Signalling Pathways And Neuroprotection
Protein-protein interactions
Upstream JNK activators: We found a rapid and selective activation of MKK7 after NMDA-stress, while the only other JNK activator, MKK4, was unaffected.
Our aim: To inhibit JNK signalling pathway targeting the upstream JNK’s activator MKK7 (Fig. 2).
1. MKK7 Silencing
- To inactivate MKK7 we use lentiviruses because of the particular capability of integrating genetic material into the genome of non-dividing cells stably. Our lentiviral vector will carry a single si-RNA duplex of MKK7 (a kind gift of Prof. Franzoso).
2. Gadd45β MKK7 Inhibitor
The aim is to test in vitro the specific inhibitor: Gadd45β, a molecule active on the upstream JNK activator MKK7 (Fig. 3). Gadd45β binds to MKK7 directly and blocks its catalytic activity. Furthermore, this protein is of particular interest to us since it is a powerful inhibitor of MKK7, the binding between Gadd45β/MKK7 being tighter than JIP1/MKK7. The endogenous Gadd45β interacts to MKK7 through direct, high-affinity contact (Papa et al 2004) but not with the other JNK upstream kinase, MKK4. For this study we initially plan to use a viral system that will allow us to observe the role of this pathway in excitotoxicity, with a final goal of producing a cell-permeable peptide with a more specific effect in the prevention of neuronal death.
3. JNK3 specific inhibition
β-arrestin2/JNK3 interaction
The JNK3 isoform is of particular interest in our studies since it is specifically expressed in the brain, and to a much lesser extent to other tissues, i.e heart. Notably, it has been shown that JNK3 deletion is protective against ischemia/hypoxia induced brain injury. The aim of this project is to identify ways for inhibiting the JNK3 action.
Interestingly, it was recently shown that the b-arrestin-2 scaffold protein promotes c-JNK3 activation through binding. We plan to deepen the understanding of such interaction by using computer DOCKING programs (see Prof. Cosentino) and thus identify sites of binding. The final goal of this study would be to produce cell-permeable peptides to block specifically the JNK3 isoform and explore their effects on neuroprotection.