Secretogranin III binds to cholesterol in the secretory granule membrane as an adapter for chromogranin A
Secretogranin III binds to cholesterol in the secretory granule membrane as an adapter for chromogranin A
复制标题
DOI：
10.1074/jbc.m310104200
复制
发表时间：
2004-01-30
影响因子：
4.8
通讯作者：
Takeuchi, T
中科院分区：
文献类型：
--
作者：
Hosaka, M;Suda, M;Takeuchi, T
颗粒蛋白家族蛋白，包括嗜铬粒蛋白 A (CgA) 和分泌颗粒蛋白 III (SgIII)，被转运至神经内分泌细胞中的分泌颗粒 (SG)。我们之前表明，SgIII 在颗粒内环境中与 CgA 强烈结合，并将 CgA 靶向垂体和胰腺内分泌细胞中的 SG。在这项研究中，我们证明，在大鼠胰岛素瘤来源的 INS-1 细胞匀浆的蔗糖密度梯度下，尽管缺乏跨膜区域，SgIII 仍定位于 SG 级分并被分级到 SG 膜 (SGM)。使用甲基-β-环糊精去除 SGM 中的胆固醇后，SgIII 与 SGM 的结合受到损害。 SgIII 和 CgA 均被 Triton X-100 从 SGM 中溶解，而 Triton X-100 不溶解羧肽酶 E。SgIII 和羧肽酶 E 在颗粒内条件下与 SGM 型脂质体强烈结合，但 CgA 则不然。相反，CgA 仅在 SgIII 存在的情况下才与 SGM 型脂质体结合。免疫细胞化学和脉冲追踪实验表明，删除 N 末端脂质结合区的 SgIII 与小鼠促肾上腺皮质激素来源的 AtT-20 细胞的组成型通路发生错误分选。因此，我们认为 SgIII 直接与 SGM 的胆固醇成分结合，并将 CgA 靶向垂体和胰腺内分泌细胞中的 SG。
Granin-family proteins, including chromogranin A (CgA) and secretogranin III (SgIII), are transported to secretory granules (SGs) in neuroendocrine cells. We previously showed that SgIII binds strongly to CgA in an intragranular milieu and targets CgA to SGs in pituitary and pancreatic endocrine cells. In this study, we demonstrated that with a sucrose density gradient of rat insulinoma-derived INS-1 cell homogenates, SgIII was localized to the SG fraction and was fractionated to the SG membrane (SGM) despite lacking the transmembrane region. With depletion of cholesterol from the SGM using methyl-beta-cyclodextrin, SgIII was impaired to bind to the SGM. Both SgIII and CgA were solubilized from the SGM by Triton X-100 in contrast to the Triton X-100 insolubility of carboxypeptidase E. SgIII and carboxypeptidase E strongly bound to the SGM-type liposome in intragranular conditions, but CgA did not. Instead, CgA bound to the SGM-type liposome only in the presence of SgIII. Immunocytochemical and pulse-chase experiments revealed that SgIII deleting the N-terminal lipid-binding region missorted to the constitutive pathway in mouse corticotroph-derived AtT-20 cells. Thus, we suggest that SgIII directly binds to cholesterol components of the SGM and targets CgA to SGs in pituitary and pancreatic endocrine cells.