Neuronal PARP activity in fetal alcohol spectrum disorders

胎儿酒精谱系障碍（FASD）是智力障碍的主要已知和可预防的原因。 残疾。它们会损害执行功能，包括工作记忆，这是一种高度依赖于大脑的功能。 内侧前额叶皮质（mPFC）。这个大脑区域是最后成熟的区域之一，在人类的第三个三个月 以及出生后的前10天。这是mPFC树突的脆弱时期，它们的生长 易受环境污染如乙醇（EtOH）的破坏。聚ADP核糖聚合酶 蛋白质涉及几种细胞功能，包括调节基因表达。PARP合成 并将聚（ADP-核糖）（PAR）链（PARylating）附着到其靶上。PARP酶可以影响基因 通过对表观遗传酶KDM 4D进行PAR化来表达。这降低了KDM 4D删除 在组蛋白H3（H3 K9 me 2）处的转录抑制的二甲基化赖氨酸9。PARP介导的基因沉默 也可以通过PARylating转录因子过氧化物酶体增殖物激活 受体γ（PPARγ）。我们的基本假设是EtOH诱导PARP 1活性， 将PAR基团添加到已知的PARP 1靶点，如PPARγ和KDM 4D。这种翻译后 修饰将降低PPARγ和KDM 4D结合DNA或染色质的能力，从而导致 神经发育基因表达、树突分支和工作记忆。这种假设是 我们的初步数据支持，其中EtOH增加PARP活性并减少Bdnf IV，IXa和Klf 4 原代皮层神经元培养物中的mRNA表达。使用PARP抑制剂时，这些变化是可逆的。 作为PARP和PPARγ之间的直接联系，我们发现PARP抑制增加了PPARγ与 体外的BDNF IV和Klf 4启动子。在体内，新生儿EtOH处理诱导PARP活性，这与 PPARγ DNA结合能力降低，Bdnf IV mRNA表达减少。31天后， 在最终剂量时，Bdnf IV表达的减少在EtOH暴露的大鼠中持续存在。在第一个目标中，我们计划 剖析PARP与发育基因表达变化的分子机制， mPFC，重点关注神经元特异性变化。为了确定PARP在EtOH诱导的BDNF中的作用， 和Klf 4基因表达沉默，我们将尝试通过施用PARP来防止表达变化。 抑制剂ABT-888对EtOH处理的大鼠的影响。我们还将分析PARP介导的转录抑制是否 通过使用神经元培养物对PPARγ和KDM 4D的翻译后修饰发生。第二个目标，我们 将确定PARP在mPFC树突状分支中的重复缺陷中的作用， 在FASD模型中观察到神经突发生。在第三个目标中，我们将研究PARP在晚期妊娠中的作用 等效EtOH暴露诱导的空间工作记忆缺陷。已知PARP抑制剂是 神经保护，目前正在进行其他疾病的临床试验，并穿过血脑屏障。 因此，这些研究的结果可能是未来药理学发展的一个有前途的途径。

Fetal alcohol spectrum disorders (FASD) are the leading known and preventable causes of intellectual disability. They impair executive functions including working memory, a function highly dependent on the medial prefrontal cortex (mPFC). This brain region is one of the last to mature, the third trimester in humans and the first 10 days after birth in rats. This is a vulnerable period for mPFC dendrites where their growth is prone to disruption from environmental insults, such as ethanol (EtOH). Poly ADP ribose polymerases (PARP) proteins are implicated in several cellular functions, including regulating gene expression. PARP synthesizes and attaches poly (ADP-ribose) (PAR) chains (PARylating) to its targets. PARP enzymes can affect gene expression by PARylating the epigenetic enzyme KDM4D. This reduces KDM4D’s ability to remove the transcriptionally repressive, dimethylated lysine 9 at histone H3 (H3K9me2). PARP-mediated gene silencing can also be accomplished by PARylating the transcription factor Peroxisome proliferator-activated receptor gamma (PPARγ). Our underlying hypothesis is that EtOH induces PARP1 activity, promoting the addition of PAR groups to known PARP1 targets such as PPARγ and KDM4D. This post-translational modification would then reduce PPARγ and KDM4D’s ability to bind DNA or chromatin resulting in changes to neurodevelopmental gene expression, dendritic arborization, and working memory. This hypothesis is supported by our preliminary data in which EtOH increased PARP activity and reduced Bdnf IV, IXa, and Klf4 mRNA expression in primary cortical neuron cultures. These changes were reversible with a PARP inhibitor. As a direct connection between PARP and PPARγ we found that PARP inhibition increased PPARγ binding to Bdnf IV and Klf4 promoters in vitro. In vivo, neonatal EtOH treatment induced PARP activity, and this coincided with a decrease in PPARγ DNA binding ability and reduced Bdnf IV mRNA expression. Thirty-one days after the final dose, the reduction in Bdnf IV expression persisted in EtOH exposed rats. In the first aim, we plan to dissect the molecular mechanisms connecting PARP to changes in developmental gene expression in the mPFC with a focus on neuron-specific changes. In order to establish the role of PARP in EtOH induced Bdnf and Klf4 gene expression silencing we will attempt to prevent expression changes by administering a PARP inhibitor ABT-888 to EtOH treated rats. We will also dissect whether PARP mediated transcriptional repression occurs via post-translational modifications to PPARγ and KDM4D using neuron cultures. In the second aim, we will establish the role of PARP in the much-replicated deficits in mPFC dendritic arborization and neuritogenesis observed in FASD models. In the third aim, we will study the role of PARP in third trimester equivalent EtOH exposure-induced spatial working memory deficits. PARP inhibitors are known to be neuroprotective, are currently undergoing clinical trials for other disorders, and cross the blood-brain barrier. Therefore, the results of these studies may be a promising avenue for future pharmacological development.