Temporal Proteomic Profiling of Pheromone‐Induced Cell Cycle Re‐Entry in <i>Saccharomyces cerevisiae</i>

The regulation of cell cycle progression in response to environmental cues is essential for cellular adaptation. In Saccharomyces cerevisiae, the BAR1 gene modulates sensitivity to the mating pheromone α-factor, which induces cell cycle arrest in G1. Here, we investigated the dynamic proteomic response in the bar1 deletion strain using a 27-plex experimental design with TMTproD isobaric labeling. Asynchronous bar1Δ cells were treated with α-factor and then released from the pheromone-induced cell cycle arrest in G1. Using higher-order TMTpro sample multiplexing, we generated global temporal profiles of protein abundance associated with recovery from this arrest, with triplicate samples collected at eight time points from 0 to 165 min after washing out the pheromone. We identify specific proteins involved in cell cycle re-entry and in the attenuation of the pheromone signal, providing insights into the regulatory mechanisms of mating response in yeast. This study also contributes significantly to dynamic proteomic analysis of cell cycle progression. We present a versatile approach for investigating complex cellular processes and showcase cell cycle progression following release from pheromone-induced arrest in yeast.