DPYSL2/CRMP2isoform B knockout in human iPSC-derived glutamatergic neurons confirms its role in mTOR signaling and neurodevelopmental disorders

ABSTRACT DPYSL2/CRMP2 is a microtubule-stabilizing protein crucial for neurogenesis and associated with numerous psychiatric and neurodegenerative disorders. DPYSL2 has multiple RNA and protein isoforms, but few studies have differentiated between them or explored their individual functions. We previously demonstrated in HEK293 cells that a schizophrenia -associated variant in the DPYSL2 B isoform ( DPYSL2 - B ) reduced the length of cellular projections, created a transcriptomic disturbance that captured schizophrenia etiology, and was acted upon by the mTOR pathway. In the present study, we follow up on these results by creating, to our knowledge, the first models of endogenous DPYSL2 - B knockout in human induced pluripotent stem cells and excitatory glutamatergic neurons. We use CRISPR/Cas9 to specifically knock out DPYSL2 - B and observe corresponding reduction of its RNA and protein. The average length of dendrites in knockout neurons was reduced up to 58% compared to controls. Transcriptome analysis reveals disruptions in pathways highly relevant to psychiatric disease including mTOR signaling, cytoskeletal dynamics, immune function, calcium signaling, and cholesterol biosynthesis. We also observed a significant enrichment of our differentially expressed genes in schizophrenia GWAS-associated loci. Our findings clarify the functions of the human DPYSL2 - B isoform and confirm its involvement in molecular pathologies shared between many psychiatric diseases.