Terpenoid indole alkaloids (TIA) are important pharmaceuticals produced in low amounts through the terpenoid indole alkaloid pathway in Catharanthus roseus. Production of these alkaloids is controlled by biosynthetic and regulatory genes. CrMYC1 is a chief transcription factor that regulates terpenoid indole alkaloid production. In this study, we cloned CrMYC1 and overexpressed it in C. roseus hairy roots to investigate its effect on the production of the TIAs ajmalicine and catharanthine. qRT-PCR results revealed that CrMYC1 expression in transgenic CrMYC1-overexpressing hairy root lines was 3-6 times higher than that of the wild-type hairy root control line. Also, we detected a significant increase in strictosidine beta-D-glucosidase (SGD) expression, a critical enzyme in TIA biosynthesis, in the overexpression lines. HPLC analysis showed that CrMYC1 overexpression significantly improved the accumulation of ajmalicine (13 to 14-fold) and catharanthine (3 to 4.4-fold) compared to the control hairy root line. These results provide insight into the role of this understudied transcription factor in TIA pathway regulation and emphasize further research objectives.