Marchantia polymorpha has a unique phenylpropanoid biosynthetic pathway for producing bis(bibenzyl) compounds, such as marchantins. Previous tracer studies have suggested that dihydro-p-coumaroyl-CoA, an intermediate in the M. polymorpha bis(bibenzyl) biosynthetic pathway, is derived from dihydro-p-coumaric acid, which may result from the reduction of p-coumaric acid. However, the enzymes responsible for reducing the α,β-unsaturated double bond in p-coumaric acid have not yet been identified in bryophytes or in vascular plants. To clarify the biosynthetic pathway to dihydro-p-coumaroyl-CoA in M. polymorpha, the present study characterized a 4-coumarate:CoA ligase (Mp4CL3) and a double-bond reductase (MpDBR1), both of which were upregulated in M. polymorpha line overexpressing MpMYB2 with high accumulation of bis(bibenzyl) compounds. Enzyme assays using Escherichia coli showed that Mp4CL3 has the esterification activity toward both p-coumaric acid and dihydro-p-coumaric acid with producing their corresponding CoA thioesters, whereas the reduction activity of MpDBR1 was specific to p-coumaroyl-CoA, but not to p-coumaric acid. CRISPR/Cas9-mediated knock out of MpDBR1 resulted in a significant reduction in bis(bibenzyl) content. These findings provide clear evidence for a biosynthetic route from p-coumaric acid to dihydro-p-coumaroyl-CoA via p-coumaroyl-CoA in the bis(bibenzyl) biosynthetic pathway of M. polymorpha.
Kobayashi et al. (Mon,) studied this question.