We identified three JAM-type genes, CrJAM2, CrJAM3, and CrRMT1 in C. roseus. In a transient expression screen, the overexpression of CrJAM2 and CrJAM3 repressed several promoters of enzymes and TFs involved in TIA biosynthesis. Similarly, we observed the stable overexpression of CrJAM2 and CrJAM3 in hairy root cultures repressed the basal expression of most enzymes and transcriptional activators involved in TIA biosynthesis. To increase TIA biosynthesis, we used CRISPR/Cas9 to introduce mutations in the native CrJAMs in C. roseus hairy root cultures. CRISPR/Cas9 mediated gene editing was extremely efficient in C. roseus hairy roots, resulting in 100% mutations in the native CrJAMs at every guide location. This was the first time CRISPR/Cas9 was reported in C. roseus. These mutant lines exhibited increased basal expression of most TIA biosynthetic genes. Our results also suggest that the CrJAM family may be the missing player in the regulatory triad with the activator MYC2a and the JAZ repressor family that confers the tight regulation of TIA biosynthesis observed in the literature. These results highlight the role of CrJAMs as repressors of TIA biosynthesis in C. roseus and of CRISPR/Cas9 as a strategy for unJAMming the tight regulation of TIA biosynthesis and potentially of other plant specialized metabolic pathways.