Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria, Australia.
PLoS One. 2013 Apr 18;8(4):e59763. doi: 10.1371/journal.pone.0059763. Print 2013.
Cyclic AMP Response Element-Binding Protein 1 (Creb1) is a transcription factor that mediates cyclic adenosine 3', 5'-monophosphate (cAMP) signalling in many tissues. Creb1(-/-) mice die at birth due to respiratory failure and previous genome-wide microarray analysis of E17.5 Creb1(-/-) fetal mouse lung identified important Creb1-regulated gene targets during lung development. The lipogenic enzymes stearoyl-CoA desaturase 1 (Scd1) and fatty acid synthase (Fasn) showed highly reduced gene expression in Creb1(-/-) lungs. We therefore hypothesized that Creb1 plays a crucial role in the transcriptional regulation of genes involved in pulmonary lipid biosynthetic pathways during lung development. In this study we confirmed that Scd1 and Fasn mRNA levels were down regulated in the E17.5 Creb1(-/-) mouse lung while the lipogenic-associated transcription factors SrebpF1, C/ebpα and Pparγ were increased. In vivo studies using germline (Creb1(-/-) ) and lung epithelial-specific (Creb1(EpiΔ/Δ) ) Creb1 knockout mice showed strongly reduced Scd1, but not Fasn gene expression and protein levels in lung epithelial cells. In vitro studies using mouse MLE-15 epithelial cells showed that forskolin-mediated activation of Creb1 increased both Scd1 gene expression and protein synthesis. Additionally, MLE15 cells transfected with a dominant-negative ACreb vector blocked forskolin-mediated stimulation of Scd1 gene expression. Lipid profiling in MLE15 cells showed that dominant-negative ACreb suppressed forskolin-induced desaturation of ether linked lipids to produce plasmalogens, as well as levels of phosphatidylethanolamine, ceramide and lysophosphatidylcholine. Taken together these results demonstrate that Creb1 is essential for the induction and maintenance of Scd1 in developing fetal mouse lung epithelial cells.
环磷酸腺苷反应元件结合蛋白 1(Creb1)是一种转录因子,可介导许多组织中的环腺苷酸 3',5'-单磷酸(cAMP)信号传导。由于呼吸衰竭,Creb1(-/-)小鼠在出生时死亡,此前对 E17.5 Creb1(-/-)胎鼠肺的全基因组微阵列分析确定了肺发育过程中重要的 Creb1 调节基因靶标。脂肪生成酶硬脂酰辅酶 A 去饱和酶 1(Scd1)和脂肪酸合酶(Fasn)在 Creb1(-/-)肺中的基因表达明显降低。因此,我们假设 Creb1 在肺发育过程中参与肺脂生物合成途径相关基因的转录调控中发挥关键作用。在这项研究中,我们证实 Scd1 和 Fasn 的 mRNA 水平在 E17.5 Creb1(-/-)小鼠肺中下调,而脂肪生成相关转录因子 SrebpF1、C/ebpα 和 Pparγ 增加。体内研究使用生殖系(Creb1(-/-))和肺上皮特异性(Creb1(EpiΔ/Δ))Creb1 敲除小鼠表明,肺上皮细胞中 Scd1 但不是 Fasn 的基因表达和蛋白水平明显降低。体外研究使用小鼠 MLE-15 上皮细胞表明, forskolin 介导的 Creb1 激活增加了 Scd1 的基因表达和蛋白合成。此外,用显性失活的 ACreb 载体转染的 MLE15 细胞阻断了 forskolin 介导的 Scd1 基因表达的刺激。MLE15 细胞中的脂质分析表明,显性失活的 ACreb 抑制了 forskolin 诱导的醚连接脂质的去饱和以产生磷脂酰乙醇胺、神经酰胺和溶血磷脂酰胆碱。总之,这些结果表明 Creb1 对于诱导和维持发育中的胎鼠肺上皮细胞中的 Scd1 是必不可少的。