Specific plant species produce unique isoquinoline alkaloids (IQAs); however, the mechanism of their evolution and the regulation of their biosynthesis are largely unknown. We report here the isolation of a novel basic helix-loop-helix protein, CjbHLH1, from IQA-producing Coptis japonica. A BLAST search indicated that CjbHLH1 homologs were only found in plant species that produce IQAs. Transient RNA interference (RNAi) and overexpression of CjbHLH1 in C. japonica protoplasts revealed the activity of CjbHLH1 in transcription of IQA biosynthetic genes, and little activity in the transcription of genes involved in primary metabolism or the stress response. A chromatin immunoprecipitation experiment using CjbHLH1-specific antibodies revealed the direct interaction of CjbHLH1 with promoter sequences of IQA biosynthetic genes in vivo. We discuss the unique role of CjbHLH1 in IQA biosynthesis.