SMAP2 Regulates Retrograde Transport from Recycling Endosomes to the Golgi

Tatsuyuki Matsudaira,Toshio Watanabe,Tomohiko Taguchi,Shunsuke Kon,Hiroyuki Arai,Yasunori Uchida,Kenji Tanabe,Christophe Lamaze

doi:10.1371/journal.pone.0069145

Abstract

Retrograde transport is where proteins and lipids are transported back from the plasma membrane (PM) and endosomes to the Golgi, and crucial for a diverse range of cellular functions. Recycling endosomes (REs) serve as a sorting station for the retrograde transport and we recently identified evection-2, an RE protein with a pleckstrin homology (PH) domain, as an essential factor of this pathway. How evection-2 regulates retrograde transport from REs to the Golgi is not well understood. Here, we report that evection-2 binds to SMAP2, an Arf GTPase-activating protein. Endogenous SMAP2 localized mostly in REs and to a lesser extent, the trans-Golgi network (TGN). SMAP2 binds evection-2, and the RE localization of SMAP2 was abolished in cells depleted of evection-2. Knockdown of SMAP2, like that of evection-2, impaired the retrograde transport of cholera toxin B subunit (CTxB) from REs. These findings suggest that evection-2 recruits SMAP2 to REs, thereby regulating the retrograde transport of CTxB from REs to the Golgi.

Highlights

Synthesized proteins that are destined for secretion or for residence within organelles move from the endoplasmic reticulum (ER), through the Golgi, to their final destination [1]
Among several combinations of fixation and permeabilization reagents tested, we found that fixation with trichloroacetic acid (TCA) and permeabilization with saponin or Triton X-100 yielded a decent signal of SMAP2
We identified a second protein SMAP2, an Arf GAP, which functions in the retrograde transport of cholera toxin B subunit (CTxB) as evection-2

Summary

Introduction

Synthesized proteins that are destined for secretion or for residence within organelles move from the endoplasmic reticulum (ER), through the Golgi, to their final destination [1]. This membrane outflow is counteracted by retrograde membrane flow that originates from either PM or endosomal system [2,3]. In cells depleted of evection-2, the retrograde transport of CTxB to the Golgi was impaired in REs, and the Golgi localization of TGN46 and GP73 was abolished. The molecular mechanism of how evection-2 regulates retrograde transport is not well understood

Methods

Results

Conclusion