Yersinia pseudotuberculosis is a significant psychrotrophic food pathogen whose cold tolerance mechanisms are poorly understood. Signal transduction systems serve to monitor the environment, but no systematic investigation of their role at cold temperatures in Y. pseudotuberculosis has yet been undertaken. The relative expression levels of 54 genes predicted to encode proteins belonging to signal transduction systems in Y. pseudotuberculosis IP32953 were determined at 28°C and 3°C by quantitative real-time reverse transcription-PCR. The relative expression levels of 44 genes were significantly (p<0.05) higher at 3°C than at 28°C. Genes encoding the two-component system CheA/CheY had the highest relative expression levels at 3°C. Mutational analysis revealed that cheA is important for growth and motility at 3°C. The relative expression level of one gene, rssB, encoding an RpoS regulator, was significantly (p<0.05) lower at 3°C than at 28°C. The results suggest that several signal transduction systems might be used during growth at low temperature, and at least, CheA/CheY two-component system is important for low-temperature growth.