The yeast Ste2p G protein-coupled receptor dimerizes on the cell plasma membrane

Abstract

Dimerization of G protein-coupled receptors (GPCR) may play an important role in maturation, internalization, signaling and/or pharmacology of these receptors. However, the location where dimerization occurs is still under debate. In our study, variants of Ste2p, a yeast mating pheromone GPCR, were tagged with split EGFP (enhanced green fluorescent protein) fragments inserted between transmembrane domain seven and the C-terminus or appended to the C-terminus. Bimolecular Fluorescence Complementation (BiFC) assay was used to determine where receptor dimerization occurred during protein trafficking by monitoring generation of EGFP fluorescence, which occurred upon GPCR dimerization. Our results suggest that these tagged receptors traffic to the membrane as monomers, undergo dimerization or higher ordered oligomerization predominantly on the plasma membrane, and are internalized as dimers/oligomers. This study is the first to provide direct in vivo visualization of GPCR dimerization/oligomerization, during trafficking to and from the plasma membrane.

Mating yeast cells use an intrinsic polarity site to assemble a pheromone-gradient tracking machine

“…S3 and S4 and Tables S3, S4, S5, S6, and S7). (1) The receptor is dimeric (Overton and Blumer, 2000;Gehret et al, 2012;Sridharan et al, 2016;Cevheroglu et al, 2017), and gradientstimulation generates three species: the active-active homodimer (AA), the inactive-active heterodimer (IA), and the inactive-inactive homodimer (II). This allows for pheromoneinduced internalization of the G protein with the receptor (Suchkov et al, 2010) without violating the GPCR-G protein trafficking paradigm, i.e., that activation of GPCRs triggers their internalization while heterotrimeric G proteins stably interact only with inactive receptors.…”

“…To allow for pheromoneinduced internalization of the G protein with the receptor (Suchkov et al, 2010) without violating the GPCR-G protein trafficking paradigm, pheromone receptors were modeled as dimers. The existence of both Ste2 homo-and heterooligomers has been well established (Overton and Blumer, 2000;Sridharan et al, 2016;Cevheroglu et al, 2017). Pheromone (L, ligand)-receptor association and dissociation generate a dynamic population of inactive, partially active, and fully active dimers (II, IA, and AA, respectively).…”

Regulation of G protein‐coupled receptor signaling by plasma membrane organization and endocytosis

“…Heteromers such as the μ‐/κ‐opioid receptor (μOR, κOR) dimer might couple to different effectors and induce functional effects distinct from their monomers . Receptors can oligomerize at multiple steps throughout the biosynthetic trafficking of GPCRs, with the γ‐aminobutyric acid receptor type B (GABABR) requiring dimerization for endoplasmic reticulum (ER) export whereas Ste2p dimerizes only at the plasma membrane …”

“…[33][34][35][36] Receptors can oligomerize at multiple steps throughout the biosynthetic trafficking of GPCRs, with the γ-aminobutyric acid receptor type B (GABABR) requiring dimerization for endoplasmic reticulum (ER) export 37,38 whereas Ste2p dimerizes only at the plasma membrane. 39 Receptor oligomerization regulates the diffusion of the receptors within the plasma membrane. The GABAB receptor, an obligate dimer, 23,24 diffuses slowly within the plasma membrane and primarily exists as dimers and tetramers.…”

Cell Communications among Microorganisms, Plants, and Animals: Origin, Evolution and Interplays

“…The membrane receptors in bacteria often directly respond to nutrients [11], or to Quorum Sensing (QS) signaling molecules [12], and those in yeasts can besides respond to pheromones [13] cytoplasmic transcription factors (QseB) which direct alterations in gene expression [12,25]. The QseC receptor responds to AI-3 but also to AI-2 and, additionally, to epinephrine/ norepinephrine from eukaryotic hosts.…”