Further understanding of vesicular pH regulation and its impact on growth factor signaling

进一步了解囊泡 pH 调节及其对生长因子信号传导的影响

• 批准号: RGPIN-2016-03928
• 负责人: Dubois, Claire
• 金额: $ 2.4万
• 依托单位: Université de Sherbrooke
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2016
• 资助国家: 加拿大
• 起止时间: 2016-01-01 至 2017-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=614726
• 关键词: Further; understanding; vesicular; pH; regulation

The regulation of intracellular pH in all eukaryotic cells is crucial for the control of physiological processes that include cell growth and division. Cytoplasmic pH is maintained close to neutrality by efficient ion plasma membrane transport mechanisms. The situation is quite different in cellular vacuoles, which generally have a rather variable pH in the range of 4.5-6.5. Although much is known about transporters at the plasma membrane, our understanding of intravesicular ion transport and its consequences on cellular functions is at the present rudimentary. Two recently identified vesicular Na+/H+ exchangers, SLC9A6/NHE6 and SLC9A9/NHE9, are specifically located to intracellular vesicles and may contribute to the establishment and/or fine-tuning of intravesicular pH. Although of obvious importance, the contribution of vesicular NHEs as well as a direct link between their activity and an ensuing physiologically relevant effect remains to be established. We have found that incubation of fibroblastic cells under hypoxic conditions resulted in NHE-dependent acidification of intracellular vesicles. This event was associated with the activation of the transforming growth factor beta (TGFß) signal transducer, Smad3. Based on these observations, we propose the novel hypothesis that 1) the modulation of organellar exchanger activity by metabolic changes such as hypoxia is a key event in intravesicular pH regulation and, 2) enhanced functions of pH-sensitive proteins involved in TGFß signaling is part of the mechanism by which hypoxia influences cell functions that include cell growth. Over the next 5 years, we propose to identify some of the components and the rules governing vacuolar pH regulation and their role in the regulation of growth factor signaling using the universal TGFß system as a model. To achieve this, our specific objectives are: 1) to revisit the cellular components involved in the pump-and-leak model for regulation of intravesicular pH with a focus on the newly identified vesicular-type NA+/H+ exchangers; 2) to define how these exchangers adapt to metabolic changes such as hypoxia through identification of binding partners and regulators; 3) to define the impact of endosomal pH regulation on growth factor signaling using pH-sensitive and pH-insensitive TGFß signal regulators. Results from these studies will increase our understanding on the nature and the regulation of the components involved in intravesicular pH regulation. They will also reveal how endosomal pH and its regulation affect growth factor signaling pathways. This will be a key achievement that will link for the first time endosomal pH regulation and cell growth.

所有真核细胞中细胞内pH值的调节对于控制包括细胞生长和分裂在内的生理过程至关重要。通过有效的离子质膜转运机制，细胞质pH值保持接近中性。在细胞空泡中，情况完全不同，其通常具有在4.5-6.5范围内的相当可变的pH。虽然我们对质膜上的转运蛋白有很多了解，但我们对囊泡内离子转运及其对细胞功能的影响的了解目前还很不成熟。最近确定的两个囊泡Na+/H+交换器，SLC 9A 6/NHE 6和SLC 9A 9/NHE 9，是专门位于细胞内囊泡，并可能有助于建立和/或微调的囊泡内pH值。虽然明显的重要性，囊泡NHE的贡献，以及它们的活性和随后的生理相关的效果之间的直接联系仍然有待建立。我们已经发现成纤维细胞在缺氧条件下孵育导致细胞内囊泡的NHE依赖性酸化。该事件与转化生长因子β（TGF β）信号转导子Smad 3的激活有关。基于这些观察结果，我们提出了新的假设，即1）通过代谢变化（如缺氧）调节细胞器交换活性是囊内pH调节的关键事件，2）参与TGF β信号传导的pH敏感蛋白的功能增强是缺氧影响细胞功能（包括细胞生长）的机制的一部分。在接下来的5年里，我们建议确定一些组件和规则的液泡pH值调节和它们的作用，在调节生长因子信号转导的通用TGF β 1系统作为一个模型。为了实现这一点，我们的具体目标是：1）重新审视泵和泄漏模型中涉及的细胞成分，以调节囊泡内pH，重点是新发现的囊泡型NA+/H+交换器; 2）通过识别结合伴侣和调节剂来确定这些交换器如何适应代谢变化，如缺氧; 3）使用pH敏感性和pH不敏感性TGF β信号调节剂来定义内体pH调节对生长因子信号传导的影响。这些研究的结果将增加我们对囊泡内pH调节所涉及的组分的性质和调节的理解。他们还将揭示内体pH及其调节如何影响生长因子信号通路。这将是首次将内体pH调节与细胞生长联系起来的关键成就。