Role of post-transcriptional gene regulation in differentiating retinal ganglion cells

神经元是一种特殊的细胞，通过树突和轴突相互交流。在它们的生长和分化过程中，神经元树突和轴突响应营养因子信号从细胞体中伸出和伸长，直到它们到达目标。这一过程被称为神经新生，对于神经元和目标细胞之间建立功能性和可存活的连接至关重要。不能建立这种联系的神经元会发生细胞死亡。神经突需要蛋白质、脂质和其他分子的持续供应，以延长和正常运作。这些分子通常在细胞体中产生，然后通过复杂的分子网络沿着神经突运送到突触。除了这种穿梭机制，最近的研究表明，许多蛋白质实际上是在需要的时候和地方在树突中局部产生的。这个过程使神经元对特定的刺激有更快更精确的反应。蛋白质是由信使RNA （mRNA）分子产生的，这种分子通常在靠近细胞核的地方被发现。最近在神经突的远端发现了mrna，在那里它们被翻译成蛋白质。mrna稳定性和半衰期的调控是细胞调节蛋白质合成的机制之一。在用于调节这一过程的几种方法中，人们发现microRNAs，即非常短的RNA分子，起着非常重要的作用。通过与mRNA的特定部分结合，它们触发mRNA的降解，从而间接减少特定蛋白质的产生。因此，在我们的实验建议中，我们希望研究亚细胞定位和microrna在神经突伸长中的作用。这将通过分析Dicer的定位和表达来补充，Dicer是参与成熟microrna产生的关键酶之一。我们特别想了解Dicer和microrna是否在延长的神经突中被重新定位，以及它们是否在这一过程中起致病作用。这将使我们更好地了解生长中的神经元是如何调节蛋白质合成的，并通过实验调节Dicer的活性，了解这种机制的任何损害是否会对神经细胞造成任何损害。此外，根据我们最近关于microRNAs调控Brn-3b转录因子mRNA的研究结果，我们想分析在视网膜神经节细胞（RGC）诱导的延长神经突中发生的类似机制。Brn-3b在RGC发育过程中表达，对于它们的生存至关重要，在缺乏Brn-3b的小鼠中，高达70%的RGC损失证明了这一点。因此，对外植RGC中Brn-3b转录后调控的分析将成为RGC中转录后调控整体作用的范例。

Neurons are specialized cells which communicate with each other and other cells via processes called dendrites and axons. During their growth and differentiation, neuronal dendrites and axons protrude and elongate from the cell body in response to trophic factor cues until they reach their target. This process, called neuritogenesis, is essential for the proper establishment of functional and viable connections between neurons and their targets. Neurons that fail to establish such contact undergo cell death. Neurites need a constant supply of proteins, lipids and other molecules in order to elongate and function properly. These molecules are generally produced in the cell body and then transported along the neurite to the synapse by an elaborate network of molecules. Alongside this shuttling mechanism, it has been recently shown that many proteins are actually produced locally in the dendrite when and where they are needed. This process allows neurons to have a faster and more precise response to specific stimuli. Proteins are produced from molecules called messenger RNA or mRNA which are normally found close to the cellular core, or nucleus. Very recently mRNAs have been found in distal parts of neurites where they are translated into proteins. The regulation of mRNAs stability and half-life is one of the mechanisms used by cells to modulate protein synthesis. Amongst the several means used to regulate this process, microRNAs, very short RNA molecules, have been found to play a very important part. By binding to a specific part of the mRNA they trigger its degradation and thus indirectly reduce the production of specific proteins. In our experimental proposal we would thus like to study the sub-cellular localization and the role played by microRNAs in the elongation of neurites. This will be complemented by analyzing the localization and expression of Dicer, one of the key enzymes involved in the production of mature microRNAs. In particular we want to understand whether Dicer and microRNAs are relocated in elongating neurites and whether they play a causative role in this process. This will allow us to better understand how protein synthesis is regulated in growing neurons and by experimentally modulating Dicer's activity, whether any impairment of this mechanism can cause any damage to nerve cells. Furthermore, following our recent results on the regulation of the Brn-3b transcription factor mRNA by microRNAs, we would like to analyze the occurrence of a similar mechanism in retinal ganglion cells (RGC) induced to elongate neurites. Brn-3b is expressed during RGC development, and is essential for their survival as evidenced by the loss of up to 70% of RGC in mice devoid of Brn-3b. The analysis of Brn-3b post-transcriptional regulation in explanted RGC will be thus be a paradigm of the overall role of post-transcriptional regulation in RGC.