Louvain Institute of Biomolecular Science and Technology, Université catholique de Louvain, B-1348, Louvain-la-Neuve, Belgium.
Institute of Physiology, Czech Academy of Sciences, CZ-14220, Prague, Czech Republic.
Sci Rep. 2020 Feb 5;10(1):1881. doi: 10.1038/s41598-020-58795-w.
The gradual acidification of the secretory pathway is conserved and extremely important for eukaryotic cells, but until now there was no pH sensor available to monitor the pH of the early Golgi apparatus in Saccharomyces cerevisiae. Therefore, we developed a pHluorin-based sensor for in vivo measurements in the lumen of the Golgi. By using this new tool we show that the cis- and medial-Golgi pH is equal to 6.6-6.7 in wild type cells during exponential phase. As expected, V-ATPase inactivation results in a near neutral Golgi pH. We also uncover that surprisingly Vph1p isoform of the V-ATPase is prevalent to Stv1p for Golgi acidification. Additionally, we observe that during changes of the cytosolic pH, the Golgi pH is kept relatively stable, mainly thanks to the V-ATPase. Eventually, this new probe will allow to better understand the mechanisms involved in the acidification and the pH control within the secretory pathway.
分泌途径的逐渐酸化在真核细胞中是保守且极其重要的，但直到现在，还没有可用的 pH 传感器来监测酿酒酵母中高尔基体早期的 pH 值。因此，我们开发了一种基于 pHluorin 的传感器，用于高尔基体腔室的体内测量。通过使用这个新工具，我们表明在指数生长期的野生型细胞中，顺面和中间高尔基体的 pH 值等于 6.6-6.7。正如预期的那样，V-ATPase 的失活导致高尔基体 pH 值接近中性。我们还发现，令人惊讶的是，V-ATPase 的 Vph1p 同工型比 Stv1p 更有利于高尔基体酸化。此外，我们观察到在细胞质 pH 值变化时，高尔基体 pH 值相对稳定，这主要归功于 V-ATPase。最终，这个新的探针将有助于更好地理解分泌途径中酸化和 pH 值控制的相关机制。
