The retrograde transport inhibitor Retro-2 has a protective effect on cells and in mice against Shiga-like toxins and ricin. Retro-2 causes toxin accumulation in early endosomes and relocalization of the Golgi SNARE protein syntaxin-5 to the endoplasmic reticulum. The molecular mechanisms by which this is achieved remain unknown. Here, we show that Retro-2 targets the endoplasmic reticulum exit site component Sec16A, affecting anterograde transport of syntaxin-5 from the endoplasmic reticulum to the Golgi. The formation of canonical SNARE complexes involving syntaxin-5 is not affected in Retro-2-treated cells. By contrast, the interaction of syntaxin-5 with a newly discovered binding partner, the retrograde trafficking chaperone GPP130, is abolished, and we show that GPP130 must indeed bind to syntaxin-5 to drive Shiga toxin transport from the endosomes to the Golgi. We therefore identify Sec16A as a druggable target and provide evidence for a non-SNARE function for syntaxin-5 in interaction with GPP130.

逆行转运抑制剂 Retro-2 对细胞和小鼠具有抗志贺样毒素和蓖麻毒素的保护作用。Retro-2 导致早期内体中的毒素积累和高尔基体 SNARE 蛋白 syntaxin-5 重新定位到内质网。实现这一目标的分子机制仍然未知。在这里，我们显示Retro-2 靶向内质网出口站点组件Sec16A，影响syntaxin-5 从内质网顺行运输到高尔基体。涉及syntaxin-5 的典型SNARE 复合物的形成在Retro-2 处理的细胞中不受影响。相比之下，syntaxin-5 与新发现的结合伙伴逆行运输伴侣 GPP130 的相互作用被废除，我们证明 GPP130 确实必须与 syntaxin-5 结合以驱动志贺毒素从内体转运到高尔基体。因此，我们将 Sec16A 确定为可药用靶标，并为 syntaxin-5 与 GPP130 相互作用的非 SNARE 功能提供证据。