Lombardi R, Riezman H
Biozentrum of the University of Basel, CH-4056 Basel, Switzerland.
J Biol Chem. 2001 Feb 23;276(8):6016-22. doi: 10.1074/jbc.M008735200. Epub 2000 Nov 28.
Mutations in RVS161 and RVS167, the two yeast amphiphysin homologs, cause very similar growth phenotypes, a depolarized actin cytoskeleton, and a defect in the internalization step of endocytosis. Rvs161p and Rvs167p have been shown to interact in the two-hybrid system, but their localization in the cell may be different thus raising the question whether the interaction is physiologically relevant. Here we demonstrate that the two proteins function together in vivo. We find that the steady state level of Rvs167p is strongly reduced in an rvs161Delta strain. Similarly, the level of Rvs161p is strongly reduced in an rvs167Delta strain. We demonstrate that these reduced protein levels at steady state are due to a decreased stability of either Rvs protein in the absence of the other protein. Furthermore, we find that the amount and ratio of Rvs161p and Rvs167p are critical parameters for receptor-mediated endocytosis. In addition, by using the two-hybrid system we show that the interaction of Rvs167p with actin is not abolished in an abp1Delta strain suggesting that Abp1p is not essential for this interaction.
酵母中两个发动蛋白同源物RVS161和RVS167的突变会导致非常相似的生长表型、肌动蛋白细胞骨架去极化以及内吞作用内化步骤中的缺陷。Rvs161p和Rvs167p已被证明在双杂交系统中相互作用，但它们在细胞中的定位可能不同，因此引发了这种相互作用在生理上是否相关的问题。在这里，我们证明这两种蛋白在体内共同发挥作用。我们发现，在rvs161Δ菌株中，Rvs167p的稳态水平大幅降低。同样，在rvs167Δ菌株中，Rvs161p的水平也大幅降低。我们证明，这些稳态下降低的蛋白水平是由于在缺乏另一种蛋白时，任一Rvs蛋白的稳定性降低所致。此外，我们发现Rvs161p和Rvs167p的量和比例是受体介导的内吞作用的关键参数。另外，通过使用双杂交系统，我们表明在abp1Δ菌株中，Rvs167p与肌动蛋白的相互作用并未消除，这表明Abp1p对于这种相互作用并非必不可少。
