Skip to Content
Merck
  • Transport of the cholera toxin B-subunit from recycling endosomes to the Golgi requires clathrin and AP-1.

Transport of the cholera toxin B-subunit from recycling endosomes to the Golgi requires clathrin and AP-1.

Journal of cell science (2015-07-03)
Tatsuyuki Matsudaira, Takahiro Niki, Tomohiko Taguchi, Hiroyuki Arai
ABSTRACT

The retrograde pathway is defined by the transport of proteins and lipids from the plasma membrane through endosomes to the Golgi complex, and is essential for a variety of cellular activities. Recycling endosomes are important sorting stations for some retrograde cargo. SMAP2, a GTPase-activating protein (GAP) for Arf1 with a putative clathrin-binding domain, has previously been shown to participate in the retrograde transport of the cholera toxin B-subunit (CTxB) from recycling endosomes. Here, we found that clathrin, a vesicle coat protein, and clathrin adaptor protein complex 1 (AP-1) were present at recycling endosomes and were needed for the retrograde transport of CTxB from recycling endosomes to the Golgi, but not from the plasma membrane to recycling endosomes. SMAP2 immunoprecipitated clathrin and AP-1 through a putative clathrin-binding domain and a CALM-binding domain, and SMAP2 mutants that did not interact with clathrin or AP-1 could not localize to recycling endosomes. Moreover, knockdown of Arf1 suppressed the retrograde transport of CTxB from recycling endosomes to the Golgi. These findings suggest that retrograde transport is mediated by clathrin-coated vesicles from recycling endosomes and that the role of the coat proteins is in the recruitment of Arf GAP to transport vesicles.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Dimethyl sulfoxide, anhydrous, ≥99.9%
Sigma-Aldrich
Dimethyl sulfoxide, ≥99.6%, ReagentPlus®
Sigma-Aldrich
Dimethyl sulfoxide, for molecular biology
Sigma-Aldrich
Dimethyl sulfoxide, PCR Reagent
Sigma-Aldrich
Dimethyl sulfoxide, ≥99.5% (GC), suitable for plant cell culture
Sigma-Aldrich
Dimethyl sulfoxide, BioUltra, for molecular biology, ≥99.5% (GC)
Sigma-Aldrich
Dimethyl sulfoxide, sterile-filtered, BioPerformance Certified, meets EP, USP testing specifications, suitable for hybridoma
Sigma-Aldrich
Dimethyl sulfoxide, meets EP testing specifications, meets USP testing specifications
Sigma-Aldrich
Ethidium bromide solution, BioReagent, for molecular biology, 500 μg/mL in H2O
Sigma-Aldrich
Ethidium bromide solution, BioReagent, for molecular biology, 10 mg/mL in H2O
Sigma-Aldrich
MISSION® esiRNA, targeting mouse Clint1
Sigma-Aldrich
MISSION® esiRNA, targeting human PLEKHB2
Supelco
Pyridaben, PESTANAL®, analytical standard
Sigma-Aldrich
Ethidium bromide solution, for fluorescence, ~1% in H2O
Sigma-Aldrich
MISSION® esiRNA, targeting human SMAP2
Sigma-Aldrich
MISSION® esiRNA, targeting human SMOC2
Sigma-Aldrich
MISSION® esiRNA, targeting mouse Golim4
Sigma-Aldrich
MISSION® esiRNA, targeting mouse Smoc2
Sigma-Aldrich
MISSION® esiRNA, targeting human ARF1
Sigma-Aldrich
Dimethyl sulfoxide, Hybri-Max, sterile-filtered, BioReagent, suitable for hybridoma, ≥99.7%
Sigma-Aldrich
MISSION® esiRNA, targeting mouse Arf1
Sigma-Aldrich
MISSION® esiRNA, targeting human BRD8
Sigma-Aldrich
MISSION® esiRNA, targeting mouse Brd8
Sigma-Aldrich
Ethidium bromide, ~95% (HPLC)
Sigma-Aldrich
Ethidium bromide, BioReagent, for molecular biology, powder