Shaw A C, Røssel Larsen M, Roepstorff P, Justesen J, Christiansen G, Birkelund S
Department of Medical Microbiology and Immunology, University of Aarhus, Denmark.
Electrophoresis. 1999 Apr-May;20(4-5):984-93. doi: 10.1002/(SICI)1522-2683(19990101)20:4/5<984::AID-ELPS984>3.0.CO;2-R.
Interferon gamma (IFN-gamma) is a potent immunomodulatory lymphokine, secreted by activated T-lymphocytes and NK-cells during the cellular immune response. Actions of IFN-gamma are mediated through binding to the IFN-gamma-receptor, present on most cells, and the subsequent activation of a great magnitude of IFN-gamma responsive genes has been reported previously. Our goal is to identify and map IFN-gamma-regulated HeLa cell proteins to the two-dimensional polyacrylamide gel electrophoresis with the immobilized pH gradient (IPG) two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) system. A semiconfluent layer of HeLa cells was grown on tissue culture plates, and changes in protein expression due to 100 U/mL IFN-gamma were investigated at different periods after treatment, using pulse labeling with [35S]methionine/cysteine in combination with 2-D PAGE (IPG). The identity of eight protein spots was elucidated by matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS), and several variants of the IFN-gamma-inducible tryptophanyl-tRNA synthetase (hWRS) were detected by immunoblotting.
干扰素γ（IFN-γ）是一种强效的免疫调节性淋巴因子，在细胞免疫反应期间由活化的T淋巴细胞和自然杀伤细胞分泌。IFN-γ的作用是通过与大多数细胞上存在的IFN-γ受体结合来介导的，并且先前已经报道了大量IFN-γ反应性基因的随后激活。我们的目标是使用固定化pH梯度（IPG）二维聚丙烯酰胺凝胶电泳（2-D PAGE）系统，将IFN-γ调节的HeLa细胞蛋白鉴定并映射到二维聚丙烯酰胺凝胶电泳上。在组织培养板上培养半汇合层的HeLa细胞，并在处理后的不同时间段，使用[35S]甲硫氨酸/半胱氨酸脉冲标记结合2-D PAGE（IPG）研究由于100 U/mL IFN-γ引起的蛋白质表达变化。通过基质辅助激光解吸/电离质谱（MALDI-MS）阐明了八个蛋白质斑点的身份，并通过免疫印迹检测到了IFN-γ诱导的色氨酰-tRNA合成酶（hWRS）的几种变体。
