FUNCTIONAL ANALYSIS OF IPS CELL-DERIVED NEURONS FROM A PATIENT WITH 3Q29 DELETION

Abstract Background Since the prevalence of patients with psychiatric disorders has recently been increasing worldwide (Patel, 2018; Rehm, 2019) the understanding of molecular and cellular pathogenesis of psychiatric disorders is an urgent issue. Recent genetic studies have identified genetic mutations and genome copy number variants (CNVs) that are strongly associated with psychiatric disorders. Among these genetic variants, recurrent CNVs have been suggested to greatly contribute to genetic vulnerability to psychiatric disorders. The 3q29 deletion is an approximately 1.6 Mb deletion on chromosome 3 and contains 21 protein-coding genes, of which incidence of this deletion is 1 in 30,000 to 40,000 births (Cox, 2015; Mulle, 2015). Importantly, the 3q29 deletion is strongly associated with multiple psychiatric disorders, of which core symptoms are reduced verbal communication, hallucination, and delusion, suggesting that the molecular and cellular pathogenesis of 3q29 deletion is an important clue for elucidating the mechanism of psychiatric disorders. However, the detailed pathogenesis of 3q29 deletion remains largely unclear (Baba, 2019; Takemoto, 2021). In this study, using previously established iPS cells from a patient with 3q29 deletion, we analyzed the pathological effect of 3q29 deletion on developmental and functional abnormalities of neurons. Methods The patient-derived and control neural stem cells from iPS cells were seeded in a differentiation medium. On day 2, which was in an early stage of neural differentiation, the cells were subjected to immunocytochemical analysis with antibodies against neural marker MAP2 and HuC/D. Furthermore, total RNAs from neurons were isolated and were subjected to reverse transcription and real-time PCR to analyze mRNA expression levels of these neural marker genes. For analyzing synaptic function of iPS cell- derived neurons, whole-cell patch-clamp recordings were made from iPS cell-derived neurons at 4 weeks after differentiation. We also employed the MED64 recording system to investigate synaptic function of iPS cell-derived neurons. Results The proportion of MAP2 and HuC/D neurons was significantly lower in the patient-derived cells than in the control cells. We also demonstrated that the mRNA expressions of MAP2 and ELAVL3/4 (encoding HuC/D) genes were decreased in the patient-derived cells as compared to those in control cells, suggesting that neural differentiation from neural stem cells into neurons is impaired in patient-derived cells. Using electrophysiological methods, we demonstrated multiple altered synaptic functions in patient-derived neurons as compared to control neurons. Discussion & Conclusion Since 3q29 deletion confers a high risk for psychiatric disorders, including autism spectrum disorders and schizophrenia, our current study may provide important insights into the pathogenesis of psychiatric disorders. For further understanding the molecular pathogenesis of 3q29 deletion, we need to identify responsible genes involved in the impaired neurodevelopment as well as altered synaptic function in the 3q29 deleted region. Considering that psychiatric disorders are etiologically and genetically heterogeneous, molecular elucidation of strongly disease-associated 3q29 deletion leads to stratification of psychiatric disorders by molecular and cellular pathogenesis and ultimately to the development of personalized medicine. References 1V. Patel et al., Lancet, 2018 Oct 27; 392(10157):1553-1598 2M. Rahimi-Balaei et al., Curr. Psychiatry Rep., 2018 Dec 17; 12:484 3Devin M Cox et al., Clin. Dysmorphol., 2015 Jul; 24(3):89-94 4J. G. Mulle, Mol. Psychiatry, 2015 Sep; 20(9):1028-1029 5Baba et al., Neuropsychopharmacol., 2019 Nov; 44(12):2125-2135 6Takemoto et al., Mol. Brain, 2022 Apr 11; 15(1):34