Sakai Hiroshi, Shen Xiang, Koga Takahisa, Park Bum-Chan, Noskina Yelina, Tibudan Martin, Yue Beatrice Y J T
Department of Ophthalmology and Visual Sciences, University of Illinois at Chicago College of Medicine, Chicago, Illinois 60612, USA.
J Cell Physiol. 2007 Dec;213(3):775-84. doi: 10.1002/jcp.21147.
The trabecular meshwork (TM), an ocular tissue next to the cornea, is a major site for regulation of the aqueous humor outflow. Malfunctioning of this tissue is believed to be responsible for development of glaucoma, a major blinding disease. Myocilin is a gene directly linked to the most common form of glaucoma. Its protein product has been localized to both intra- and extra-cellular sites in TM cells. This study was to investigate the association of myocilin with mitochondria in TM cells. In vitro mitochondrial import assays showed that myocilin was imported to the TM mitochondria, targeting to mitochondrial membranes and/or the intermembrane space. The targeting was mediated mostly via the amino-terminal region of myocilin. When myocilin expression was induced either by treatment with dexamethasone or transfection with a myocilin construct, the mitochondrial membrane potential in TM cells, as assessed by JC-1 staining, was lowered. Subcellular fractionation and Western blot analyses confirmed that a portion of myocilin sedimented with the mitochondrial fractions. Upon anti-Fas treatment to provoke apoptosis, an increase of myocilin distribution in cytosolic fraction was observed, suggesting that myocilin was partially released from mitochondrial compartments. These results confirmed the association of myocilin with TM cell mitochondria and indicated that myocilin may have a proapoptotic role in TM cells.
小梁网（TM）是紧邻角膜的眼组织，是调节房水流出的主要部位。该组织功能失调被认为是导致青光眼（一种主要致盲疾病）的原因。肌纤蛋白是与最常见形式的青光眼直接相关的基因。其蛋白质产物已定位在TM细胞的细胞内和细胞外位点。本研究旨在探讨肌纤蛋白与TM细胞中线粒体的关联。体外线粒体导入试验表明，肌纤蛋白被导入到TM线粒体中，定位于线粒体外膜和/或膜间隙。这种定位主要通过肌纤蛋白的氨基末端区域介导。当通过地塞米松处理或用肌纤蛋白构建体转染诱导肌纤蛋白表达时，通过JC-1染色评估，TM细胞中的线粒体膜电位降低。亚细胞分级分离和蛋白质印迹分析证实，一部分肌纤蛋白与线粒体分级分离物一起沉淀。在用抗Fas处理引发细胞凋亡后，观察到细胞溶质部分中肌纤蛋白分布增加，表明肌纤蛋白从线粒体区室中部分释放。这些结果证实了肌纤蛋白与TM细胞线粒体的关联，并表明肌纤蛋白可能在TM细胞中具有促凋亡作用。
