Regulation of gene expression in developing cortex by the transcription factor Pax6

我们对大脑在胚胎发生过程中如何控制自身发育的知识仍然相对贫乏。我们所知道的是，一组相对较小的基因，编码被称为转录因子的蛋白质，控制着细胞在胚胎中分裂、移动和成熟以形成大脑的过程。这些转录因子通常被认为是发育的主要调控因子。在人类或其他动物物种中，这些基因中的任何一种的活动异常都可能导致严重的缺陷，这表明它们对正常的大脑发育至关重要。转录因子通过控制其他基因的活性起作用；但是哪些基因，它们有多直接受到控制不同的基因在发育的不同时期是否受到控制？这些问题在很大程度上仍未得到解答，尽管我们有足够的知识认识到答案将是复杂的，因为每个转录因子控制着数百个基因。我们的研究旨在加深对其中一种转录因子Pax6的作用的理解。多年来，我们和其他人一直对分析这种特殊的转录因子感兴趣。众所周知，它的破坏会导致严重的人类疾病。在许多情况下，它是神经细胞发育的高水平调节剂，它可以编程干细胞成为神经细胞，这在未来基于干细胞开发新疗法的开发中可能是有用的。这项申请是及时的，因为我们最近已经制作并验证了新的转基因小鼠。这些新的转基因品系将使我们能够避免在以前的研究中遇到的一些主要问题，这些研究旨在发现和调查Pax6在发育过程中不同时期调节的基因。首先，它们将允许我们锁定那些表达Pax6基因的细胞，在这些细胞中，Pax6的直接作用将被发现；我们将能够忽略那些不再表达Pax6的细胞，因此，不再受Pax6的直接控制，即使它们可能仍然在早期和/或在其他细胞中间接受到Pax6作用的影响。其次，它们将使我们能够测试在不同发育年龄移除Pax6对其他基因活性的影响，从而提供在不同年龄哪些基因受Pax6控制的信息。第一部分工作的目的将是使用实验技术，然后使用计算（生物信息学）方法来生成和分析大量数据，以了解Pax6在不同年龄的丧失如何影响其他基因的活性。这方面预期的重要结果包括对不同发育年龄受Pax6调控的基因及其调控性质的强有力鉴定，例如哪些基因被Pax6抑制，哪些被激活，它们受Pax6缺失影响的速度有多快等。第二部分工作的目的是测试Pax6是否直接调节特定基因的特定假设，并将Pax6对基因活性的作用与其对发育中的脑细胞如何分裂、移动和成熟的影响联系起来。这将是朝着全面了解大脑发育中关键转录因子的分子功能这一具有挑战性的目标迈出的重要一步。这项工作很重要，不仅因为它将告诉我们Pax6的作用机制：更一般地说，这里的进展将为相关领域的研究提供信息，例如遗传疾病、癌症和干细胞生物学的分析。

Our knowledge of how the brain controls its own development during embryogenesis is still relatively poor. What we do know is that a relatively small group of genes, encoding proteins called transcription factors, control the processes by which cells divide, move and mature in the embryo to make the brain. These transcription factors are often thought of as master regulators of development. Severe defects can result from abnormalities in the activity of any one of these genes in humans or in other animal species, indicating their fundamental importance for normal brain development. Transcription factors work by controlling the activity of other genes; but which genes, how directly are they controlled and are different genes controlled at different times in development? These questions remain largely unanswered, although we know enough to realise that the answer will be complex, with many hundreds of genes falling under the control of each individual transcription factor.Our research is aimed at deepening understanding of the actions of one such transcription factor, Pax6. There has been sustained interest in the analysis of this particular transcription factor over many years by our group and others. Its disruption is well-known to cause severe human disease. It is a high-level regulator of the development of nerve cells in many contexts and it can programme stem cells to become nerve cells, which could be useful in the future development of new therapies based on the exploitation of stem cells. This application is timely since we have recently made and validated new transgenic mice for this work.These new transgenic lines will allow us to circumvent some of the major problems encountered in previous studies aimed at discovering and investigating the genes that Pax6 regulates at different times during development. First, they will allow us to home in on just those cells that express the Pax6 gene, in which Pax6's direct actions will be found; we shall be able to ignore those cells that no longer express Pax6 and are, therefore, no longer under its direct control even though they might still be influenced indirectly by Pax6's actions at earlier times and/ or in other cells. Second, they will allow us to test the effects of removing Pax6 at different developmental ages on the activity of other genes, providing information on which genes are controlled by Pax6 at different ages. The aim of the first part of the work will be to use experimental techniques followed by computational (bioinformatic) methods to generate and analyse large amounts of data on how loss of Pax6 at different ages affects the activity of other genes. The important outcome expected from this aspect includes robust identification of genes that are regulated by Pax6 at different developmental ages and the nature of the regulation, e.g. which genes are suppressed by Pax6, which are activated, how quickly are they affected by loss of Pax6, etc. The aims of the second part of the work will be to test specific hypotheses on whether Pax6 regulates particular genes directly or not and to correlate Pax6's actions on gene activity with its effects on how developing brain cells divide, move and mature. This will be a vital step towards the challenging goal of gaining a comprehensive understanding of the molecular functions of a key transcription factor in brain development. This work is important not only because it will tell us about Pax6's mechanisms of action: more generally, progress here will inform research in related fields, for example the analysis of genetic diseases, cancer and stem cell biology.