Regulation of the divalent metal ion transporter via membrane budding.

Kimberlyd Mackenzie,Sushma Anand,Hazel E Dalton,Natasha Chaudhary,Brett M Collins,Sharad Kumar,Natalie J Foot,Suresh Mathivanan

doi:10.1038/celldisc.2016.11

Abstract

The release of extracellular vesicles (EVs) is important for both normal physiology and disease. However, a basic understanding of the targeting of EV cargoes, composition and mechanism of release is lacking. Here we present evidence that the divalent metal ion transporter (DMT1) is unexpectedly regulated through release in EVs. This process involves the Nedd4-2 ubiquitin ligase, and the adaptor proteins Arrdc1 and Arrdc4 via different budding mechanisms. We show that mouse gut explants release endogenous DMT1 in EVs. Although we observed no change in the relative amount of DMT1 released in EVs from gut explants in Arrdc1 or Arrdc4 deficient mice, the extent of EVs released was significantly reduced indicating an adaptor role in biogenesis. Furthermore, using Arrdc1 or Arrdc4 knockout mouse embryonic fibroblasts, we show that both Arrdc1 and Arrdc4 are non-redundant positive regulators of EV release. Our results suggest that DMT1 release from the plasma membrane into EVs may represent a novel mechanism for the maintenance of iron homeostasis, which may also be important for the regulation of other membrane proteins.

Highlights

The regulation of plasma membrane proteins is critical for maintaining cellular homeostasis and for the regulation of signaling responses
Arrdc1 or Arrdc4 overexpression significantly decreased DMT1 transport activity, whereas small interfering RNA-mediated knockdown of endogenous Arrdc1 (490% knockdown) lead to an increase in DMT1 activity compared with the control (Figure 1A)
This effect on DMT1 is specific to Arrdc1 and Arrdc4 as other α-arrestin family members (Arrdc3 and TXNIP) did not alter DMT1 activity (Supplementary Figure S1)

Summary

Introduction

The regulation of plasma membrane proteins is critical for maintaining cellular homeostasis and for the regulation of signaling responses. These data show that DMT1 trafficking into EVs is dependent, at least partially, on the recruitment of Nedd4 E3 ligases and the plasma membrane localization of Arrdc1 and Arrdc4.

Results

Conclusion