Hydrogen sulfide (H2S) is a gasotransmitter that regulates multiple cellular processes, such as carbon metabolism, endoplasmic reticulum stress response, redox homeostasis, and cellular survival. Saccharomyces cerevisiae is one of the few organisms capable of producing H2S through both major pathways-sulfur assimilation and transsulfuration-making it a unique model to study the cellular impact of H2S. Because this compound can be lethal at high concentrations by inhibiting cellular respiration, its production must be tightly regulated. However, at physiological levels, H2S can regulate protein activity, localization, and function, as well as protect against oxidative stress. The mechanism through which this compound acts on proteins is S-persulfidation, a post-translational modification (PTM) on cysteine residues characterized by the formation of a disulfide-like bond on cysteine thiols. Research on S-persulfidation has progressed thanks to techniques capable of distinguishing it from other oxidative states in cysteine residues. Proteins involved in carbon metabolism (such as GAPDH and Pyk1), amino acid biosynthesis (for example, Cys4), the translation machinery, and stress-response pathways have been shown to undergo S-persulfidation, and these cysteines play an important regulatory role. Despite these advances, information on S. cerevisiae remains limited, leaving this field with many possibilities for discovering new processes regulated by H2S and S-persulfidation.