Characterizing the Role of Yeast Cyclin Pcl1 in the Establishment of Pheromone Receptor Polarity

Chemotaxis (directed cell movement) and chemotropism (directed cell growth) are vital mechanisms essential to a variety of biological processes, including cell development, cancer metastasis, angiogenesis, and axon guidance. Both chemotaxis and chemotropism depend on the ability of the cell to interpret shallow and complex chemical gradients. One of the best‐studied models of eukaryotic directional sensing is the mating response of Saccharomyces cerevisiae (budding yeast). Yeast exist as two haploid mating types, MAT a and MAT α, that can sense pheromone gradients, chemotrop toward the closest mating partner, and eventually fuse at their tips to form diploid cells. In vegetative cells, the G protein coupled receptor (GPCR) that binds pheromone and its associated heterotrimeric G protein are uniformly distributed on the plasma membrane. Upon receptor activation, Gβγ is released from Gα and Gβ is rapidly phosphorylated on multiple residues. Free Gβγ signals through a MAP kinase cascade to trigger cell‐cycle arrest and changes in gene expression. Pheromone‐activated receptors are globally internalized and then reappear as polarized crescents that mark the chemotropic growth site, where the cell ultimately forms its mating projection. Previous studies have shown that the receptor crescents are visible before detectable actin polarization and that the receptors can polarize without actin‐dependent directed secretion [1]. On the contrary, inhibiting receptor internalization confers a moderate defect in gradient sensing [2]. A key question raised by these observations is how does the yeast cell interpret the external signals and establish the chemotropic growth site? From a directed genetic screen, we found that Pcl1 is critical for the establishment of receptor polarity. Pcl1 is a cyclin required by the cyclin dependent kinase Pho85, which has been implicated in polarization during budding. It is known that Gβ phosphorylation is crucial to receptor polarization, and Gβ is predicted to be a substrate of Pho85‐Pcl1 [3]. Additionally, our data indicated a genetic interaction between Gβ and Pcl1. Here, we show that Pcl1 localizes to the mating projection during the mating response. Moreover, Pcl1 and Gβ interacted directly on the plasma membrane of pheromone‐treated cells, as indicated by a Bimolecular fluorescence complementation assay. Lastly, Pho85 inactivation also appears to affect receptor polarity. Taken together, these results suggest that Pho85‐Pcl1 regulates polarization of the receptor by phosphorylating Gβ. Support or Funding Information National Science Foundation