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Although cell size regulation is crucial for cellular functions in a variety of organisms, from bacteria to humans, the underlying mechanisms remain elusive. Here, we identify Rim21, a component of the pH-sensing Rim101 pathway, as a positive regulator of cell size through a genome-wide screen of Saccharomyces cerevisiae deletion mutants. We found that mutants defective in the Rim101 pathway were consistently smaller than wild-type cells in the log and stationary phases. The expression of the active form of Rim101 increased the size of wild-type cells. Furthermore, the size of wild-type cells increased in response to external alkalization, which was associated with changes in both vacuolar and cytoplasmic volume. These volume changes were dependent on Rim21 and Rim101. A mutant lacking Vph1, a component of V-ATPase that is transcriptionally regulated by Rim101, was also smaller than wild-type cells, with no increase in size in response to alkalization. The loss of Vph1 suppressed the Rim101-induced increase in cell size under physiological pH conditions. Our results suggest that the cell size of budding yeast is regulated by the Rim101 and V-ATPase axis under physiological conditions as well as in response to alkaline stresses.