Department of Biochemistry and Molecular Biology, University of Louisville School of Medicine, 580 South Preston Street, Louisville, KY 40202, USA.
Oxid Med Cell Longev. 2013;2013:628615. doi: 10.1155/2013/628615. Epub 2013 Feb 5.
This laboratory previously identified an albumin-like protein (denoted as p70) as a component of the macromolecular complex assembled within the 5'-regulatory region of redox-sensitive genes in vascular smooth muscle cells (vSMCs). Here we show that p70 is present in the cytosolic and nuclear compartments of vSMCs and dynamically responsive to redox status. Intense cytoplasmic and perinuclear staining, coupled with enhanced nuclear localization, was observed in vSMCs, but not HepG2 cells, treated with benzo(a)pyrene (BaP), H(2)O(2), or N-acetylcysteine, agents known to modulate redox status. 3' RACE indicated that p70 is not generated as a product of endogenous gene expression, but rather taken up from the extracellular compartment. While p70 was undetectable in cells grown for 24 hours under serum-free conditions, cell-associated, acid-resistant albumin was detected 30 min after the addition of exogenous albumin. vSMCs incubated at 4°C with 100 μ g/mL unlabeled BSA and 10 μ g/mL FITC-BSA for 60 minutes and switched to 37°C to examine temperature-sensitive label uptake showed punctate structures throughout the cell consistent with albumin internalization at the higher temperature. Albumin was found to influence redox-signaling, as evidenced by modulation of cyp1a1 gsta1 and Ha-ras gene inducibility. Together, these results implicate albumin and albumin-like proteins as critical regulators of vascular redox signaling.
本实验室先前鉴定出一种白蛋白样蛋白（表示为 p70），作为血管平滑肌细胞（vSMCs）中氧化还原敏感基因 5'调控区组装的大分子复合物的组成部分。在这里，我们表明 p70存在于 vSMCs 的细胞质和核区室中，并且对氧化还原状态具有动态响应。在用苯并（a）芘（BaP）、H2O2 或 N-乙酰半胱氨酸处理的 vSMCs 中观察到强烈的细胞质和核周染色，同时增强了核定位，但在 HepG2 细胞中未观察到这种情况，这些试剂已知可调节氧化还原状态。3' RACE 表明，p70不是作为内源性基因表达的产物产生的，而是从细胞外区室摄取的。虽然在无血清条件下培养 24 小时的细胞中无法检测到 p70，但在添加外源性白蛋白 30 分钟后检测到细胞相关的、耐酸的白蛋白。将 vSMCs 在 4°C 下与 100 μg/mL 未标记的 BSA 和 10 μg/mL FITC-BSA 孵育 60 分钟，然后切换至 37°C 以检查温度敏感的标记摄取，结果显示整个细胞中都有点状结构，这与在较高温度下的白蛋白内化一致。白蛋白被发现影响氧化还原信号，这可以通过调节 cyp1a1 gsta1 和 Ha-ras 基因的诱导性来证明。总之，这些结果表明白蛋白和白蛋白样蛋白是血管氧化还原信号的关键调节剂。
