Indirect Regulation of Presenilins in CREB-mediated Transcription
Indirect Regulation of Presenilins in CREB-mediated Transcription
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DOI：
10.1074/jbc.m809168200
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发表时间：
2009-05-15
影响因子：
4.8
通讯作者：
Shen, Jie
中科院分区：
文献类型：
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作者：
Watanabe, Hirotaka;Smith, Miriam J.;Shen, Jie
早老素是突触功能、记忆形成和神经元存活所必需的。以前，我们报道了cAMP反应元件结合蛋白（CREB）靶基因的表达减少早老素（PS）条件性双基因敲除（CDKO）小鼠的大脑皮层。为了确定CREB靶基因在这些突变小鼠中的表达减少是否是由于早老素的损失直接或继发于受损的神经元活性，我们建立了一个敏感的荧光素酶报告系统，以评估培养细胞中的直接转录调控。我们首先使用永生化的PS缺陷小鼠胚胎成纤维细胞（MEFs），发现CREB介导的转录和Notch介导的HES 1转录均降低。然而，泛素-C启动子介导的转录也被降低，并且在这三种报告基因中，外源PS1的转染仅能拯救Notch介导的HES 1转录。进一步的北方分析揭示了Creb，泛素-C和其他管家基因在PS缺陷的MEFs中的转录改变，表明这些细胞中的转录失调。然后，我们使用的Cre/loxP系统开发一个出生后PS缺陷的皮层神经元培养。令人惊讶的是，在这些无PS的神经元中，CREB介导的转录没有显著降低，总CREB蛋白和磷酸化CREB蛋白的水平也没有变化。Notch介导的HES 1转录显著减少，并且这种减少可以通过外源PS1来挽救。总之，我们的研究结果表明，CREB介导的转录是由PS在成人大脑皮层间接调节，和衰减的CREB靶基因的表达PS cDKO小鼠可能是由于这些突变体的大脑神经元活动减少。
Presenilins are essential for synaptic function, memory formation, and neuronal survival. Previously, we reported that expression of cAMP response element-binding protein (CREB) target genes is reduced in the cerebral cortex of presenilin (PS) conditional double knock-out (cDKO) mice. To determine whether the reduced expression of the CREB target genes in these mutant mice is due to loss of presenilin directly or secondary to the impaired neuronal activity, we established a sensitive luciferase reporter system to assess direct transcriptional regulation in cultured cells. We first used immortalized PS-deficient mouse embryonic fibroblasts (MEFs), and found that both CREB-mediated transcription and Notch-mediated HES1 transcription are decreased. However, the ubiquitin-C promoter-mediated transcription is also reduced, and among these three reporters, transfection of exogenous PS1 can rescue only the Notch-mediated HES1 transcription. Further Northern analysis revealed transcriptional alterations of Creb, ubiquitin-C, and other housekeeping genes in PS-deficient MEFs, indicating transcriptional dysregulation in these cells. We then used the Cre/loxP system to develop a postnatal PS-deficient cortical neuronal culture. Surprisingly, in these PS-null neurons, CREB-mediated transcription is not significantly decreased, and levels of total and phosphorylated CREB proteins are unchanged as well. Notch-mediated HES1 transcription is markedly reduced, and this reduction can be rescued by exogenous PS1. Together, our findings suggest that CREB-mediated transcription is regulated indirectly by PS in the adult cerebral cortex, and that attenuation of CREB target gene expression in PS cDKO mice is likely due to reduced neuronal activity in these mutant brains.