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Institute for Research in Biomedicine
Istituto di Ricerca in Biomedicina

Via Vincenzo Vela 6 - CH-6500 Bellinzona
Tel. +41 91 820 0300 - Fax +41 91 820 0302 - info [at] irb [dot] usi [dot] ch

Comparative Interactomics to Identify Novel ER-Resident Quality Control Players

Research area: Protein Folding and Quality Control

Group leaders: Maurizio Molinari


Status: In progress

The aim of the project is to identify new players that intervene in protein folding, quality control and ERAD in the mammalian ER lumen. We generated a collection of human cell lines expressing epitope-tagged folding-competent and folding-defective proteins. The model proteins are used as baits to capture interacting partners in the same immuno-complexes. The proteins co-immunoisolated with the individual baits are subjected to tryptic digestion and fragments are separated by nano-HPLC followed by tandem mass spectrometry. Fragmentation spectra of the samples are matched to a human protein database sequence with the Mascot software. These analytic steps are performed in collaboration with Manfredo Quadroni, coordinator of the Center for Integrative Genomics, University of Lausanne. Involvement in protein quality control of the interacting partners of the model proteins will be validated in 2 steps: i) confirmation of interaction by co-immunoisolation followed by western blot; ii) evaluation of the role of the interactors by monitoring consequences on the substrate fate upon silencing of their expression or upon co-expression with the model substrate of their dominant negative mutants.