SYNTHETIC K+ CHANNELS FOR NEURONAL SILENCING (ERC AdG 695078)
We use a simple viral protein, the K+-selective channel Kcv (Plugge et al, Science 2000), to engineer synthetic light-gated ion channels for optogenetics. We have created BLINK1 (Cosentino et al, Science 2015) and BLINK2 (Alberio et al, Nat Methods 2018) and have validated them in vivo in zebrafish and mouse. We are currently working at engineering other channels that respond to other external stimuli to overcome the inherent limitation of low tissue penetration of visible light. Specifically, we are working at channels that can be activated by temperature, ultrasounds and magnetic fields (this project is supported by ERC Grant noMAGIC).
cAMP MODULATION IN HCN CHANNELS
A second line of research concerns cAMP modulation of Hyperpolarization activated Cyclic Nucleotide gated (HCN) channels which control heart rate and neuronal excitability in several brain areas. To this end we combine electrophysiology to structural approaches, X-ray (Lolicato et al, Nat Chem Biol 2014), NMR (Saponaro et al, PNAS 2014) and, more recently, cryoEM (Saponaro et al, Mol Cell 2021) to uncover the allosteric pathway of the cAMP signal within the HCN tetramer.
HCN FUNCTIONS AND CLINCIAL APPLICATIONS (Telethon_GGP20021)
Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels control neuronal excitability and their dysfunction has been linked to epileptogenesis but few individuals with neurological disorders related to variants altering HCN channels have been reported so far. Recently, patients with de novo or inherited HCN1 variants have been identified in a cohort of patients with catastrophic neonatal or infantile epileptic encephalopathy (EIEE) (Marini et al, Brain 2018; Nava et al, Nat Genet 2014). This strongly supports our central hypothesis that HCN1-related EIEE patients who are resistant to conventional antiepileptic drugs may be better treated with drugs that specifically target HCN channels. To this end, we will use our expertise in HCN channel biophysics, structural biology and Molecular Dynamics (MD) simulation together with an array of peptide tools and noncommercial molecules developed by us to correct or attenuate mutation-induced defects in HCN1 channels. Initial results obtained in vitro show that this strategy is successful and will be further validated in human iPSC-derived neurons from patients and in knock in mice with HCN1 mutations. The goal is to provide clinicians with robust indications and new tools to treat individual patients.