Delayed Degradation and Impaired Dendritic Delivery of Intron-Lacking EGFP-Arc/Arg3.1 mRNA in EGFP-Arc Transgenic Mice.
Delayed Degradation and Impaired Dendritic Delivery of Intron-Lacking EGFP-Arc/Arg3.1 mRNA in EGFP-Arc Transgenic Mice.
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DOI：
10.3389/fnmol.2017.00435
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发表时间：
2017
影响因子：
4.8
通讯作者：
Bito H
中科院分区：
文献类型：
--
作者：
Steward O;Matsudaira Yee K;Farris S;Pirbhoy PS;Worley P;Okamura K;Okuno H;Bito H
Arc是一种独特的立即早期基因（IEG），其表达在学习过程中突触被修饰时被诱导。新合成的Arc mRNA在整个树突中快速运输，并定位在最近激活的突触附近。Arc mRNA水平受快速降解的调节，而快速降解在突触依赖性过程中被突触活动加速。一种可能的机制是无义介导的mRNA衰变（NMD），这取决于3′UTR中剪接点的存在。在这里，我们使用表达EGFP-Arc的转基因小鼠来验证这一假设。由于转基因是由Arc cDNA构建的，因此它在3′UTR中缺乏内源性Arc基因中存在的内含子结构。NMD依赖于终止密码子下游外显子连接复合物（EJC）蛋白的存在，因此EGFP-Arc mRNA不应经历NMD。在转录抑制剂放线菌素-D存在下评估Arc mRNA下降证实了EGFP-Arc mRNA的延迟降解。EGFP-Arc mRNA和蛋白在基础和活化条件下在转基因小鼠中以高得多的水平表达，但EGFP-Arc mRNA不能有效地进入树突。在生理测定中，放线菌酮（CHX）注入后，诱导的弧癫痫发作，有增加的内源性Arc mRNA水平与癫痫依赖性Arc mRNA衰减一致，但这是没有看到与EGFP-Arc mRNA。综上所述，我们的结果表明：（1）Arc mRNA的降解是通过具有NMD特征的机制发生的;（2）诱导后新合成的Arc mRNA的快速树突状递送可能部分依赖于3′UTR的预先剪接。
Arc is a unique immediate early gene (IEG) whose expression is induced as synapses are modified during learning. Newly-synthesized Arc mRNA is rapidly transported throughout dendrites and localizes near recently activated synapses. Arc mRNA levels are regulated by rapid degradation, which is accelerated by synaptic activity in a translation-dependent process. One possible mechanism is nonsense-mediated mRNA decay (NMD), which depends on the presence of a splice junction in the 3′UTR. Here, we test this hypothesis using transgenic mice that express EGFP-Arc. Because the transgene was constructed from Arc cDNA, it lacks intron structures in the 3′UTR that are present in the endogenous Arc gene. NMD depends on the presence of proteins of the exon junction complex (EJC) downstream of a stop codon, so EGFP-Arc mRNA should not undergo NMD. Assessment of Arc mRNA rundown in the presence of the transcription inhibitor actinomycin-D confirmed delayed degradation of EGFP-Arc mRNA. EGFP-Arc mRNA and protein are expressed at much higher levels in transgenic mice under basal and activated conditions but EGFP-Arc mRNA does not enter dendrites efficiently. In a physiological assay in which cycloheximide (CHX) was infused after induction of Arc by seizures, there were increases in endogenous Arc mRNA levels consistent with translation-dependent Arc mRNA decay but this was not seen with EGFP-Arc mRNA. Taken together, our results indicate: (1) Arc mRNA degradation occurs via a mechanism with characteristics of NMD; (2) rapid dendritic delivery of newly synthesized Arc mRNA after induction may depend in part on prior splicing of the 3′UTR.
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影响因子：
16.2
作者：
Steward, O;Worley, PF
通讯作者：
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