and then analyse their gene expression profile in these cells in comparison with control cells to reconstruct the involved molecular pathways.
3. We will mine existing interactome databases for predicted interactors of the BBS/OFD1/NPHP proteins.