Matlin K S, Simons K
J Cell Biol. 1984 Dec;99(6):2131-9. doi: 10.1083/jcb.99.6.2131.
In Madin-Darby canine kidney (MDCK) cells (a polarized epithelial cell line) infected with influenza virus, the hemagglutinin behaves as an apical plasma membrane glycoprotein. To determine biochemically the domain on the plasma membrane, apical or basolateral, where newly synthesized hemagglutinin first appears, cells were cultured on Millipore filters to make both cell surface domains independently accessible. Hemagglutinin in virus-infected cells was pulse-labeled, chased, and detected on the plasma membrane with a sensitive trypsin assay. Under all conditions tested, newly made hemagglutinin appeared simultaneously on both domains, with the bulk found in the apical membrane. When trypsin was continuously present on the basolateral surface during the chase, little hemagglutinin was cleaved relative to the amount transported apically. In addition, specific antibodies against the hemagglutinin placed basolaterally had no effect on transport to the apical domain. These observations suggested that most newly synthesized hemagglutinin does not transiently appear on the basolateral surface but rather is delivered directly to the apical surface in amounts that account for its final polarized distribution.
在感染流感病毒的犬肾传代细胞（MDCK，一种极化上皮细胞系）中，血凝素表现为顶端质膜糖蛋白。为了从生化角度确定质膜上新合成的血凝素首先出现的区域是顶端还是基底外侧，细胞在微孔滤膜上培养，以使两个细胞表面区域都能独立被检测。对病毒感染细胞中的血凝素进行脉冲标记、追踪，并通过灵敏的胰蛋白酶检测法在质膜上进行检测。在所有测试条件下，新合成的血凝素同时出现在两个区域，大部分存在于顶端膜中。当在追踪过程中基底外侧表面持续存在胰蛋白酶时，相对于顶端运输的量，很少有血凝素被裂解。此外，基底外侧放置的针对血凝素的特异性抗体对向顶端区域的运输没有影响。这些观察结果表明，大多数新合成的血凝素不会短暂出现在基底外侧表面，而是直接以与其最终极化分布相符的量被递送至顶端表面。
