Faculdade de Ciências Farmacêuticas de Ribeirão, Preto Universidade de São Paulo, São Paulo, Brazil.
BMC Complement Altern Med. 2012 Oct 24;12:194. doi: 10.1186/1472-6882-12-194.
Propolis is a natural product of plant resins collected by honeybees (Apis mellifera) from various plant sources. Our previous studies indicated that propolis sensitivity is dependent on the mitochondrial function and that vacuolar acidification and autophagy are important for yeast cell death caused by propolis. Here, we extended our understanding of propolis-mediated cell death in the yeast Saccharomyces cerevisiae by applying systems biology tools to analyze the transcriptional profiling of cells exposed to propolis.
We have used transcriptional profiling of S. cerevisiae exposed to propolis. We validated our findings by using real-time PCR of selected genes. Systems biology tools (physical protein-protein interaction [PPPI] network) were applied to analyse the propolis-induced transcriptional bevavior, aiming to identify which pathways are modulated by propolis in S. cerevisiae and potentially influencing cell death.
We were able to observe 1,339 genes modulated in at least one time point when compared to the reference time (propolis untreated samples) (t-test, p-value 0.01). Enrichment analysis performed by Gene Ontology (GO) Term finder tool showed enrichment for several biological categories among the genes up-regulated in the microarray hybridization such as transport and transmembrane transport and response to stress. Real-time RT-PCR analysis of selected genes showed by our microarray hybridization approach was capable of providing information about S. cerevisiae gene expression modulation with a considerably high level of confidence. Finally, a physical protein-protein (PPPI) network design and global topological analysis stressed the importance of these pathways in response of S. cerevisiae to propolis and were correlated with the transcriptional data obtained thorough the microarray analysis.
In summary, our data indicate that propolis is largely affecting several pathways in the eukaryotic cell. However, the most prominent pathways are related to oxidative stress, mitochondrial electron transport chain, vacuolar acidification, regulation of macroautophagy associated with protein target to vacuole, cellular response to starvation, and negative regulation of transcription from RNA polymerase II promoter. Our work emphasizes again the importance of S. cerevisiae as a model system to understand at molecular level the mechanism whereby propolis causes cell death in this organism at the concentration herein tested. Our study is the first one that investigates systematically by using functional genomics how propolis influences and modulates the mRNA abundance of an organism and may stimulate further work on the propolis-mediated cell death mechanisms in fungi.
蜂胶是由蜜蜂（Apis mellifera）从各种植物源中采集的植物树脂的天然产物。我们之前的研究表明，蜂胶敏感性取决于线粒体功能，液泡酸化和自噬对于蜂胶引起的酵母细胞死亡很重要。在这里，我们通过应用系统生物学工具来分析暴露于蜂胶的细胞的转录谱，从而扩展了我们对蜂胶介导的酵母 Saccharomyces cerevisiae 细胞死亡的理解。
我们已经使用了暴露于蜂胶的 S. cerevisiae 的转录谱。我们通过对选定基因的实时 PCR 验证了我们的发现。系统生物学工具（物理蛋白质-蛋白质相互作用[PPPI]网络）被应用于分析蜂胶诱导的转录行为，旨在确定哪些途径在 S. cerevisiae 中被蜂胶调节并可能影响细胞死亡。
与参比时间（未处理的蜂胶样品）相比，我们能够观察到至少一个时间点有 1339 个基因被调节（t 检验，p 值<0.01）。通过 Gene Ontology（GO）术语查找工具进行的富集分析显示，在微阵列杂交中上调的基因中，许多生物学类别都被富集，例如运输和跨膜运输以及应激反应。通过我们的微阵列杂交方法进行的选定基因的实时 RT-PCR 分析表明，它能够提供有关 S. cerevisiae 基因表达调节的信息，置信度相当高。最后，物理蛋白质-蛋白质（PPPI）网络设计和全局拓扑分析强调了这些途径在 S. cerevisiae 对蜂胶的反应中的重要性，并与通过微阵列分析获得的转录数据相关。
总之，我们的数据表明，蜂胶在真核细胞中广泛影响着几个途径。然而，最突出的途径与氧化应激、线粒体电子传递链、液泡酸化、与蛋白靶向液泡相关的巨自噬的调节、细胞对饥饿的反应以及 RNA 聚合酶 II 启动子的转录负调节有关。我们的工作再次强调了 S. cerevisiae 作为一个模型系统的重要性，可用于在分子水平上理解蜂胶在此浓度下引起该生物体细胞死亡的机制。我们的研究是第一个使用功能基因组学系统地研究蜂胶如何影响和调节生物体的 mRNA 丰度，并可能刺激进一步研究蜂胶介导的真菌细胞死亡机制的研究。
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