Wang Lixin, Duncan George
School of Biological Sciences, University of East Anglia, Norwich, NR4 7TJ, UK.
Exp Cell Res. 2006 May 1;312(8):1439-46. doi: 10.1016/j.yexcr.2006.01.004. Epub 2006 Feb 9.
Sigma receptors have no known homology with other receptor systems, have no known natural ligands, but appear to play a critical role in a large diversity of cell functions. In the absence of a conventional pharmacology, siRNA technology provides a direct means of elucidating the major cell signaling pathways influenced by this receptor system. The non-transformed human lens cell line FHL124 was found to express the sigma-1 receptor (Sig-1R) and was employed for these studies. 72 h of transfection with either of the two siRNA directed against the sigma-1 receptor reduced messenger RNA and protein levels by over 70 and 60% respectively. Subsequent incubation for 96 h in culture medium (EMEM) supplemented with 5% serum gave a partial recovery of message, but there was no significant increase in protein. LDH leakage assays showed that significant cell death occurred during this time with an increased expression of caspase-3. Thrombin (10 nM) drives the growth of lens cells with a concomitant increase in ERK and Akt phosphorylation. These increases were inhibited in the cells where knockdown had occurred but not in cells exposed to scrambled siRNA. This study establishes a central role for Sig-1R in cell survival and death.
西格玛受体与其他受体系统没有已知的同源性，没有已知的天然配体，但似乎在多种细胞功能中发挥关键作用。在缺乏传统药理学方法的情况下，小干扰RNA（siRNA）技术提供了一种直接手段，用于阐明受该受体系统影响的主要细胞信号通路。发现未转化的人晶状体细胞系FHL124表达西格玛-1受体（Sig-1R），并用于这些研究。用两种针对西格玛-1受体的siRNA之一进行72小时转染后，信使核糖核酸（mRNA）和蛋白质水平分别降低了70%以上和60%以上。随后在补充有5%血清的培养基（伊格尔最低必需培养基，EMEM）中培养96小时，mRNA有部分恢复，但蛋白质没有显著增加。乳酸脱氢酶（LDH）泄漏试验表明，在此期间发生了显著的细胞死亡，同时半胱天冬酶-3的表达增加。凝血酶（10纳摩尔）驱动晶状体细胞生长，同时细胞外信号调节激酶（ERK）和蛋白激酶B（Akt）磷酸化增加。在发生基因敲低的细胞中这些增加受到抑制，但在暴露于乱序siRNA的细胞中没有受到抑制。这项研究确立了Sig-1R在细胞存活和死亡中的核心作用。
