Argonaute2 in excitatory neurons functionally regulates cellular metabolism and cerebral vascularization during brain development

Argonaute2 （Ago2）参与核糖核酸内切酶活性，参与microRNA通路，在病理生理胁迫下调控基因表达起重要作用。Ago2缺失导致小鼠中枢神经系统异常形成和产前胚胎早期死亡，提示Ago2在中枢神经系统发育中起重要作用。我们的初步数据显示，表达vglut2的兴奋性神经元（Vglut2-Cre， Ago2flox/flox小鼠）中Ago2的缺失，而表达vgat -抑制性神经元（Vgat-Cre， Ago2flox/flox小鼠）中Ago2的缺失，1)会干扰小鼠的发育并引发出生后小鼠的早期死亡；2)损害海马神经元的细胞代谢，包括线粒体功能；3)强烈破坏发育中的大脑皮层的脑血管形成；4)改变Cadm1， Mecp2和Ucp2基因的表达，这些基因在发育过程中对神经元和突触功能至关重要，并且包含ago2依赖性mirna的靶位点，如microRNA-22 (MiR-22), miR-24， miR-124和miR-128。根据我们的初步研究，我们假设Ago2作为核糖核酸内切酶蛋白和mirna介导的基因沉默的调节因子，有助于神经元发育和脑血管形成。因此，研究Ago2在兴奋性神经元发育过程中的功能作用是本课题研究的总体科学目标。我们将研究Ago2作为核糖核酸内切酶蛋白和mirna介导的基因沉默的调节剂如何通过调节其基本靶基因来促进神经元发育和脑血管形成。具体来说，我们将讨论1)Ago2在兴奋性神经元中对神经发生、神经元迁移和分化的功能作用；2)病理生理应激下Ago2及其靶向mirna对神经元的细胞代谢调控；3)兴奋性神经元中的Ago2如何调控脑血管生成和血管形成。此外，尽管Vgat-Cre、Ago2flox/flox小鼠直到P16才表现出明显的病理现象并正常存活，但我们仍有兴趣研究4)Ago2在表达vgat的抑制性神经元中对神经回路和小鼠行为的功能作用。本项目的研究结果将扩展我们对Ago2及其相关mirna在大脑发育过程中的功能作用和机制的理解。

Argonaute2 (Ago2) processes an endoribonuclease activity and is involved in microRNA pathway, which plays an important role in regulation of the gene expression upon pathophysiological stresses. Ago2 deletion in mice results in abnormal formation of the CNS and early death of prenatal embryos, suggesting an essential role of Ago2 in the CNS development. Our promising preliminary data show that loss of Ago2 in Vglut2-expressing excitatory neurons (Vglut2-Cre, Ago2flox/flox mice), but not in Vgat-expressing inhibitory neurons (Vgat-Cre, Ago2flox/flox mice), 1) disturbs mouse development and triggers early death of postnatal mice; 2) impairs cellular metabolism, including mitochondrial function, in hippocampus neurons; 3) strongly disrupts the brain blood vessel formation in the developing cerebral cortex; 4) alters the expression of Cadm1, Mecp2, and Ucp2 genes, which are critical for neuronal and synaptic function during development and contain target sites for Ago2-dependent miRNAs, like microRNA-22 (MiR-22), miR-24, miR-124, and miR-128. Based on our preliminary studies, we hypothesize that Ago2 as an endoribonuclease protein and a regulator of miRNA-mediated gene silencing contributes to neuronal development and brain vascularization. Therefore, the overall scientific goal of this proposal is to find out the functional roles of Ago2 in excitatory neurons during development. We will investigate how Ago2 as an endoribonuclease protein and a regulator of miRNA-mediated gene silencing contributes to neuronal development and brain vascularization via regulating its essential target genes. Specifically we will address 1) the functional role of Ago2 in excitatory neurons on neurogenesis, neuronal migration and differentiation; 2) the cellular metabolic regulation of Ago2 and its targeted miRNAs on neurons upon pathophysiological stress; and 3) how Ago2 in excitatory neurons regulates brain angiogenesis and vascularization. Furthermore, although Vgat-Cre, Ago2flox/flox mice did not show obviously pathological phenomenon until P16 and survive normally, it is still interest to investigate 4) the functional role of Ago2 in Vgat-expressing inhibitory neurons on neural circuit and mouse behavior. The findings of this project will extend our understanding of the functional role and mechanism of Ago2 and its associated miRNAs during brain development.