Towards a temperature-sensitive proteome: developing a Drosophila-friendly degron

在这里，我们建议开发一种新技术，使生物学家能够简单地通过改变温度来开启和关闭基因的活动，只需在较高的培养温度下针对他们的产品进行降解。这项技术基于成熟的生物机制，细胞使用该机制来实现对不需要的蛋白质的有控制的去除。这种受调控的降解是通过将降解信号附着到靶标上来调节的。在酵母中发现的一些蛋白质(降解子)，当它们的结构在更高的温度下被破坏时，可以诱导这些信号附着到自己身上，从而导致它们自身和它们所属的任何融合蛋白的降解。我们和其他人已经成功地证明了该系统可以用于更复杂的模式生物--果蝇。然而，果蝇或其他冷血生物不能很好地耐受酵母中诱导目标降解的温度。因此，我们建议分离酵母降解信号的新变体，这些变体在果蝇和其他生物学家广泛使用的模式生物的正常温度范围内工作得更好，并在培养的果蝇细胞和基因操纵的果蝇中进行测试。

Here we propose to develop a new technology that enables biologists to turn activity of genes on and off simply by changing the temperature, by targeting their products for degradation only at higher culture temperatures. This technology is based on well-established biological mechanisms employed by the cell to achieve regulated removal of unwanted proteins. This regulated degradation is mediated by the attachment of degradation signals to the target. Some proteins (degrons), identified in yeast, can induce attachment of these signals to themselves when their structure is disrupted at higher temperatures, thus inducing degradation of themselves and any fusion proteins that they are part of. We and others have successfully shown proof-of-principle that the system can be used in a more complex model organism, Drosophila (fruitfly). However, the temperatures at which degradation of the target is induced in yeast are not well-tolerated by Drosophila, or by other 'cold-blooded' organisms. Therefore, we propose to isolate novel variants of the yeast degron signal, that work better in the normal temperature ranges of Drosophila and of other model organisms widely used by biologists, and to test these in cultured Drosophila cells and in genetically manipulated flies.