![]() Targeting of an improved pHluorin-based probe to the Golgi apparatus of S. It could also give guidelines to design other probes very specifically targeted to the different steps of the secretory pathway. In the future, this new probe will allow to better understand how the acidification occurs and is controlled in the Golgi apparatus. With this tool, we reveal that Vph1p plays a prevalent role compared to Stv1p for the sustenance of an acidic Golgi lumen and that the Golgi pH is actively kept acidic when the cytosolic pH fluctuates, mainly due to the V-ATPase. For this reason, we engineered a pH probe for the Golgi lumen. However, there is no sensor suitable to measure precisely the early Golgi pH. Recently, another sensor has been developed for pH measurements within the ER 20. cerevisiae, one probe is already available to measure the pH of the trans-Golgi network/endosomes lumen 17, 18 and the chemical probe BCECF is commonly used to measure the vacuolar pH 19. 16), it is essential to possess appropriate tools to accurately measure this parameter in vivo. Given the importance of pH homeostasis within the cell and the secretory pathway (reviewed in Casey et al. When these acidification mechanisms are not perfectly functional at the Golgi level, it may lead to various diseases such as congenital disorders of glycosylation, Cutis laxa or non-syndromic intellectual disability 12, 13, 14, 15. The vacuolar H +-ATPase (V-ATPase) is the main pump responsible for the acidification of the secretory pathway and the electrochemical balance is controlled by a Golgi pH regulator which is an anion channel 10, probably in collaboration with a still unidentified proton leak channel 11. While at the plasma membranes the nature of this electrochemical gradient differs between the different kingdoms of life, the pH gradient is the main electrochemical gradient used in organelles of all eukaryotes by secondary transporters. Furthermore, the pH gradient across biological membranes serves as the driving force for many secondary transporters. It is particularly well described for several plasma membrane receptors which bind to their target at the plasma membrane and dissociate once the pH drops in endosomes 6, for the delivery of lysosomal proteases to their destination thanks to the mannose-6-phosphate receptor 7, or for the retrieval of ER-resident proteins that are recycled from the Golgi to the ER thanks to the KDEL receptor 8, 9. Many receptors have pH-dependent affinity for their ligand. In addition, the pH also controls the trafficking and localization of these enzymes within the secretory pathway 5. Similarly, some glycosidases and glycosyltransferases become active once they face the appropriate pH, in a specific compartment of the secretory pathway 4. For example, it is the case for many proteases which are turned “ON” when they reach the acidic vacuole 2, 3. This gradual acidification is crucial to trigger the activation of some enzymes involved in post-translational modifications and degradation processes. ![]() In the secretory pathway, the pH is becoming gradually more acidic from the endoplasmic reticulum (ER) to secretory vesicles or to the vacuole/lysosomes. An acute regulation of the intracellular pH is particularly important for most of the biological processes because protein structures as well as enzyme activities rely on this parameter 1.
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