Hülser D F, Rehkopf B, Traub O
Biologisches Institut, Abt. Biophysik, Universität Stuttgart,
Exp Cell Res. 1997 Jun 15;233(2):240-51. doi: 10.1006/excr.1997.3568.
An indirect immunogold labeling technique was applied to replicas of freeze-fractured membranes of rapidly frozen unfixed cells. The endogenous gap junction protein Cx43 of BICR/M1Rk rat mammary tumor cells was preferentially identified in quasi-crystalline gap junction plaques as were the transfected connexins Cx40, Cx43, and Cx45 in HeLa (human cervical carcinoma) cells. With this method we also detected contact areas with dispersed gap junction channels which are the only structural correlation for endogenous Cx45 in HeLa wild-type cells where no gap junction plaques exist. In double-transfected HeLa cells a colocalization of Cx40 and Cx43 was occasionally detected in quasi-crystalline gap junction plaques, whereas in contact areas with dispersed particles only one Cx type was present. Our results indicate that functional gap junction channels exist outside the quasi-crystalline plaques.
一种间接免疫金标记技术应用于快速冷冻未固定细胞的冷冻断裂膜复制品。在准晶体间隙连接斑中优先鉴定出BICR/M1Rk大鼠乳腺肿瘤细胞的内源性间隙连接蛋白Cx43，在HeLa（人宫颈癌）细胞中转染的连接蛋白Cx40、Cx43和Cx45也是如此。用这种方法，我们还检测到了具有分散间隙连接通道的接触区域，这是HeLa野生型细胞中内源性Cx45的唯一结构关联，在这些细胞中不存在间隙连接斑。在双转染的HeLa细胞中，偶尔在准晶体间隙连接斑中检测到Cx40和Cx43的共定位，而在具有分散颗粒的接触区域中仅存在一种Cx类型。我们的结果表明，功能性间隙连接通道存在于准晶体斑之外。
