NEDDYLATION IS ESSENTIAL FOR ASTROCYTE DEVELOPMENT <EM>IN VITRO</EM> AND <EM>IN VIVO</EM>

Astrocytes, the star-shaped glial cells, are vital architects of brain development and function. Their development from radial glial progenitors follows a tightly regulated temporal sequence and spatial patterning. A promising and underexplored regulatory mechanism in astrocyte biology is neddylation, a post-translational modification involving the conjugation of the ubiquitin-like protein NEDD8 to target substrates. Neddylation regulates protein turnover, signal transduction, and cell cycle progression. Not surprisingly, neddylation has been linked to many diseases, including neurodegenerative diseases, cancer, and cardiac disease. Although its impact on neuronal development has been documented, its impact on the development of astrocytes remains unknown. Using different techniques, such as cytotoxicity assays, Western Blot, immunostainings, and pharmacological inhibition or activation of neddylation, we have demonstrated that neddylation is essential for astrocyte viability, differentiation, and maturation in vitro. Experiments in cultured astrocytes showed that acute and chronic activation and inhibition of neddylation activated astrocytes by increasing the number of amoeboid astrocytes, altering their morphology, and inducing cell death. Further, we showed that Neddylation needs to be finely regulated in human brain organoids to ensure the correct development of astrocytes. Lastly, we corroborated the data in vivo, inhibiting or over-activating Neddylation pharmacologically in pups and exploring the astrocytes in the brain one week after. Thus, we have demonstrated that neddylation is crucial for the development of astrocytes. It may uncover novel mechanisms by which astrocyte maturation and reactivity are controlled, with potential implications for modulating astrocyte responses in diseases from early neurodevelopmental disorders to adult neuroinflammatory diseases.