Gap junction endocytosis and lysosomal degradation of connexin43-P2 in WB-F344 rat liver epithelial cells treated with DDT and lindane
Gap junction endocytosis and lysosomal degradation of connexin43-P2 in WB-F344 rat liver epithelial cells treated with DDT and lindane
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DOI：
10.1093/carcin/17.9.1791
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发表时间：
1996-09-01
期刊：
影响因子：
4.7
通讯作者：
Ruch, RJ
中科院分区：
文献类型：
--
作者：
Guan, XJ;Ruch, RJ
DDT（1,1-二（对氯苯基）-2,2,2-三氯乙烷）或林丹处理WB-F344大鼠肝上皮细胞诱导间隙连接斑块的丢失和磷酸化的间隙连接蛋白connexin43-P2 （Cx43-P2）的减少，这与斑块有关。在本研究中，我们考虑了几种机制，连接斑块的丢失可能是由于连接颗粒的分解或斑块的内吞作用；而Cx43-P2的缺失可能是由于去磷酸化或降解。DDT或林丹处理细胞的免疫组化分析显示，质膜性cx43阳性间隙连接斑块减少，与cx43阳性点状细胞质结构的出现一致，细胞质cx43阳性结构在处理4小时后最终消失。DDT或林丹处理后未见弥漫性cx43阳性质膜染色，这些细胞的Western blot分析表明，Cx43-P2以一种时间依赖性的方式减少，与质膜间隙连接斑块的消失平行。Cx43-P2的丧失不是由于去磷酸化，因为非磷酸化（Cx43-NP）或其他磷酸化（Cx43-P1）形式的蛋白没有明显增加。秋水仙碱和氯喹阻止了Cx43-P2的减少和细胞质中cx43 -阳性结构的消失，这表明含有Cx43-P2的斑块在溶酶体中被内化和降解，此外，Western blots中出现了两条与Cx43-P2丢失一致的小条带（类似于18和22 kDa），可能是该蛋白的降解产物。这些免疫组织化学和生化数据强烈提示，DDT和林丹处理的WB-F344细胞中间隙连接斑块和Cx43-P2的丢失是由于斑块的内吞作用和溶酶体中Cx43-P2的降解。
Treatment of WB-F344 rat liver epithelial cells with DDT (1,1-bis(p-chlorophenyl)-2,2,2-trichloroethane) or lindane induces a loss of gap junction plaques and a decrease in the phosphorylated gap junction protein connexin43-P2 (Cx43-P2), which is associated with the plaques, In this study we have considered several mechanisms, The loss of junctional plaques could be due to disaggregation of junctional particles or to endocytosis of the plaques, while the loss of Cx43-P2 could be due to dephosphorylation or degradation. Immunohistochemical analyses of DDT- or lindane-treated cells revealed a reduction in plasma membranous Cx43-positive gap junction plaques coincident with the appearance of Cx43-positive punctate cytoplasmic structures, The cytoplasmic Cx43-positive structures eventually disappeared after 4 h treatment. Diffuse Cx43-positive plasma membranous staining was not seen following DDT or lindane treatment, Western blot analyses of these cells indicated that Cx43-P2 decreased in a time-dependent manner that paralleled the disappearance of gap junction plaques from the plasma membrane, The loss of Cx43-P2 was not due to dephosphorylation, since no increase in non-phosphorylated (Cx43-NP) or other phosphorylated (Cx43-P1) forms of the protein were evident, The decrease in Cx43-P2 and the disappearance of cytoplasmic Cx43-positive structures were prevented by colchicine and chloroquine, which suggests that Cx43-P2-containing plaques were internalized and degraded in lysosomes, In addition, two small (similar to 18 and similar to 22 kDa) bands appeared in Western blots coincident with the loss of Cx43-P2 and may be degradation products of the protein, These immunohistochemical and biochemical data strongly suggest that the loss of gap junction plaques and of Cx43-P2 in WB-F344 cells treated with DDT and lindane were due to endocytosis of the plaques and degradation of Cx43-P2 in lysosomes.