GPCR receptor phosphorylation and endocytosis are not necessary to switch polarized growth between internal cues during pheromone response in S. cerevisiae

GUSTAVO VASEN; PAULA DUNAYEVICH; ANDREAS CONSTANTINOU; ALEJANDRO COLMAN LERNER

Communicative and Integrative Biology

Landes Bioscience

Año: 2020

1942-0889

Chemotactic/chemotropic cells follow accurately the direction of gradients of regulatory molecules. Many G-protein-coupled receptors (GPCR) function as chemoattractant receptors to guide polarized responses. In ?a? mating type yeast, the GPCR Ste2 senses the -cell?s pheromone. Previously, phosphorylation and trafficking of this receptor has been implicated in the process of gradient sensing, where cells dynamically correct growth. Correction is often necessary since yeast have intrinsic polarity sites that interfere with a correct initial gradient decoding. We have recently showed (Vasen et al., 2020) that when actively dividing (not in G1) yeast are exposed to a uniform pheromone concentration, they initiate a pheromone-induced polarization next to the mother-daughter cytokinesis site. Then, they reorient their growth to the intrinsic polarity site. Here, to study if Ste2 phosphorylation and internalization are involved in this process, we generated receptor variants combining 3 types of mutated signals for the first time: phosphorylation, ubiquitylation and the NPFX1,2D Sla1-binding motif. We first characterized their effect on endocytosis, and found that these processes regulate internalization in a more complex manner than previously shown. Interestingly, we showed that receptor phosphorylation can drive internalization independently of ubiquitylation and the NPFX1,2D motif. When tested in our assays, cells expressing either phosphorylation or endocytosis deficient receptors were able to switch away from the cytokinesis site to find the intrinsic polarity site as efficiently as their WT counterparts. Thus, we conclude that these processes are not necessary for the reorientation of polarization.