Effects of FUS/TLS and its mutations on the in vitro properties of Neural Progenitor cell lines and their neural progeny

Amyotrophic Lateral Sclerosis (ALS) is a neurodegenerative disorder affecting upper and lower motor neurons in the brainstem and spinal cord. Mutations in sarcoma/translocated in lyposarcoma protein (FUS/TLS) cause ALS and these mutations are mainly dominant. FUS shows both nuclear and cytoplasmatic localization, although several mutations cause its accumulation in the cytoplasm, where it may form inclusions. FUS is a RNA/DNA binding protein, which regulates transcription, RNA, and miRNA processing and transport. Although ALS is an adult-onset neurodegenerative disorder, understanding the role of FUS in nervous system development can help to unveil several aspects of the pathology. Neural progenitor (NP) cultures represent a useful cellular model to recapitulate some events of nervous system formation. With the aim to clarify the effects of FUS on NP fate we established and characterized NP lines derived from E13.5 CD1 mice spinal cord or cortex. These cell lines were stably engineered with doxycycline inducible plasmids, containing either human FUS wild type (WT-FUS), or a FUS variant causing ALS (mFUS; substitution P525L), or a control plasmid (RFP). This system allows us to efficiently express, in a pharmacologically regulated manner, the transgenes either in undifferentiated NP cells or in their differentiated progeny (neurons and glia). We found that WT-FUS is mainly localized in the nucleus, whereas m-FUS is clearly mislocalized, and accumulated in the cytoplasm. Over-expression of WT-FUS or mFUS reduces the number of differentiated neurons. We are investigating whether this effect is related to neurotoxicity or to changes in NP commitment.