The Angiotensin II Type 1 Receptor C-Terminal Lys Residues Interact with Tubulin and Modulate Receptor Export Trafficking

Shu Zhu,Guangyu Wu,Catherine Faivre-Sarrailh,Matthew T. Duvernay,Xiaoping Zhang,Hong Wang

doi:10.1371/journal.pone.0057805

Abstract

The physiological and pathological functions of angiotensin II are largely mediated through activating the cell surface angiotensin II type 1 receptor (AT1R). However, the molecular mechanisms underlying the transport of newly synthesized AT1R from the endoplasmic reticulum (ER) to the cell surface remain poorly defined. Here we demonstrated that the C-terminus (CT) of AT1R directly and strongly bound to tubulin and the binding domains were mapped to two consecutive Lys residues at positions 310 and 311 in the CT membrane-proximal region of AT1R and the acidic CT of tubulin, suggestive of essentially ionic interactions between AT1R and tubulin. Furthermore, mutation to disrupt tubulin binding dramatically inhibited the cell surface expression of AT1R, arrested AT1R in the ER, and attenuated AT1R-mediated signaling measured as ERK1/2 activation. These data demonstrate for the first time that specific Lys residues in the CT juxtamembrane region regulate the processing of AT1R through interacting with tubulin. These data also suggest an important role of the microtubule network in the cell surface transport of AT1R.

Highlights

Angiotensin II (Ang II), an octapeptide hormone, modulates the physiological function of virtually all organs and plays an important role in the development of a number of human diseases such as diabetes, hypertension, myocardial infarction, congestive heart failure, and stroke
The CTs of Ang II type 1 receptor (AT1R), b2-AR and a2B-AR were generated as Glutathione S-transferase (GST) fusion proteins and their interaction with brain extracts and purified tubulin was determined by GST fusion protein pulldown assay
The AT1R CT interacted with tubulin from brain extracts and purified tubulin (Fig. 1B) and the amount of tubulin bound to the AT1R CT was significantly higher than that bound to the a2B-AR CT (Fig. 1C)

Summary

Introduction

Angiotensin II (Ang II), an octapeptide hormone, modulates the physiological function of virtually all organs and plays an important role in the development of a number of human diseases such as diabetes, hypertension, myocardial infarction, congestive heart failure, and stroke. Similar to many other GPCRs, the proper function of AT1R relies on the dynamic and highly regulated intracellular trafficking of the receptors, including anterograde export of newly synthesized receptors from the endoplasmic reticulum (ER) to the cell surface, Ang II-evoked internalization of the cell surface receptor, recycling of the internalized receptors from endosomes back to the cell surface, and targeting of the receptor to the degradation pathways. A number of studies have shown that the cell surface targeting of nascent AT1R is regulated by several regulatory proteins [5,6,7,8,9,10] and specific motifs in the different locations within the receptor [7,11,12,13,14]

Methods

Results

Conclusion