Department of Pediatrics, University of California at San Diego, La Jolla, CA 92093, USA.
Proc Natl Acad Sci U S A. 2011 Feb 8;108(6):2349-54. doi: 10.1073/pnas.1010643108. Epub 2011 Jan 24.
Through long-term laboratory selection (over 200 generations), we have generated Drosophila melanogaster populations that tolerate severe, normally lethal, levels of hypoxia. Because of initial experiments suspecting genetic mechanisms underlying this adaptation, we compared the genomes of the hypoxia-selected flies with those of controls using deep resequencing. By applying unique computing and analytical methods we identified a number of DNA regions under selection, mostly on the X chromosome. Several of the hypoxia-selected regions contained genes encoding or regulating the Notch pathway. In addition, previous expression profiling revealed an activation of the Notch pathway in the hypoxia-selected flies. We confirmed the contribution of Notch activation to hypoxia tolerance using a specific γ-secretase inhibitor, N-[N-(3,5-Difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester (DAPT), which significantly reduced adult survival and life span in the hypoxia-selected flies. We also demonstrated that flies with loss-of-function Notch mutations or RNAi-mediated Notch knockdown had a significant reduction in hypoxia tolerance, but those with a gain-of-function had a dramatic opposite effect. Using the UAS-Gal4 system, we also showed that specific overexpression of the Notch intracellular domain in glial cells was critical for conferring hypoxia tolerance. Unique analytical tools and genetic and bioinformatic strategies allowed us to discover that Notch activation plays a major role in this hypoxia tolerance in Drosophila melanogaster.
通过长期的实验室选择（超过 200 代），我们生成了能够耐受严重、通常致命的缺氧水平的黑腹果蝇种群。由于最初的实验怀疑这种适应的遗传机制，我们使用深度重测序将缺氧选择的果蝇基因组与对照进行了比较。通过应用独特的计算和分析方法，我们鉴定了一些受选择的 DNA 区域，主要在 X 染色体上。缺氧选择的几个区域包含编码或调节 Notch 途径的基因。此外，先前的表达谱分析显示 Notch 途径在缺氧选择的果蝇中被激活。我们使用特定的γ-分泌酶抑制剂 N-[N-(3,5-二氟苯乙酰基)-L-丙氨酰]-S-苯甘氨酸叔丁酯（DAPT）证实了 Notch 激活对缺氧耐受性的贡献，该抑制剂显著降低了缺氧选择的果蝇的成虫存活率和寿命。我们还证明，具有 Notch 功能丧失突变或 RNAi 介导的 Notch 敲低的果蝇对缺氧耐受性有显著降低，但具有功能获得性的果蝇则有相反的显著效果。我们还使用 UAS-Gal4 系统表明，胶质细胞中 Notch 细胞内结构域的特异性过表达对于赋予果蝇缺氧耐受性至关重要。独特的分析工具以及遗传和生物信息学策略使我们能够发现 Notch 激活在黑腹果蝇的这种缺氧耐受性中起着重要作用。
