Prolonged exposure to elevated free fatty acids induces β-cell failure (lipotoxicity) and contributes to the pathogenesis of type 2 diabetes. In vitro exposure of β-cells to the saturated free fatty acid palmitate is a valuable model of lipotoxicity, reproducing features of β-cell failure observed in type 2 diabetes. In order to map the β-cell response to lipotoxicity, we combined RNA-sequencing of palmitate-treated human islets with iTRAQ proteomics of insulin-secreting INS-1E cells following a time course exposure to palmitate. Crossing transcriptome and proteome of palmitate-treated β-cells revealed 85 upregulated and 122 downregulated genes at both transcript and protein level. Pathway analysis identified lipid metabolism, oxidative stress, amino-acid metabolism and cell cycle pathways among the most enriched palmitate-modified pathways. Palmitate induced gene expression changes compatible with increased free fatty acid mitochondrial import and β-oxidation, decreased lipogenesis and modified cholesterol transport. Palmitate modified genes regulating endoplasmic reticulum (ER) function, ER-to-Golgi transport and ER stress pathways. Furthermore, palmitate modulated cAMP/protein kinase A (PKA) signaling, inhibiting expression of PKA anchoring proteins and downregulating the GLP-1 receptor. SLC7 family amino-acid transporters were upregulated in response to palmitate but this induction did not contribute to β-cell demise. To unravel critical mediators of lipotoxicity upstream of the palmitate-modified genes, we identified overrepresented transcription factor binding sites and performed network inference analysis. These identified LXR, PPARα, FOXO1 and BACH1 as key transcription factors orchestrating the metabolic and oxidative stress responses to palmitate. This is the first study to combine transcriptomic and sensitive time course proteomic profiling of palmitate-exposed β-cells. Our results provide comprehensive insight into gene and protein expression changes, corroborating and expanding beyond previous findings. The identification of critical drivers and pathways of the β-cell lipotoxic response points to novel therapeutic targets for type 2 diabetes.

中文翻译：

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胰腺β细胞对棕榈酸酯反应的转录组和蛋白质组分析相结合，揭示了β细胞脂毒性的关键途径。

长时间暴露于升高的游离脂肪酸中会诱发β细胞衰竭（脂毒性），并导致2型糖尿病的发病。β细胞在饱和游离脂肪酸棕榈酸酯中的体外暴露是脂毒性的宝贵模型，可再现在2型糖尿病中观察到的β细胞衰竭特征。为了绘制β细胞对脂毒性的反应，我们将棕榈酸酯处理的人胰岛的RNA测序与胰岛素分泌的INS-1E细胞的iTRAQ蛋白质组学结合起来，然后逐渐暴露于棕榈酸酯。棕榈酸酯处理过的β细胞的转录组和蛋白质组交叉显示，在转录和蛋白质水平上共有85个上调的基因和122个下调的基因。途径分析在最丰富的棕榈酸酯修饰途径中鉴定出脂质代谢，氧化应激，氨基酸代谢和细胞周期途径。棕榈酸酯诱导的基因表达变化与增加的游离脂肪酸线粒体输入和β-氧化，降低的脂肪生成和修饰的胆固醇转运相兼容。棕榈酸酯修饰的基因调节内质网（ER）功能，ER向高尔基体的转运和ER应激途径。此外，棕榈酸酯调节cAMP /蛋白激酶A（PKA）信号传导，抑制PKA锚定蛋白的表达并下调GLP-1受体。SLC7家族氨基酸转运蛋白响应棕榈酸酯被上调，但这种诱导并没有促进β细胞的死亡。为了揭示棕榈酸酯修饰基因上游脂毒性的关键介体，我们鉴定了过度表达的转录因子结合位点并进行了网络推断分析。这些确定了LXR，PPARα，FOXO1和BACH1是关键的转录因子，负责协调对棕榈酸酯的代谢和氧化应激反应。这是首次将棕榈酸酯暴露的β细胞的转录组学和敏感的时程蛋白质组学分析相结合的研究。我们的结果提供了对基因和蛋白质表达变化的全面洞察力，证实并扩大了以前的发现。β细胞脂毒性反应的关键驱动器和途径的鉴定指向2型糖尿病的新型治疗靶标。证实并扩大了以前的发现。β细胞脂毒性反应的关键驱动器和途径的鉴定指向2型糖尿病的新型治疗靶标。证实并扩大了以前的发现。β细胞脂毒性反应的关键驱动器和途径的鉴定指向2型糖尿病的新型治疗靶标。