Involvement of syntaxin 18, an endoplasmic reticulum (ER)-localized SNARE protein, in ER-mediated phagocytosis
Involvement of syntaxin 18, an endoplasmic reticulum (ER)-localized SNARE protein, in ER-mediated phagocytosis
复制标题
DOI：
10.1091/mbc.e05-12-1174
复制
发表时间：
2006-09-01
影响因子：
3.3
通讯作者：
Wada, Ikuo
中科院分区：
文献类型：
--
作者：
Hatsuzawa, Kiyotaka;Tamura, Taku;Wada, Ikuo
内质网（ER）被认为在吞噬作用中发挥重要的结构和功能作用。根据这一模型，ER和质膜或吞噬体膜之间的直接膜融合必须先于进一步的内陷，但确切的机制仍然难以捉摸。在这里，我们研究了各种ER定位的SNARE蛋白是否参与了这一融合过程。当从小鼠J774巨噬细胞中分离出吞噬体时，我们发现ER定位的SNARE蛋白（syntaxin 18，D12和Sec 22 b）在吞噬体中显著富集。荧光和免疫-EM分析证实了突触融合蛋白18在J774细胞的吞噬体膜中的定位，所述J774细胞稳定表达标记到GFP变体的这种蛋白。为了检查这些SNARE蛋白是否是吞噬作用所需的，我们产生了稳定表达Fc γ受体的293 T细胞，其中吞噬作用以IgG介导的方式发生。在这些细胞中，突触融合蛋白18或D12缺乏跨膜结构域的显性阴性突变体的表达，但不是Sec 22 b突变体，损害了吞噬作用。Syntaxin 18小干扰RNA（siRNA）选择性地降低吞噬效率，而稳定过表达syntaxin 18可显著提高J774细胞的吞噬率。因此，我们得出结论，syntaxin 18参与ER介导的吞噬作用，大概是通过调节ER和质膜或吞噬体膜的特异性和直接融合。
The endoplasmic reticulum (ER) is thought to play an important structural and functional role in phagocytosis. According to this model, direct membrane fusion between the ER and the plasma or phagosomal membrane must precede further invagination, but the exact mechanisms remain elusive. Here, we investigated whether various ER-localized SNARE proteins are involved in this fusion process. When phagosomes were isolated from murine J774 macrophages, we found that ER-localized SNARE proteins (syntaxin 18, D12, and Sec22b) were significantly enriched in the phagosomes. Fluorescence and immuno-EM analyses confirmed the localization of syntaxin 18 in the phagosomal membranes of J774 cells stably expressing this protein tagged to a GFP variant. To examine whether these SNARE proteins are required for phagocytosis, we generated 293T cells stably expressing the Fcy receptor, in which phagocytosis occurs in an IgG-mediated manner. Expression in these cells of dominant-negative mutants of syntaxin 18 or D12 lacking the transmembrane domain, but not a Sec22b mutant, impaired phagocytosis. Syntaxin 18 small interfering RNA (siRNA) selectively decreased the efficiency of phagocytosis, and the rate of phagocytosis was markedly enhanced by stable overexpression of syntaxin 18 in J774 cells. Therefore, we conclude that syntaxin 18 is involved in ER-mediated phagocytosis, presumably by regulating the specific and direct fusion of the ER and plasma or phagosomal membranes.