Selective and specific internalization of clostridial C3 ADP-ribosyltransferases into macrophages and monocytes
Selective and specific internalization of clostridial C3 ADP-ribosyltransferases into macrophages and monocytes
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DOI：
10.1111/j.1462-5822.2009.01393.x
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发表时间：
2010-02-01
影响因子：
3.4
通讯作者：
Barth, Holger
中科院分区：
文献类型：
--
作者：
Fahrer, Joerg;Kuban, Jasmin;Barth, Holger
来自肉毒梭菌（C3 bot）和Clostridium limosum（C3 lim）的C3转移酶单-ADP-核糖基化，从而使真核细胞的RhoA、-B和-C活化。由于它们极差的细胞摄取，C3转移酶被认为是外切酶而不是外毒素，挑战了它们在发病机制中的作用。在这里，我们首次报道，低浓度的C3 lim和C3 bot在不到3小时内被选择性地内化到巨噬细胞/单核细胞中，诱导肌动蛋白细胞骨架通过ADP-核糖基化Rho进行重组。我们证明C3转移酶通过酸化的早期内体内化到巨噬细胞/单核细胞的胞质溶胶中。巴弗洛霉素A1是一种内体酸化抑制剂，可保护J774A.1巨噬细胞和人早幼粒白血病细胞（HL-60）免受C3中毒。此外，共聚焦激光扫描显微镜显示C3与早期内体共定位。细胞外酸性脉冲使细胞表面结合的C3直接易位穿过细胞质膜到胞质溶胶。与该发现一致，两种C3蛋白仅在酸性条件（pH < 5.5）下在脂质双层膜中表现出膜活性。总之，我们确定了巨噬细胞/单核细胞作为梭菌C3转移酶的靶细胞，并阐明了它们的选择性摄取机制，这可能有助于理解C3转移酶在发病机制中的作用。
The C3 transferases from Clostridium botulinum (C3bot) and Clostridium limosum (C3lim) mono-ADP-ribosylate and thereby inactivate RhoA, -B and -C of eukaryotic cells. Due to their extremely poor cellular uptake, C3 transferases were supposed to be exoenzymes rather than exotoxins, challenging their role in pathogenesis. Here, we report for the first time that low concentrations of both C3lim and C3bot are selectively internalized into macrophages/monocytes in less than 3 h, inducing the reorganization of the actin cytoskeleton by ADP-ribosylation of Rho. We demonstrate that C3 transferases are internalized into the cytosol of macrophages/monocytes via acidified early endosomes. Bafilomycin A1, an inhibitor of endosomal acidification, protected J774A.1 macrophages and human promyelotic leukaemia cells (HL-60) from intoxication by C3. Moreover, confocal laser scanning microscopy revealed colocalization of C3 with early endosomes. An extracellular acidic pulse enabled direct translocation of cell surface-bound C3 across the cytoplasmic membrane to the cytosol. In line with this finding, both C3 proteins exhibited membrane activity in lipid bilayer membranes only under acidic conditions (pH < 5.5). In conclusion, we identified macrophages/monocytes as target cells for clostridial C3 transferases and shed light on their selective uptake mechanism, which might contribute to understand the role of C3 transferases in pathogenesis.