PROJECT SUMMARY Many cognitive disorders arise from abnormalities at the early stage of brain development. Alternative splicing, the inclusion or exclusion of specific exons in mRNAs, is a key component in controlling normal neuronal development but affected in neuropsychiatric diseases represented by schizophrenia (SCZ). Recent discoveries indicate that long non-coding RNAs (lncRNAs), a class of non-coding RNAs longer than 200 nucleotides which do not encode for proteins, play sophisticated roles in gene regulation including alternative splicing. IncRNAs are poorly conserved in general and highly expressed in the human brain. Abnormalities in lncRNA expression are implicated in neurodegenerative and neuropsychiatric diseases. A particular human lncRNA of interest is GOMAFU, which is abundantly expressed in human iPSC-derived neural progenitor cells (hNPCs) and brain neurons but negligible in glia. GOMAFU is a nuclear lncRNA affected in SCZ and known to regulate alternative splicing of a number of risk factor transcripts involved in SCZ. However, molecular mechanisms regulating GOMAFU are undefined. Moreover, although GOMAFU was postulated to regulate alternative splicing through sequestering neuronal RNA-binding protein (RBP) splicing factors, RBPs interacting with GOMAFU still remain elusive. Emerging evidence, including our preliminary data, indicates that GOMAFU may form a functional pathway with an RBP called Quaking I (QKI), another SCZ risk factor known to regulate alternative splicing in neural progenitor cells (NPCs) and neuron-glia lineage development. Recombinant QKI was shown to interact with GOMAFU in vitro. Furthermore, during development of human iPSC-derived cortical neurons, the decline of QKI conversely associates with increased GOMAFU expression. Importantly, elimination of QKI leads to GOMAFU up-regulation in a human NPC cell line. Thus, I hypothesize that QKI-5 suppresses GOMAFU expression in human NPCs whereas GOMAFU controls splicing in human neuron development through sequestration of nuclear splicing factors, including QKI-5. The goal of this project is to delineate the function of the QKI-GOMAFU SCZ risk factor pathway in alternative splicing during human neuronal development. In Aim 1, I will determine whether QKI-5 binds and suppresses GOMAFU biogenesis in a hNPC cell line and human iPSC-derived NPCs. In Aim 2, I will determine whether GOMAFU regulates QKI nuclear distribution and splicing function in hNPCs. Moreover, I will utilize the recently developed comprehensive identification of RNA binding proteins (ChIRP) assay to identify GOMAFU-bound RBP splicing factors and elucidate how GOMAFU modulate their function in alternative splicing during hNPC development.