Loss of function of FIP200 in human pluripotent stem cell-derived neurons leads to axonal pathology and hyperactivity.

FIP200 plays important roles in homeostatic processes such as autophagy and signaling pathways such as focal adhesion kinase (FAK) signaling. Furthermore, genetic studies suggest an association of FIP200 mutations with psychiatric disorders. However, its potential connections to psychiatric disorders and specific roles in human neurons are not clear. We set out to establish a human-specific model to study the functional consequences of neuronal FIP200 deficiency. To this end, we generated two independent sets of isogenic human pluripotent stem cell lines with homozygous FIP200KO alleles, which were then used for the derivation of glutamatergic neurons via forced expression of NGN2. FIP200KO neurons exhibited pathological axonal swellings, showed autophagy deficiency, and subsequently elevated p62 protein levels. Moreover, monitoring the electrophysiological activity of neuronal cultures on multi-electrode arrays revealed that FIP200KO resulted in a hyperactive network. This hyperactivity could be abolished by glutamatergic receptor antagonist CNQX, suggesting a strengthened glutamatergic synaptic activation in FIP200KO neurons. Furthermore, cell surface proteomic analysis revealed metabolic dysregulation and abnormal cell adhesion-related processes in FIP200KO neurons. Interestingly, an ULK1/2-specific autophagy inhibitor could recapitulate axonal swellings and hyperactivity in wild-type neurons, whereas inhibition of FAK signaling was able to normalize the hyperactivity of FIP200KO neurons. These results suggest that impaired autophagy and presumably also disinhibition of FAK can contribute to the hyperactivity of FIP200KO neuronal networks, whereas pathological axonal swellings are primarily due to autophagy deficiency. Taken together, our study reveals the consequences of FIP200 deficiency in induced human glutamatergic neurons, which might, in the end, help to understand cellular pathomechanisms contributing to neuropsychiatric conditions.