Roles for the stem cell factor LIN41 in ependymal cell function and hydrocephalus

脑积水可定义为脑室内脑脊液过度蓄积。这种情况可以是先天性的，也可以是后天的，由过量的脑脊液产生、吸收减少或体液流动障碍引起。儿童脑积水在出生时的发病率约为每1000个儿童中就有2个，这是公认的，儿童脑积水具有重要的遗传成分。目前已知的风险基因突变很可能只是非综合征性发育性脑积水的一部分。综合征形式的脑积水经常与神经管缺陷(NTDS)或纤毛疾病有关，如Bardet-Biedl综合征。在小鼠身上的研究表明，神经管和脑室系统的形态发生所需的遗传路径存在显著重叠。此外，在没有闭合缺陷的情况下，与NTDS相关的几个通路的破坏，包括平面细胞极性，可能会导致发育性脑积水。纤毛病可以影响原发纤毛，也可以影响活动纤毛。虽然初级纤毛被认为协调生长和形态发生信号，但活动的纤毛有助于脑脊液的流动。初级纤毛或活动纤毛的缺陷可导致脑积水。我们一直在研究Lin41基因产物的分子功能和发育作用。LIN41在小鼠体内的零突变是胚胎致死的，并导致神经管关闭失败。NTD的表型被归因于神经元的过早分化，然而，神经前体细胞靶向缺失的结果尚未报道。为了更好地了解Lin41在胚胎和出生后皮质发育中的功能，我们产生了一个条件突变小鼠，并使用Emx1-Cre特异性地删除了端脑背侧放射状胶质细胞中的Lin41。使用这个模型，我们没有观察到对皮质神经发生本身的任何有害影响。然而，超过25%的受影响突变体会在出生后发展为脑积水。这一结果与我们之前的发现一致，即Lin41在出生后脑室管膜细胞中特异表达。在我们的初步数据中，我们提出了LIN41是室管膜细胞功能成熟所必需的实验证据。我们还发现，在培养的室管膜细胞中，Lin41基因的缺失会导致纤毛长度和搏动频率的减少。这项研究旨在更全面地描述脑积水的表型以及室管膜纤毛发生和功能的缺陷。我们的实验将利用我们的条件突变进行体内研究，并结合使用原代室管膜细胞培养模型进行分子水平的研究。从机制上，我们将测试LIN41作为RNA结合蛋白和作为E3泛素连接酶在室管膜细胞分化中的几个拟议功能的相关性。总之，这些模型系统和我们的实验计划将使我们能够探索发育性脑积水的新途径和发病机制。

Hydrocephalus can be defined as excessive accumulation of cerebrospinal fluid (CSF) in the brain ventricles. The condition can be either congenital or acquired and is caused by excess CSF production, reduced absorption or impaired fluid flow. With an incidence of about 2 per 1000 children at birth, it is well established that pediatric hydrocephalus has a significant heritable component. Mutations in currently known risk genes most likely account for only a portion of nonsyndromic developmental hydrocephalus. Syndromic forms of hydrocephalus are frequently associated with neural tube defects (NTDs) or ciliopathies such as Bardet-Biedl Syndrome. Studies in the mouse have revealed significant overlap in the genetic pathways required for morphogenesis of the neural tube and the ventricular system. Moreover, disruption of several pathways associated with NTDs, including planar cell polarity, can lead to developmental hydrocephalus in the absence of closure defects. Ciliopathies can affect either primary or motile cilia. Whereas the primary cilium is thought to coordinate growth and morphogenic signals, motile cilia contribute to CSF flow. Defects in either primary or motile cilia can lead to hydrocephalus. We have been studying the molecular functions and developmental roles of the Lin41 gene product. Null mutations of Lin41 in the mouse are embryonic lethal and cause failure of neural tube closure. The NTD phenotype has been attributed to premature neuronal differentiation, however the results of targeted deletion in neural progenitors have not yet been reported. To better understand Lin41 function in embryonic and postnatal cortical development, we have generated a conditional mouse mutant and used Emx1-Cre to specifically delete Lin41 in radial glia of the dorsal telencephalon. Using this model, we have not observed any deleterious effect on cortical neurogenesis per se. However, more than 25% of the affected mutants develop postnatal hydrocephalus. This result is consistent with our previous finding that Lin41 is specifically expressed in ependymal cells of the postnatal brain. In our preliminary data, we present experimental evidence that LIN41 is required for functional maturation of ependymal cells. We also show that deletion of Lin41 in cultured ependymal cells leads to a reduction in cilia length and beating frequency. This research proposal is designed to more fully characterize the hydrocephalus phenotype and the defects in ependymal ciliogenesis and function. Our experiments will make use of our conditional mutant for in vivo studies coupled to studies at the molecular level using a primary ependymal cell culture model. Mechanistically, we will test the relevance of several proposed functions for LIN41 as RNA binding protein and as E3 ubiquitin ligase for ependymal cell differentiation. Together, these model systems and our experimental program will allow us to explore new pathways and pathomechanisms for developmental hydrocephalus.