Dissecting the molecular and cellular mechanisms of neurite formation through two understudied molecules in neurodevelopment and exploration on a potential peptide therapy in a mouse model for autism study

Xiaonan Liu

doi:10.17918/00001815

Neuritogenesis is the first step for the neuron to establish the polarity after migration. Malformation of neurite structures may lead to abnormalities in neuronal activity and connectivity. Altered neuronal activity and connectivity are seen as major defects in neurodevelopmental disorders, such as autism spectrum disorders. We dissected roles of several molecules in neuritogenesis to expand our understanding of this critical step of neurodevelopment. Glutathione S-transferase Pi (GSTP) protein belongs to the superfamily of glutathione transferases and conventionally is known for catalytic function in phase II metabolism. Three isoforms of Gstp are expressed in the developing mouse cerebral cortex. Knocking down of the two major isoforms at the early stage of neurite formation impeded the outgrowth of mouse cortical neurons in multiple aspects, including the altered growth dynamics in apical dendrites, misoriented apical dendrites, and decreased neurite numbers. We were the first to study Gstp in neurodevelopment at the time of the publication. Referencing studies in other research fields, we found Gstp might regulate activity of several kinases, including CDK5 and JNK. By applying a JNK inhibitor, we found that the reduced neurite number in the Gstp 1/2 depleted neurons was rectified to the control level. Further experiments confirmed that JNK phosphorylation was increased by Gstp depletion in the cells. This work identified that Gstp 1 and 2 are involved in neurite formation through regulating JNK activity. Myosin 1C (Myo1c) is an unconventional Myosin associated with a neurodevelopmental disorder, Miller-Dieker Syndrome. No previous study had investigated Myo1c in neurodevelopment. We studied Myo1c expression in cortical neurons and its function in neurite formation. Myo1c was expressed in the early developmental stage of mouse cerebral cortex, and a shift of cellular localization was identified over the four stages of neurite formation. By depleting Myo1c using shRNA, we found that Myo1c was essential for the soma orientation at early stages of neurite formation, for establishing neurite number, and for regulation of dendritic spine formation in mouse upper layer cortical pyramidal neurons. In matured neurons, Myo1c depletion led to an altered spontaneous calcium activity. Literature show that Myo1c has an extensive role in F-actin regulation in other cell types. Using fixed cells and live cells to study Myo1c's function in F-actin organization in the mouse cortical neurons, we found that Myo1c regulated F-actin localization at the peripheral regions of neurite-initiating neurons. The motor activity of Myo1c was required for this regulation. Moreover, F-actin retrograde flow in the filopodia was also reduced in the Myo1c-depleted cells. Our findings in neurobiology will help to uncover the etiology of neurodevelopmental disorders. Another part of this dissertation stemmed from our lab's discovery in the role of PEDF in neuritogenesis. Our previous study identified PEDF as a positive regulator for neurite formation (Blazejewski, Bennison et al. 2021). For exploring the therapeutic potential in treating neurodevelopmental disorders, we used short fractions of PEDF peptides containing different functional domains in a mouse model for autism study. The short peptides showed different effects on neurite outgrowth. In the course of further investigation, we found that the treatment with one short peptide restored changes in spine morphology in the cultured neurons from the mouse model of autism, as well as the altered spontaneous calcium activity. Our PEDF peptide study also showed the potential of the PEDF peptide treatment in ameliorating neuronal morphological and spontaneous activity abnormalities. Overall, we advanced the understanding of the molecular and cellular mechanisms of neuronal morphogenesis and provided evidence that supports further discovery of a novel treatment for neurodevelopmental disorders with neuronal morphogenesis abnormalities.

Dissecting the molecular and cellular mechanisms of neurite formation through two understudied molecules in neurodevelopment and exploration on a potential peptide therapy in a mouse model for autism study

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Dissecting the molecular and cellular mechanisms of neurite formation through two understudied molecules in neurodevelopment and exploration on a potential peptide therapy in a mouse model for autism study

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