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A genetically encoded biosensor for visualising hypoxia responses?in vivo

机译：用于体内低氧反应可视化的遗传编码生物传感器

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摘要

Cells experience different oxygen concentrations depending on location, organismal developmental stage, and physiological or pathological conditions. Responses to reduced oxygen levels (hypoxia) rely on the conserved hypoxia-inducible factor 1 (HIF-1). Understanding the developmental and tissue-specific responses to changing oxygen levels has been limited by the lack of adequate tools for monitoring HIF-1?in vivo.?To visualise and analyse HIF-1 dynamics in?Drosophila, we used a hypoxia biosensor consisting of GFP fused to the oxygen-dependent degradation domain (ODD) of the HIF-1 homologue Sima. GFP-ODD responds to changing oxygen levels and to genetic manipulations of the hypoxia pathway, reflecting oxygen-dependent regulation of HIF-1 at the single-cell level. Ratiometric imaging of GFP-ODD and a red-fluorescent reference protein reveals tissue-specific differences in the cellular hypoxic status at ambient normoxia. Strikingly, cells in the larval brain show distinct hypoxic states that correlate with the distribution and relative densities of respiratory tubes. We present a set of genetic and image analysis tools that enable new approaches to map hypoxic microenvironments, to probe effects of perturbations on hypoxic signalling, and to identify new regulators of the hypoxia response.
机译：细胞会经历不同的氧气浓度，具体取决于位置，生物发育阶段以及生理或病理状况。对降低的氧气水平（缺氧）的反应依赖于保守的缺氧诱导因子1（HIF-1）。由于缺乏足够的体内监测HIF-1的工具，对了解氧水平变化的发育和组织特异性反应受到了限制。为了可视化和分析果蝇中HIF-1的动态，我们使用了一个由GFP融合到HIF-1同系物Sima的氧依赖性降解域（ODD）。 GFP-ODD响应不断变化的氧气水平和缺氧途径的遗传操作，反映了单细胞水平上HIF-1的氧气依赖性调节。 GFP-ODD和红​​色荧光参考蛋白的比例成像显示在环境常氧下细胞缺氧状态的组织特异性差异。令人惊讶的是，幼虫大脑中的细胞表现出明显的低氧状态，与呼吸管的分布和相对密度有关。我们提供了一套遗传和图像分析工具，这些工具使人们能够采用新方法来绘制低氧微环境，探测微扰对低氧信号的影响，并确定低氧反应的新调节剂。

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