Makorin RING zinc-finger protein 1-dependent translational control of dendritic mRNAs in mammalian neurons

在神经细胞中，不同形式的学习和记忆需要新的蛋白质合成，这可以通过位于树突中的选择mRNA种类的翻译在突触处局部发生。这些新的蛋白质允许响应于突触激活的突触特异性重塑。然而，到目前为止，控制树突翻译的分子机制只知之甚少。在哺乳动物脑神经元中，Makorin RING锌指蛋白1（MKRN 1）是poly（A）结合蛋白的相互作用伴侣，与几种树突状定位的转录本相关。在大鼠大脑中，它特异性地积聚在激活的突触处。此外，功能测定显示，MKRN 1刺激神经元中报告mRNA的翻译起始。这一证据和其他证据表明，MKRN 1调节突触处树突状mRNA的活性依赖性翻译。我们的目标是表征神经元中MKRN 1介导的翻译控制机制。我们将特别处理以下问题：我们将确定不同的突触信号级联是否调节MKRN 1的活性。具体来说，我们想分析如何选择性刺激或抑制神经元中的单个信号成分影响MKRN 1介导的报告mRNA的翻译控制。二.翻译是一个高度复杂的过程，涉及到许多因素。我们打算表征在结构上和功能上将MKRN 1与翻译机制联系起来的分子。将对携带MKRN 1的细胞复合物进行生物化学纯化，并通过质谱法鉴定单个组分。随后，将使用小干扰（si）RNA技术评估单个蛋白质的生理意义。三. MKRN 1是一种真正的RNA结合蛋白。通过在蛋白质/RNA复合物的体内UV交联后进行免疫沉淀，我们想要鉴定啮齿动物脑中的MKRN 1 mRNA靶点。之后，我们将研究MKRN 1是否调节这些mRNA的翻译。我们希望我们的结果是相关的活性依赖性的翻译控制机制，更好地了解树突状mRNA的运作。这将为哺乳动物学习和记忆以及认知缺陷的分子过程提供新的见解。

In nerve cells, distinct forms of learning and memory require new protein synthesis, which can occur locally at synapses by translation of select mRNA species residing in dendrites. These new proteins permit synapse-specific remodelling in response to synaptic activation. However, to date the molecular mechanisms controlling dendritic translation are only sparsely understood. In mammalian brain neurons, makorin RING zinc-finger protein 1 (MKRN1), an interaction partner of poly(A)-binding protein, associates with several dendritically localized transcripts. In vivo in the rat brain, it specifically accumulates at activated synapses. Moreover, functional assays show that MKRN1 stimulates translation initiation of reporter mRNAs in neurons. This and additional evidence indicate that MKRN1 regulates activity-dependent translation of dendritic mRNAs at synapses. Our goal is to characterize the MKRN1-mediated translational control mechanisms in neurons. In particular, we will address the following issues: i. We will determine whether distinct synaptic signaling cascades modulate the activity of MKRN1. Specifically, we want to analyze how the selective stimulation or inhibition of individual signaling components in neurons influences MKRN1-mediated translational control of reporter mRNAs. ii. Translation is a highly complex process with numerous factors involved. We intend to characterize molecules that structurally and functionally link MKRN1 to the translation machinery. Cellular complexes harbouring MKRN1 will be biochemically purified and individual components will be identified by mass spectrometry. Subsequently, the physiological significance of individual proteins will be assessed using small interfering (si)RNA technology. iii. MKRN1 is a bona fide RNA-binding protein. By performing immunoprecipitations after in vivo UV-crosslinking of protein/RNA complexes we want to identify MKRN1 mRNA targets in rodent brain. Afterwards we will investigate if MKRN1 regulates translation of these mRNAs. We expect our results to be relevant for a better understanding of activity-dependent translational control mechanisms operating on dendritic mRNAs. This will provide new insights into the molecular processes underlying learning and memory as well as cognitive deficits in mammals.