Cortactin and Spine Dysfunction in Fragile X

描述(由申请人提供)：自闭症是一种毁灭性的疾病，对患者、他们的家人和国家经济造成重大损失。目前尚无治疗方法。 影响75%以上自闭症患者的认知障碍(例如，学习和记忆障碍)。在许多自闭症相关障碍和自闭症共病的模型中，如脆性X综合征、Rett综合征和Angelman综合征，树突棘形态存在显著的异常，这与记忆编码的突触机制长时程增强(LTP)的稳定受损有关。综上所述，这些发现表明，脊椎肌动蛋白细胞骨架的缺陷可能是不同来源的自闭症相关疾病的认知缺陷的基础。脆性X综合征(FXS)的Fmr1基因敲除(KO)小鼠模型是脊柱异常和LTP损伤的最好特征，FXS是一种自闭症的高(约30%)共病情况。具体地说，这些突变体在通过Rac GTP酶传递信号方面存在缺陷，在脊丝状肌动蛋白(F)肌动蛋白活性驱动的变化中稳定下来，以及LTP的巩固。申请人的研究表明，皮质蛋白是一种脊椎蛋白，通过肌动蛋白和微管系统稳定肌动蛋白网络分支点并保护F-肌动蛋白免于降解，在LTP诱导后，在Fmr1-KO脊柱上受到损害。这表明KO中的F-肌动蛋白稳定化缺陷可能反映了对皮质肌动蛋白信号的干扰。拟议的研究将以这些发现为基础检验以下具体假设：(A)调节蛋白质亚细胞运动的皮质肌蛋白丝氨酸异常磷酸化和乙酰化均源于单个分子损伤，并导致KO小鼠LTP诱导后运动受损的表型；(B)(在体内)学习可激活WT而不是KO小鼠的这些突触过程。有三个具体目标。AIM 1将测试KOS中突触RAS或PP2A的基础水平或激活是否受损(这两个靶点都影响皮质蛋白的磷酸化和乙酰化途径)。Aim 2将测试通过MAPK和/或HDAC6的信号是否对活性诱导的皮质蛋白转位是必需的，这些信号有助于皮质肌蛋白丝氨酸的磷酸化和乙酰化。依赖于海马体的空间学习激活了WTS中皮质肌动蛋白的突触信号，如果该信号在活体Fmr1-KOS中减弱或缺失。通过询问与F-肌动蛋白稳定性受损相关的突触机制并确定这些异常是否也存在于行为动物中，拟议的研究将有助于我们了解正常和FXS模型小鼠的突触可塑性，并为FXS和其他自闭症条件下记忆功能的正常化提供治疗靶点。最后，Aim 3将测试依赖于海马体的空间学习是否激活WTS中皮质肌动蛋白的突触信号，以及在活体Fmr1-KO中该信号是否减弱或缺失。通过询问与F-肌动蛋白稳定性受损相关的突触机制并确定这些异常是否也存在于行为动物中，拟议的研究将有助于我们了解正常和FXS模型小鼠的突触可塑性，并为FXS和其他自闭症条件下记忆功能的正常化提供治疗靶点。

DESCRIPTION (provided by applicant): Autism is a devastating condition which takes a significant toll on patients, their families, and the national economy. There are no treatments for cognitive impairments (e.g., learning and memory deficits) that affect over 75% of autistic patients. In models of many autism-associated disorders and conditions with comorbidity for autism, such Fragile X, Rett, and Angelman Syndromes, there are significant abnormalities in dendritic spine morphology, which are associated with impairments in the stabilization of long-term potentiation (LTP), a synaptic mechanism of memory encoding. Together, these findings suggest that defects in the spine actin cytoskeleton may underlie cognitive deficits in autism-associated conditions of different origin. Spine abnormalities and LTP impairments are best characterized in the Fmr1-knockout (KO) mouse model of fragile X syndrome (FXS), a condition with high (~30%) comorbidity for autism. Specifically, these mutants exhibit defects in signaling through Rac GTPase, stabilization of activity-driven changes in spine filamentous (F) actin, and consolidation of LTP. Studies by the applicant have demonstrated that movement of cortactin, a spine protein which stabilizes actin network branch points and protects F-actin from degradation, via both actomyosin and microtubule systems, is impaired at Fmr1-KO spines following LTP-induction. This suggests that the F- actin stabilization deficits in KOs may reflect disturbances in signaling to cortactin. The proposed research will build on these findings to test the specific hypotheses that (a) abnormal cortactin serine phosphorylation and acetylation, which regulate the protein's subcellular movement, both originate from a single molecular impairment and contribute to the phenotype of impaired movement following LTP induction in KOs and that (b) learning (in vivo) activates these synaptic processes in WT but not KO mice. There are 3 specific aims. Aim 1 will test if basal levels or activation of synaptic Ras or PP2A are impaired in KOs (both of these targets influence the cortactin phosphorylation and acetylation paths). Aim 2 will test if signaling through MAPK and/or HDAC6, which contribute to cortactin serine phosphorylation and acetylation, are necessary for activity- induced cortactin translocation. hippocampus-dependent spatial learning activates synaptic signaling to cortactin in the WTs and if this signaling is attenuated or absent in Fmr1-KOs in vivo. Through interrogating synaptic mechanisms associated with impairments in F-actin stabilization and determining if these abnormalities are also present in the behaving animal, the proposed studies will contribute to our understanding of synaptic plasticity in both normal and FXS model mice and offer therapeutic targets for normalization of memory function in FXS and other autistic conditions. Finally, Aim 3 will test if hippocampus-dependent spatial learning activatessynaptic signaling to cortactin in the WTs and if this signaling is attenuated or absent in Fmr1-KOs in vivo. Through interrogating synaptic mechanisms associated with impairments in F-actin stabilization and determining if these abnormalities are also present in the behaving animal, the proposed studies will contribute to our understanding of synaptic plasticity in both normal and FXS model mice and offer therapeutic targets for normalization of memory function in FXS and other autistic conditions.