Syntaxin 16 and syntaxin 5 are required for efficient retrograde transport of several exogenous and endogenous cargo proteins

Abstract

Retrograde transport allows proteins and lipids to leave the endocytic pathway to reach other intracellular compartments, such as TGN/Golgi membranes, the endoplasmic reticulum, and in some instances, the cytosol. Here, we have used RNA interference against the SNARE proteins syntaxin 5 and syntaxin 16 combined with recently developed quantitative trafficking assays, morphological approaches, and cell intoxication analysis to show that these SNARE proteins are not only required for efficient retrograde transport of Shiga toxin, but also for that of an endogenous cargo protein, the mannose 6-phosphate receptor, and for the productive trafficking into cells of cholera toxin and ricin. We have found that the function of syntaxin 16 was specifically required for, and restricted to the retrograde pathway. Strikingly, syntaxin 5 RNA interference protected cells particularly strongly against Shiga toxin. Since our trafficking analysis showed that apart from inhibiting retrograde endosomes-to-TGN transport, the silencing of syntaxin 5 had no additional effect on Shiga toxin endocytosis or trafficking from TGN/Golgi membranes to the endoplasmic reticulum, we hypothesize that syntaxin 5 also has trafficking-independent functions. In summary, our data demonstrate that several cellular and exogenous cargo proteins use elements of the same SNARE machinery for efficient retrograde transport between early/recycling endosomes and TGN/Golgi membranes.

Asna1/TRC40 Controls β-Cell Function and Endoplasmic Reticulum Homeostasis by Ensuring Retrograde Transport

“…Although the molecular targets of Retro-2 are unknown and at what point the Retro-2 and Asna1 pathways intersect is unclear, the reduction of Stx5 in the cis Golgi and Stx6 in the TGN is likely to affect Asna1-dependent retrograde transport. Small interfering RNA-mediated knockdown of Stx5 and inhibition of Stx6 using blocking antibodies have both been shown to negatively affect EE-to-TGN recycling (35,36). However, two observations argue for a primary role for Stx5 in the context of retrograde transport and ER homeostasis.…”

Lack of the endosomal SNAREs vti1a and vti1b led to significant impairments in neuronal development

“…S9A), even though we did not observe differences in MPR steady-state distribution. Return of MPR to the TGN depends on vti1a and its partner syntaxin16 in vitro (10,17,30).…”

The Golgi apparatus: insights from filamentous fungi

“…The TlgB-labeled late Golgi compartment most possibly receives endosomal traffic: Tlg2, yeast's homolog of the A. nidulans TlgB, and its mammalian homolog, syntaxin 16, both have been implicated in the formation of SNARE bundles regulating fusion with the TGN of vesicles derived from the endosomes (Abeliovich et al 1998, Amessou et al 2007). Association of TlgB with the SNARES Vti1, TlgA and SynA observed with A. nidulans proteins (López-Berges et al 2016) is consistent with such a role.…”