Differential regulation of G protein signaling in Arabidopsis through two distinct pathways that internalize AtRGS1

Abstract

In animals, endocytosis of a seven-transmembrane GPCR is mediated by arrestins to propagate or arrest cytoplasmic G protein–mediated signaling, depending on the bias of the receptor or ligand, which determines how much one transduction pathway is used compared to another. In Arabidopsis thaliana, GPCRs are not required for G protein–coupled signaling because the heterotrimeric G protein complex spontaneously exchanges nucleotide. Instead, the seven-transmembrane protein AtRGS1 modulates G protein signaling through ligand-dependent endocytosis, which initiates derepression of signaling without the involvement of canonical arrestins. Here, we found that endocytosis of AtRGS1 initiated from two separate pools of plasma membrane: sterol-dependent domains and a clathrin-accessible neighborhood, each with a select set of discriminators, activators, and candidate arrestin-like adaptors. Ligand identity (either the pathogen-associated molecular pattern flg22 or the sugar glucose) determined the origin of AtRGS1 endocytosis. Different trafficking origins and trajectories led to different cellular outcomes. Thus, in this system, compartmentation with its associated signalosome architecture drives biased signaling.

Glucose sensing by regulator of G protein signaling 1 (RGS1) plays a crucial role in coordinating defense in response to environmental variation in tomato

“…Moreover, a study in Arabidopsis showed that flg22 initiated AtRGS1 endocytosis through clathrin‐mediated endocytosis (CME), whereas Glc activated both CME and sterol‐dependent endocytosis. Flg22 and Glc induced endocytosis of AtRGS1 from separate plasma membrane origins and had different downstream signaling pathways (Watkins et al ., 2021), implying that the two pathways do not conflict and may coexist in the plant. It should be noted that some plants, like grasses, lack homologous RGS proteins (Urano et al ., 2012, 2015).…”

G-Protein Phosphorylation: Aspects of Binding Specificity and Function in the Plant Kingdom

“…Flg22 induces AtRGS1 internalization via clathrin-mediated endocytosis (CME), while D-glucose triggers both CME and sterol-dependent endocytosis (SDE). The recruitment of the CME endocytic machinery towards GPCRs requires prior β-arrestin binding and activation, but plant genomes do not encode these proteins [ 55 , 56 ]. Nevertheless, Arabidopsis has three proteins with arrestin folds that bind as heterodimers to AtRGS1 and are required for endocytosis [ 55 ].…”

“…The recruitment of the CME endocytic machinery towards GPCRs requires prior β-arrestin binding and activation, but plant genomes do not encode these proteins [ 55 , 56 ]. Nevertheless, Arabidopsis has three proteins with arrestin folds that bind as heterodimers to AtRGS1 and are required for endocytosis [ 55 ]. These include the vacuolar sorting proteins 26 (VPS26)—AtVPS26a, AtVPS26b, and AtVPS26-like components of the retromer [ 57 ], well-known in animals for their role in endosomal to plasma membrane anterograde trafficking [ 58 ].…”

“…The candidate adaptor VPS26b forms a homodimer or a heterodimer with VPS26a, both required for flg22-mediated internalization of AtRGS1. However, those genes are not involved in AtRGS1 internalization that is induced by high concentrations of glucose [ 55 ]. Additionally, the inactivation of three cluster sites (AtRGS1 S428/435/436A ) completely abolishes flg22-induced internalization but only partially affects the glucose-mediated internalization of AtRGS1 [ 55 ].…”

“…However, those genes are not involved in AtRGS1 internalization that is induced by high concentrations of glucose [ 55 ]. Additionally, the inactivation of three cluster sites (AtRGS1 S428/435/436A ) completely abolishes flg22-induced internalization but only partially affects the glucose-mediated internalization of AtRGS1 [ 55 ]. Furthermore, a phosphatase is also required for AtRGS1 stability, and its presence reduces the in vitro identified phosphorylation by the WNKs [ 60 ].…”