Membrane Protein Rim21 Plays a Central Role in Sensing Ambient pH in Saccharomyces cerevisiae
Membrane Protein Rim21 Plays a Central Role in Sensing Ambient pH in Saccharomyces cerevisiae
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DOI：
10.1074/jbc.m112.394205
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发表时间：
2012-11-09
影响因子：
4.8
通讯作者：
Kihara, Akio
中科院分区：
文献类型：
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作者：
Obara, Keisuke;Yamamoto, Hayashi;Kihara, Akio
外部碱化激活酿酒酵母中的Rim101通路。在该通路中，三种整合膜蛋白Rim21、Dfg16和Rim9被认为是pH传感器机制的组成部分。然而，这些蛋白如何参与pH感应完全未知。在这项工作中，我们研究了Rim21、Dfg16和Rim9的定位、物理相互作用以及相互关系。发现这些蛋白形成一个复合物，并以斑片状且相互依赖的方式定位于质膜。它们的细胞水平也是相互依赖的。特别是，在dfg16Δ和rim9Δ细胞中，Rim21水平显著降低。在外部碱化时，这些蛋白被内化和降解。我们还证明Rim21的瞬时降解完全抑制了Rim101通路，但Dfg16或Rim9的降解则不会。这一发现有力地表明Rim21是pH传感器蛋白，而Dfg16和Rim9通过维持Rim21的水平并协助其在质膜上的定位发挥辅助功能。即使没有外部碱化，通过质子载体处理或质膜内叶中磷脂酰丝氨酸的消耗，Rim101通路也以依赖Rim21的方式被激活，这两种情况都像外部碱化一样导致质膜去极化。因此，质膜去极化似乎是pH传感器分子Rim21的关键信号之一。
External alkalization activates the Rim101 pathway in Saccharomyces cerevisiae. In this pathway, three integral membrane proteins, Rim21, Dfg16, and Rim9, are considered to be the components of the pH sensor machinery. However, how these proteins are involved in pH sensing is totally unknown. In this work, we investigated the localization, physical interaction, and interrelationship of Rim21, Dfg16, and Rim9. These proteins were found to form a complex and to localize to the plasma membrane in a patchy and mutually dependent manner. Their cellular level was also mutually dependent. In particular, the Rim21 level was significantly decreased in dfg16 Delta and rim9 Delta cells. Upon external alkalization, the proteins were internalized and degraded. We also demonstrate that the transient degradation of Rim21 completely suppressed the Rim101 pathway but that the degradation of Dfg16 or Rim9 did not. This finding strongly suggests that Rim21 is the pH sensor protein and that Dfg16 and Rim9 play auxiliary functions through maintaining the level of Rim21 and assisting in its plasma membrane localization. Even without external alkalization, the Rim101 pathway was activated in a Rim21-dependent manner by either protonophore treatment or depletion of phosphatidylserine in the inner leaflet of the plasma membrane, both of which caused plasma membrane depolarization like the external alkalization. Therefore, plasma membrane depolarization seems to be one of the key signals for the pH sensor molecule Rim21.