Functional dissection of the retrograde Shiga toxin trafficking inhibitor Retro-2

• Published in: Nature chemical biology Vol. 16; no. 3; pp. 327 - 336
• Main Authors: Forrester, Alison, Rathjen, Stefan J., Daniela Garcia-Castillo, Maria, Bachert, Collin, Couhert, Audrey, Tepshi, Livia, Pichard, Sylvain, Martinez, Jennifer, Munier, Mathilde, Sierocki, Raphael, Renard, Henri-François, Augusto Valades-Cruz, César, Dingli, Florent, Loew, Damarys, Lamaze, Christophe, Cintrat, Jean-Christophe, Linstedt, Adam D., Gillet, Daniel, Barbier, Julien, Johannes, Ludger
• Format: Journal Article
• Language: English
• Published: New York Nature Publishing Group US 01-03-2020; Nature Publishing Group
• Subjects: 631/326/41; 631/80/313; 631/92/555; 631/92/613; Anterograde transport; Benzamides - metabolism; Benzamides - pharmacology; Biochemical Engineering; Biochemistry; Biochemistry, Molecular Biology; Biological Transport; Bioorganic Chemistry; Cell Biology; Chemistry; Chemistry and Materials Science; Chemistry/Food Science; Endoplasmic reticulum; Endoplasmic Reticulum - drug effects; Endoplasmic Reticulum - metabolism; Endosomes; Endosomes - metabolism; Golgi apparatus; Golgi Apparatus - metabolism; HeLa Cells; Humans; Inhibitors; Life Sciences; Molecular modelling; Protein Transport; Proteins; Qa-SNARE Proteins - metabolism; Retrograde transport; Ricin; Ricin - metabolism; Shiga toxin; Shiga Toxin - metabolism; Shiga Toxins - metabolism; SNAP receptors; Syntaxin; Target recognition; Thiophenes - metabolism; Thiophenes - pharmacology; Toxins; Transport; Vesicular Transport Proteins - metabolism; Vesicular Transport Proteins - physiology; small molecule; Shiga toxin; retention using selective hooks; click chemistry; Shiga-like toxin; proximity ligation assay; Retrograde transport; STxB; syntaxin-5; SNARE; trans-Golgi network; COPII; Retro-2; GPP130; chemical biology; Sec16A; chemical genetics; mass spectrometry; anterograde trafficking
• ISSN: 1552-4450; 1552-4469
• DOI: 10.1038/s41589-020-0474-4

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.
The ER exit site component Sec16A was identified as the target of Retro-2, a small-molecule inhibitor of protein toxins and pathogens. Retro-2 treatment alters retrograde early/maturing endosomes-to-Golgi trafficking of Shiga toxin.