Institute of Environmental Health Sciences, Wayne State University, Detroit, MI, USA.
Department of Immunology and Microbiology, Wayne State University, Detroit, MI, USA.
J Proteomics. 2018 Feb 20;173:107-114. doi: 10.1016/j.jprot.2017.11.026. Epub 2017 Dec 2.
Exposure of Wehi-231 B-cells to Hg for 5min resulted in concentration dependent changes in protein phosphorylations. Phosphorylation was quantified using mass spectrometry to analyze TiO and anti-pTyr antibody selected phosphopeptides from Wehi-231 digests. The most frequent and largest amplitude responses to Hg exposure were increased phosphorylation although a decrease was observed for 1% of phosphoproteins detected in the untreated cells. A subset of proteins responded with an increase in phosphorylation to Hg exposure at low micromolar concentrations. The majority of proteins required Hg over 20μM in order to increase phosphorylation. Ser/Thr phosphorylations are prominent in the cytoskeletal organization and the GTPase signaling systems and these systems are notable as the primary ones responding to the lowest concentrations of Hg. Systems that required higher concentrations of Hg to increase phosphorylation included immune receptor signaling. The proteins for which an increase in phosphorylation occurred at Hg above 20μM have a higher proportion of pTyr sites. Anti Ig stimulation of Wehi-231 cells confirmed that cytoskeletal protein phosphorylation and GTPase signaling are modulated in physiologically relevant B-cell receptor activation. Candidate kinases that respond to Hg exposure at the low μM concentrations include MAP Kinase 1, CaM Kinase II delta and PAK2.
Mercury (Hg) is a wide spread environmental toxicant. Epidemiological and laboratory studies suggest that exposure to environmental Hg at current levels, which have been perceived to be non-toxic, may contribute to immune system dysfunction and autoimmune disease in humans and animals respectively. While we have previously shown that exposure of B lymphocytes to low levels of mercury interferes with B-cell receptor signaling mediated by post transcriptional phosphorylation events, overall the mechanism that is responsible for increased autoimmunity in mercury exposed human or animal populations is not well understood. The current study evaluated the dose dependent actions of mercury to change phosphorylation in the Wehi-231 cell line, an immature B-cell model in which actions of mercury on development of cell function can be evaluated. The study identified the cytoskeletal proteins as the most sensitive to modulation by mercury with changes in Ser/Thr phosphorylation being observed at the lowest concentrations of mercury. These findings indicate that the actions of mercury on B-cell immune function and development are at least in part likely mediated through changes in cytoskeletal protein phosphorylation.
将 Wehi-231 B 细胞暴露于汞中 5 分钟会导致蛋白质磷酸化发生浓度依赖性变化。使用质谱法定量磷酸化，以分析来自 Wehi-231 消化物的 TiO 和抗-pTyr 抗体选择的磷酸肽。对汞暴露最频繁和最大幅度的反应是磷酸化增加，尽管在未经处理的细胞中检测到的 1%磷酸蛋白中观察到减少。一小部分蛋白质对低微摩尔浓度的汞暴露表现出磷酸化增加。大多数蛋白质需要超过 20μM 的汞才能增加磷酸化。Ser/Thr 磷酸化在细胞骨架组织和 GTPase 信号系统中很突出，这些系统是对最低浓度汞的主要反应系统。需要更高浓度汞才能增加磷酸化的系统包括免疫受体信号。在高于 20μM 的汞浓度下磷酸化增加的蛋白质有更高比例的 pTyr 位点。Wehi-231 细胞的抗 Ig 刺激证实，生理相关 B 细胞受体激活会调节细胞骨架蛋白磷酸化和 GTPase 信号。在低 μM 浓度下响应汞暴露的候选激酶包括 MAP Kinase 1、CaM Kinase II delta 和 PAK2。
汞（Hg）是一种广泛存在的环境毒物。流行病学和实验室研究表明，暴露于当前被认为无毒的环境汞可能导致人类和动物的免疫系统功能障碍和自身免疫性疾病。虽然我们之前已经表明，B 淋巴细胞暴露于低水平的汞会干扰 B 细胞受体信号转导，这是由转录后磷酸化事件介导的，但总体而言，导致暴露于汞的人类或动物人群中自身免疫增加的机制尚不清楚。本研究评估了汞的剂量依赖性作用，以改变 Wehi-231 细胞系中的磷酸化，Wehi-231 细胞系是一种不成熟的 B 细胞模型，可在此模型中评估汞对细胞功能发育的作用。该研究发现细胞骨架蛋白对汞的调节最为敏感，在最低浓度的汞下观察到 Ser/Thr 磷酸化的变化。这些发现表明，汞对 B 细胞免疫功能和发育的作用至少部分可能是通过改变细胞骨架蛋白磷酸化介导的。
