Burkhardt J K, Wiebel F A, Hester S, Argon Y
Department of Immunology, Duke Medical Center, Durham, North Carolina 27710.
J Exp Med. 1993 Dec 1;178(6):1845-56. doi: 10.1084/jem.178.6.1845.
Chediak-Higashi Syndrome (CHS) is an autosomal recessive disease affecting secretory granules and lysosomes-like organelles. In CHS fibroblasts, acidic organelles are abnormally large and clustered in the perinuclear area. We have analyzed fibroblast cell lines from a CHS patient and from the murine model for CHS, the beige mouse, to determine which lysosome-like compartments are affected. Uptake of neutral red showed that in both beige and CHS cell lines, the acidic organelles were markedly clustered in the perinuclear region of the cells. Giant organelles (> 4 microns) were observed in a fraction of the cells, and these were more dramatic in the beige fibroblasts than in the CHS fibroblasts. The total dye uptake of both mutant cell lines was similar to their respective wild type fibroblasts, suggesting that the overall volume of acidic compartments is unaffected by the disorder. Histochemistry and immunofluorescence showed that the giant organelles in both beige and CHS fibroblasts were positive for cathepsin D, lysosome-associated membrane protein (LAMP) 1, LAMP 2, and a 120-kD lysosomal glycoprotein, all marker proteins for late endosomes and lysosomes. The giant organelles were also negative for transferrin receptor and mannose-6-phosphate receptor, and most of them were also negative for rab 7. This distribution of marker proteins shows that the giant organelles in both beige and CHS are derived from late compartments of the endocytic pathway. This conclusion was confirmed using endocytic tracers. BSA was transported to the giant organelles, but only after long incubation times, and only at 37 degrees C. alpha 2-Macroglobulin was taken up and degraded at similar rates by CHS or beige cells and their respective wild type control cells. Taken together, our results indicate that the mutation in CHS specifically affects late endosomes and lysosomes, with little or no effect on early endosomes. Although the mutation clearly causes mislocalization of these organelles, it appears to have little effect on their endocytic and degradative functions.
切迪阿克-希加希综合征（CHS）是一种常染色体隐性疾病，会影响分泌颗粒和类溶酶体细胞器。在CHS成纤维细胞中，酸性细胞器异常大且聚集在核周区域。我们分析了一名CHS患者以及CHS小鼠模型（米色小鼠）的成纤维细胞系，以确定哪些类溶酶体区室受到影响。中性红摄取实验表明，在米色和CHS细胞系中，酸性细胞器均明显聚集在细胞的核周区域。在一部分细胞中观察到了巨型细胞器（>4微米），且在米色成纤维细胞中比在CHS成纤维细胞中更为显著。两种突变细胞系的总染料摄取量与其各自的野生型成纤维细胞相似，这表明酸性区室的总体积不受该疾病影响。组织化学和免疫荧光显示，米色和CHS成纤维细胞中的巨型细胞器组织蛋白酶D、溶酶体相关膜蛋白（LAMP）1、LAMP 2以及一种120-kD溶酶体糖蛋白均呈阳性，这些都是晚期内体和溶酶体的标记蛋白。巨型细胞器转铁蛋白受体和甘露糖-6-磷酸受体也呈阴性，并且大多数巨型细胞器rab 7也呈阴性。标记蛋白的这种分布表明，米色和CHS中的巨型细胞器均源自内吞途径的晚期区室。使用内吞示踪剂证实了这一结论。牛血清白蛋白（BSA）被转运到巨型细胞器，但仅在长时间孵育后，且仅在37摄氏度时才会发生。α2-巨球蛋白被CHS或米色细胞及其各自的野生型对照细胞以相似的速率摄取和降解。综上所述，我们的结果表明，CHS中的突变特异性地影响晚期内体和溶酶体，对早期内体几乎没有影响。尽管该突变明显导致这些细胞器的定位错误，但似乎对其内吞和降解功能影响很小。
