HEAVY-CHAIN BINDING-PROTEIN (BIP/GRP78) AND ENDOPLASMIN ARE EXPORTED FROM THE ENDOPLASMIC-RETICULUM IN RAT EXOCRINE PANCREATIC-CELLS, SIMILAR TO PROTEIN DISULFIDE-ISOMERASE
HEAVY-CHAIN BINDING-PROTEIN (BIP/GRP78) AND ENDOPLASMIN ARE EXPORTED FROM THE ENDOPLASMIC-RETICULUM IN RAT EXOCRINE PANCREATIC-CELLS, SIMILAR TO PROTEIN DISULFIDE-ISOMERASE
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DOI：
10.1016/0003-9861(92)90554-a
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发表时间：
1992-07-01
影响因子：
3.9
通讯作者：
TASHIRO, Y
中科院分区：
文献类型：
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作者：
TAKEMOTO, H;YOSHIMORI, T;TASHIRO, Y
在大鼠胰腺外分泌细胞中，蛋白质二硫键异构酶（PDI）是许多细胞内质网（ER）腔中的主要蛋白质之一，尽管其羧基端具有ER滞留信号KDEL（Lys <$Asp <$Glu <$Leu），但PDI不仅定位于ER，而且定位于高尔基体、分泌颗粒、质膜甚至腺腔中。在这份报告中，我们研究了是否其他ER管腔蛋白轴承KDEL信号在其C-末端，如BiP/GRP 78和内质网蛋白/GRP 94也从ER输出。我们制备了两种亲和纯化的多克隆抗体;一种是针对合成肽的12个氨基酸，这是相同的羧基末端的BiP和另一个对纯化的血浆内皮素。使用这两种抗体的免疫印迹分析表明，BiP和内质蛋白存在于质膜和微粒体组分中，类似于大鼠外分泌胰腺中PDI的细胞内分布。然而，在两个级分中的三种蛋白质的量的比率是可变的，这表明携带KDEL的蛋白质如PDI、BiP和内质网蛋白以不同的效率从ER输出。着床后蛋白A-免疫电镜显示，内质网出口和分泌到细胞外空间。大鼠胰腺小叶中PDI的分泌被Brefeldin A（BFA）和胍酸酯（FOY-305）抑制，这两种物质已知是细胞内转运的抑制剂。结合Akagi et al.（1988），强烈提示在大鼠胰腺外分泌细胞中，在细胞外间隙中发现的PDI和其他携带KDEL的蛋白质在孵育期间不是通过细胞损伤人工释放的，而是通过正常分泌途径分泌的。
Previously we found that in rat exocrine pancreatic cells, protein disulfide-isomerase (PDI), one of the major resident proteins in the lumen of the endoplasmic reticulum (ER) of many cells, is localized not only in the ER but also in the Golgi apparatus, secretory granules, plasma membranes, and even in the glandular lumens, despite possessing the ER retention signal KDEL (LysAspGluLeu) at the carboxyl terminus. In this report, we examined whether other ER luminal proteins bearing the KDEL signal at their C-termini, such as BiP/GRP78 and endoplasmin/GRP94 are also exported from the ER. We prepared two kinds of affinity-purified polyclonal antibodies; one against a synthetic peptide with 12 amino acids which is identical to the carboxyl terminus of BiP and another against purified endoplasmin. Immunoblot analysis using these two antibodies showed that BiP and endoplasmin exist in both the plasma membrane and the microsomal fractions, similar to the intracellular distribution of PDI in rat exocrine pancreas. The ratios of the amount of the three proteins in the two fractions, however, were variable, suggesting that the KDEL-bearing proteins such as PDI, BiP, and endoplasmin are exported from the ER with different efficiencies. Postembedding protein A-immunogold electron microscopy revealed that endoplasmin was exported from the ER and secreted to the extracellular space. The secretion of PDI in rat pancreatic lobules was inhibited by Brefeldin A (BFA) and by guanidino acid esters (FOY-305), which are known to be the inhibitors of the intracellular transport. Taken together with the previous immunogold electron microscopic analyses by Akagiet al.(1988), it is strongly suggested that in rat exocrine pancreatic cells PDI and the other KDEL-bearing proteins found in the extracellular space were not artificially released by cell damage during incubation but were secreted via the normal secretory pathway.