Vanderbilt University Mass Spectrometry Research Center, Nashville, Tennessee, USA; Vanderbilt University Chemical and Physical Biology Program, Nashville, Tennessee, USA.
Vanderbilt University Mass Spectrometry Research Center, Nashville, Tennessee, USA; Vanderbilt University Department of Biochemistry, Nashville, Tennessee, USA.
Mol Cell Proteomics. 2023 Jan;22(1):100453. doi: 10.1016/j.mcpro.2022.100453. Epub 2022 Dec 5.
The eye lens is responsible for focusing and transmitting light to the retina. The lens does this in the absence of organelles, yet maintains transparency for at least 5 decades before onset of age-related nuclear cataract (ARNC). It is hypothesized that oxidative stress contributes significantly to ARNC formation. It is in addition hypothesized that transparency is maintained by a microcirculation system that delivers antioxidants to the lens nucleus and exports small molecule waste. Common data-dependent acquisition methods are hindered by dynamic range of lens protein expression and provide limited context to age-related changes in the lens. In this study, we utilized data-independent acquisition mass spectrometry to analyze the urea-insoluble membrane protein fractions of 16 human lenses subdivided into three spatially distinct lens regions to characterize age-related changes, particularly concerning the lens microcirculation system and oxidative stress response. In this pilot cohort, we measured 4788 distinct protein groups, 46,681 peptides, and 7592 deamidated sequences, more than in any previous human lens data-dependent acquisition approach. Principally, we demonstrate that a significant proteome remodeling event occurs at approximately 50 years of age, resulting in metabolic preference for anaerobic glycolysis established with organelle degradation, decreased abundance of protein networks involved in calcium-dependent cell-cell contacts while retaining networks related to oxidative stress response. Furthermore, we identified multiple antioxidant transporter proteins not previously detected in the human lens and describe their spatiotemporal and age-related abundance changes. Finally, we demonstrate that aquaporin-5, among other proteins, is modified with age by post-translational modifications including deamidation and truncation. We suggest that the continued accumulation of each of these age-related outcomes in proteome remodeling contribute to decreased fiber cell permeability and result in ARNC formation.
晶状体负责聚焦和将光线传输到视网膜。在没有细胞器的情况下，晶状体完成了这一过程，然而，在与年龄相关的核性白内障（ARNC）发生之前，它至少保持透明 50 年。有人假设氧化应激对 ARNC 的形成有重要贡献。此外，有人假设透明性是通过一个微循环系统来维持的，该系统将抗氧化剂输送到晶状体核，并将小分子废物输出。常见的数据依赖采集方法受到晶状体蛋白表达动态范围的限制，并且为晶状体与年龄相关的变化提供的背景信息有限。在这项研究中，我们利用非依赖性数据采集质谱分析了 16 个人晶状体的尿素不溶性膜蛋白部分，这些晶状体被细分为三个空间上不同的晶状体区域，以表征与年龄相关的变化，特别是涉及晶状体微循环系统和氧化应激反应的变化。在这个试点队列中，我们测量了 4788 个不同的蛋白质组、46681 个肽和 7592 个脱酰胺序列，比以往任何人类晶状体数据依赖采集方法都多。主要的是，我们证明了大约在 50 岁时发生了一个显著的蛋白质组重塑事件，导致代谢偏好无氧糖酵解，同时伴随着细胞器降解，与钙依赖性细胞间接触相关的蛋白质网络丰度降低，而保留与氧化应激反应相关的网络。此外，我们鉴定了多个以前在人类晶状体中未检测到的抗氧化剂转运蛋白，并描述了它们的时空和与年龄相关的丰度变化。最后，我们证明了水通道蛋白-5（aquaporin-5）和其他蛋白质一样，随着年龄的增长，会发生翻译后修饰，包括脱酰胺和截断。我们认为，这些与年龄相关的蛋白质组重塑的每一个结果的持续积累，都会导致纤维细胞通透性降低，并导致 ARNC 的形成。
