Transcriptional regulation by Egr3 in sympathetic nervous system development

描述（由申请人提供）：大多数关于交感神经系统（SNS）发育的已知信息仅限于具有基本功能的分子，如神经生长因子（NGF）。由于NGF和直接下游分子如其同源受体TrkA的损失导致SNS几乎完全损失，因此它们几乎没有提供严重程度低得多的SNS疾病的根本原因的线索。相反，对NGF信号传导下游基因的研究，如Egr 3，一种受NGF调节的转录因子，可能会更好地了解疾病。种系中Egr 3的缺失导致小鼠SNS发育异常和生理性自主神经功能障碍，与人类疾病相似。这项研究将进一步扩展我们对Egr 3功能的理解，方法是（1）检验Egr 3是SNS发育的交感神经元自主调节剂的假设，（2）定义它在交感神经元中独立于NGF信号传导的作用。这将通过产生新的小鼠模型来实现，该模型将特异性地删除交感神经元中的Egr 3并特异性地在交感神经元中过表达Egr 3。形态学和生理学检测将用于确定Egr 3的细胞自主操纵是否影响SNS发育。沿着确定Egr 3在SNS发育中的必要性，将进行研究以更好地理解其影响交感神经元分化的机制。因此，一个新发现的Egr 3靶基因，hs3st2，这是参与硫酸乙酰肝素蛋白聚糖（HSPG）的合成，将进一步检查。由于我们最初的研究已经证实Egr 3可以调节hs3st 2，并且先前的研究已经暗示了HSPGs在神经系统发育中的重要性，因此本研究的第二个目的是（1）更好地表征Egr 3对hs3st 2的调节，以及（2）确定hs3st 2是否在SNS发育中具有重要作用。将使用各种体外测定来确定Egr 3对hs3st 2的调节是直接的还是间接的，如果是直接的，将确定结合的确切位置。为了确定SNS发育是否需要hs3st2，将检查hs3st2种系缺失小鼠以评估它们是否具有任何SNS异常，这些SNS异常使人联想到在Egr 3缺陷小鼠中观察到的SNS异常。 公共卫生相关性：SNS易受各种发育和退行性疾病的影响，这些疾病困扰着全世界数百万人。一些SNS疾病已经可靠地与特定的基因突变相关，但其中绝大多数没有可识别的原因，从而突出了我们对SNS发展细节的了解仍然有限。Egr 3是一种基因，被证明参与SNS的发展，因此通过进一步阐明其作用的机制，我们将更好地了解SNS如何发展以及疾病如何损害其功能。

DESCRIPTION (provided by applicant): Most of what is known about the development of the sympathetic nervous system (SNS) is limited to molecules with essential functions such as Nerve Growth Factor (NGF). Since loss of NGF and immediate downstream molecules such as its cognate receptor, TrkA, result in near complete loss of the SNS, they offer little clues to the underlying causes of SNS diseases which are far less severe. Instead studies of genes further downstream of NGF signaling such as Egr3, a transcription factor shown to be regulated by NGF, will likely offer better insight into disease. Deletion of Egr3 in the germline results in mice with abnormalities in SNS development and physiologic dysautonomia bearing similarities to human diseases. The proposed study will further extend our understanding of Egr3 function by (1) testing the hypothesis that Egr3 is a sympathetic neuron autonomous regulator of SNS development and (2) defining the role it has independent of NGF signaling in sympathetic neurons. This will be accomplished by generating novel mouse models that will specifically delete Egr3 in sympathetic neurons and specifically over-express Egr3 in sympathetic neurons. Morphological and physiological assays will then be used to determine whether cell-autonomous manipulation of Egr3 affects SNS development. Along with determining the necessity of Egr3 in SNS development, studies will be done to better understand the mechanisms by which it affects sympathetic neuron differentiation. Thus, a newly identified Egr3 target gene, hs3st2, which is involved in heparan sulfate proteoglycan (HSPG) synthesis, will be further examined. Since our initial studies have confirmed that Egr3 can regulate hs3st2 and previous studies have implicated the importance of HSPGs in nervous system development, the second aim of this proposed study will be to (1) better characterize the regulation of hs3st2 by Egr3 and (2) determine whether hs3st2 has an important role in SNS development. Various in vitro assays will be used to determine whether regulation of hs3st2 by Egr3 is either direct or indirect and if it is direct, the exact location of binding will be established. To determine whether hs3st2 is required for SNS development, hs3st2 germline null mice will be examined to assess whether they have any SNS abnormalities that are reminiscent of those seen in Egr3-deficient mice. PUBLIC HEALTH RELEVANCE: The SNS is susceptible to a variety of developmental and degenerative diseases that afflict millions of people worldwide. A few of the SNS diseases have been reliably associated with specific gene mutations, but the vast majority of them have no identifiable cause, thus highlighting our still limited knowledge of the details of SNS development. Egr3 is a gene shown to be involved in SNS development and thus by further elucidating the mechanisms through which it acts we will gain a better understanding of how the SNS develops and how its function can be impaired by disease.