Characterization and expression profiling of cinnamate 4-hydroxylase gene from Salvia miltiorrhiza in rosmarinic acid biosynthesis pathway

A novel cinnamate 4-hydroxylase (C4H) gene (designated as SmC4H) involved in the rosmarinic acid biosynthesis pathway is cloned from Salvia miltiorrhiza. The full-length cDNA of SmC4H is 1800 bp long with an open reading frame of 1512 bp encoding a polypeptide of 504 amino acid residues. Like other C4Hs, the predicted SmC4H polypeptide includes three domains: a heme-binding domain, a proline-rich region, and a P450 (E)EFRPER-motif region, which represents a typical structure of plant C4Hs. Analysis of SmC4H genomic DNA reveals that it contains 3 exons, 2 introns, the length of exons being highly conserved. Semiquantitative RT-PCR analysis revealed that the constitutive expression of SmC4H in the root or stem was much higher than in the leaf. Further expression analysis revealed that the signaling components of defense/stress pathways, such as methyl jasmonate, abscisic acid, and ultraviolet-B radiation, up-regulated the SmC4H transcript levels over the control. Calcium chloride and hydrogen peroxide, nevertheless, had no significant effect on SmC4H expression.