脑衰老过程中长链非编码RNA对学习记忆相关基因的调控功能及机制

Learning and memory impairment is one of the early phenotypes in brain aging, but the underlying mechanism is not clear. Our preliminary data showed that, in hippocampus which is closely related to learning and memory, hippocampus-specifically expressed lncRNAs and mRNAs were almost all significantly up-regulated in the natural aged mice. These specific mRNAs are closely related to learning and memory as well as senescence, and the expression of these specific lncRNAs is also closed related to the expression of the hippocampus-specific mRNAs. Combining with these data and the synergistic effect of lncRNAs and chromatin conformation on gene regulation, we hypothesize that lncRNAs are involved in the formation of age-specific chromatin three-dimensional conformation to regulate learning and memory related genes during brain aging process, which may be one of the key molecular bases for the impairment of learning and memory during brain aging. The project will use technologies such as RNA-Seq and 3D-DSL to detect the changes of lncRNAs’ expression and chromatin conformation, as well as the three-dimensional gene regulation networks formed based on the interactions between lncRNAs and chromatin conformation in learning and memory-dependent hippocampus from young and old mice. From which we will identify how they regulate learning and memory related genes, discover the causal relationship between this gene regulation and learning and memory impairment in mice brain aging process, and explore the functional mechanisms. This study is important to reveal the molecular mechanisms of brain aging, and may lead to new ideas for slowing down the aging process and for the prevention of brain aging related diseases.

学习记忆功能减退是脑衰老早期指标之一，但机制不明。我们前期发现，在与学习记忆密切相关的脑海马区特异表达的lncRNA和mRNA几乎都在自然衰老的老年小鼠中显著上调。这些海马特异的mRNA与学习记忆及衰老密切相关，并与海马特异的lncRNA存在共表达关系。结合lncRNA与染色质构象对基因调控的协同作用，我们假设：lncRNA在脑衰老过程中参与形成年龄特异的染色质三维构象来共同调控学习记忆相关基因，这可能是脑衰老过程中学习记忆功能减退的重要分子基础之一。本项目将结合RNA-Seq和3D-DSL等技术检测海马区神经组织在小鼠青年和老年时lncRNA表达及染色质构象的变化、以及两者间形成的三维基因调控网络对脑衰老过程中学习记忆相关基因的表达调控，及其与小鼠学习记忆功能减退之间的因果关系，并探究其作用机制。本研究对揭示脑衰老分子机制有重要意义，并可能为延缓脑衰老以及防治脑衰老相关疾病提供新思路。

本项目通过对青年和自然衰老小鼠全脑的RNA 进行 RNA-Seq 测序和 lncRNA Microarray 芯片检测，鉴定出了全脑及海马、小脑、嗅球等脑区中差异表达的lncRNAs，构建了衰老过程中与学习密切相关的lncRNA表达谱。在此基础上对若干显著差异表达的lncRNAs构建敲除小鼠模型，开展一系列的分子机制研究。重要研究成果有以下几个方面：(1) lncRNA Synage调控小鼠的突触与运动功能。在小鼠体内敲除lncRNA Synage后，发现严重影响小鼠衰老关键蛋白、小鼠的神经元发育、突触的形成与突触功能的发挥。在发育与衰老过程中会明显导致小鼠运动及学习与记忆功能严重退化。(2) Malat1在年老小鼠脑尤其是嗅球区域表达量显著上调。Malat1在基因组上的一个结合位点是Cdkn2a基因，而此基因在衰老中起重要的作用。此外，我们还利用构建了Malat1的敲除鼠（敲除的片段是从未有报道的Malat1基因3’端的绝大部分，5.7kb）。结果显示Malat1敲除鼠加速机体老化并提高死亡率，其具体作用机制正在进一步研究中。(3) lncRNA AK039238在年轻和老年小鼠的不同脑区存在差异表达。它定位在多种脑细胞的细胞核内，且结合于染色体上。对其进行敲降后，我们发现KCNAB1 和LMNB1等基因的表达量显著下降。其中LMNB1编码的核纤层蛋白与细胞核的结构密切相关。LncRNA AK039238在基因组上结合在KCNAB1基因上，提示其可能对KCNAB1基因起直接的调控作用。LncRNA AK039238与nucleolin存在共定位并且可能存在相互结合。具体功能及分子机制正在研究中。本研究对揭示脑衰老过程中lncRNA对学习记忆相关基因的调控功能和分子机制有重要意义，并可能为延缓脑衰老以及防治脑衰老相关疾病提供新思路。

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