During embryogenesis of eudicots, the apical region of the embryo develops two cotyledon primordia and the shoot meristem. In Arabidopsis thaliana, this process is dependent on the functionally redundant activities of the CUP-SHAPED COTYLEDON (CUC) transcription factors, namely CUC1, CUC2, and CUC3, as well as the phytohormone auxin. However, the relationship between the CUC proteins and auxin has yet to be fully elucidated. In the present study, we examined whether the expression of auxin biosynthetic genes is dependent on CUC gene activities. Comprehensive quantitative RT-PCR analysis of the main auxin biosynthetic gene families of TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS1/TRYPTOPHAN AMINOTRANSFERASE RELATED and YUCCA (YUC) showed that YUC1 and YUC4 expression levels were lower in cuc double mutant embryos than the expression levels of these genes in wild type embryos. Reporter analysis also revealed that the expression of YUC1 and YUC4 in the cotyledon boundary region was reduced in cuc double mutant embryos. In contrast, the loss of function mutation in the SHOOT MERISTEMLESS gene, a shoot stem cell regulator that acts downstream of the CUC genes, did not markedly affect YUC1 expression levels. These results demonstrate that CUC genes play an important role in the regulation of auxin biosynthetic gene expression during embryogenesis; furthermore, they raise the possibility that the auxin produced by this regulation contributes to cotyledon boundary development.