Control of Yeast Life Span

描述(申请人提供)：人类衰老的标志之一是线粒体功能随着年龄的增长而下降。模型系统已经证明了一条信号通路，该通路将线粒体的功能状态传达到细胞核，以协调转录反应。使用一种独特的监测核转录变化的报告系统，我们发现了一条新的线粒体-核信号通路，揭示了酿酒酵母中由ATP2基因编码的F1F0-ATP合成酶复合体b亚基的重要性。我们发现缺乏ATP2基因的细胞寿命很短，这是令人惊讶的，因为在秀丽线虫中的RNAi实验表明，减少其他F1F0-ATP合成酶亚基的表达可以延长寿命。我们取得了令人兴奋的发现，ATP2表达的适度减少或增加延长了酿酒酵母的寿命。我们假设，b亚单位水平的错误调节导致了ATP水平的下降。由于F1F0-ATP合成酶从细菌到人类都是保守的，了解它如何延长酵母的寿命以及这种影响是否在进化上保守将对我们对衰老的理解产生广泛的影响。为了确定更多的长寿途径，我们在裂殖酵母中开发了一种新的老化试验，该方法概括了通过限制热量和长寿命细胞的抗应激而延长寿命的进化保守属性。这种分析方法允许从随机突变体群体中直接和无偏见地选择长寿突变体。我们已经构建了一大批S.pombe DNA插入突变体，这将使我们能够分离出长寿突变体并快速鉴定受影响的基因。我们的长期目标是了解控制衰老的遗传途径，我们将通过描述模型系统中进化保守的途径来实现这一点。我们的具体目标将回答以下问题：1.F1F0-ATP合成酶b亚基中突变的寿命延长是否与产生ATP的能力降低有关？我们将确定长寿突变体是否改变了ATP水平和线粒体功能，并测试是否必须将b亚基定位于线粒体以延长寿命。2.错误调节F1F0-ATP合成酶b亚基的寿命延长效应在进化上是保守的吗？我们将测试错误调节b亚基是否会延长果蝇或S.pombe的寿命。3.在我们独特的S.pombe老化试验中，对长寿突变体的无偏见分离是否揭示了延长时间跨度的新途径？我们将通过限制卡路里来表征S.pombe中延长寿命的机制，并使用这个系统来鉴定新的延长寿命的突变。我们的结果将为进化上保守的途径提供新的见解，这些途径将影响对人类衰老生物学的理解。

DESCRIPTION (provided by applicant): One of the hallmarks of human aging is the decline in mitochondrial function with advancing years. Model systems have demonstrated a signaling pathway that communicates the functional state of the mitochondria to the nucleus to coordinate a transcriptional response. Using a unique reporter system that monitors changes in nuclear transcription, we uncovered a new mitochondrial-nuclear signaling pathway that revealed the importance of the b-subunit of the F1F0-ATP synthase complex, encoded in Saccharomyces cerevisiae by the ATP2 gene. We found that cells lacking the ATP2 gene had a short lifespan, which was surprising since RNAi experiments in Caenorhabditis elegans indicated that reducing expression of other F1F0- ATP synthase subunits extends lifespan. We made the exciting discovery that either a modest decrease or increase in ATP2 expression extends S. cerevisiae lifespan. We hypothesize that misregulation of b-subunit levels causes a decrease in ATP levels. As the F1F0-ATP synthase is conserved from bacteria to humans, understanding how it can extend yeast lifespan and if this effect is evolutionarily conserved will have wide impact on our understanding of aging. To identify additional longevity pathways, we developed a new aging assay in Schizosaccharomyces pombe that recapitulates the evolutionarily conserved properties of lifespan extension by caloric restriction and stress resistance of long-lived cells. This assay allows the direct and unbiased selection of long-lived mutants from populations of random mutants. We have constructed a large bank of S. pombe DNA insertion mutants that will allow an isolation of long-lived mutants and rapid identification of the affected genes. Our long-term goals are to understand the genetic pathways that control aging, which we will approach by characterizing evolutionarily conserved pathways in model systems. Our specific aims will answer the following questions: 1. Is the lifespan extension of mutants misregulated in the F1F0-ATP synthase b-subunit related to a reduction in the ability to produce ATP? We will determine if the long-lived mutants have altered ATP levels and mitochondrial functions, and test whether the b-subunit must be localized to mitochondria to extend lifespan. 2. Are the lifespan extending effects of misregulating the b-subunit of the F1F0-ATP synthase evolutionarily conserved? We will test if misregulating the b-subunit increases Drosophila or S. pombe lifespan. 3. Does the unbiased isolation of long-lived mutants in our unique S. pombe aging assay reveal new pathways to extend chronological lifespan? We will characterize the mechanism of lifespan extension in S. pombe by caloric restriction and use this system to identify new lifespan extending mutations. Our results will provide new insights into evolutionarily conserved pathways that will impact the understanding of the biology of human aging.