K‐29 linked ubiquitination of Arrdc4 regulates its function in extracellular vesicle biogenesis

Abstract

Extracellular vesicles (EVs) are important mediators of intercellular communication. However, EV biogenesis remains poorly understood. We previously defined a role for Arrdc4 (Arrestin domain containing protein 4), an adaptor for Nedd4 family ubiquitin ligases, in the biogenesis of EVs. Here we report that ubiquitination of Arrdc4 is critical for its role in EV secretion. We identified five potential ubiquitinated lysine residues in Arrdc4 using mass spectrometry. By analysing Arrdc4 lysine mutants we discovered that lysine 270 (K270) is critical for Arrdc4 function in EV biogenesis. Arrdc4 K270R mutation caused a decrease in the number of EVs released by cells compared to Arrdc4 WT , and a reduction in trafficking of divalent metal transporter (DMT1) into EVs. Furthermore, we also observed a decrease in DMT1 activity and an increase in its intracellular degradation in the presence of Arrdc4 K270R . K270 was found to be ubiquitinated with K‐29 polyubiquitin chains by the ubiquitin ligase Nedd4‐2. Thus, our results uncover a novel role of K‐29 polyubiquitin chains in Arrdc4‐mediated EV biogenesis and protein trafficking.

An insulin-regulated arrestin domain protein controls hepatic glucagon action

“…ARRDC4KO mice have previously been shown to have defective extracellular vesicle release and sperm maturation ( 11 , 39 , 40 ). Here, we further show that ARRDC4KO mice exhibit low fasting glucose levels and cannot generate sufficient glucose following overnight fasting.…”

Extracellular Vesicles for Therapeutic Nucleic Acid Delivery: Loading Strategies and Challenges

“…Subsequently, the abscission of ARRDC1-containing microvesicles appears to be regulated by the VPS4 ATPase [ 219 ]. In addition to ARRDC1, the ARRDC4 protein may participate in the regulation of microvesicle biogenesis [ 220 , 221 ]. However, its mechanism of action is most likely not related to ESCRTs [ 220 ].…”

Research progress on post-translational modification of proteins and cardiovascular diseases

“…Ubiquitinated K270 has been identified by mass spectrometry, confirming that the Arrdc4 adaptor of the Nedd4 family of ubiquitin ligases, plays an important role in extracellular vesicle biogenesis. It has also been shown that NEDD4-2 poly-ubiquitiation of K-29-linked Arrdc4 at K270 is a possible new signal for ARRDC4-mediated release of extracellular vesicles [ 21 ]. The N-methyl-D-aspartate (NMDA) glutamate receptor plays important roles in the development and storage of information in the mammalian brain.…”