title: 4-Coumaroyl and Caffeoyl Shikimic Acids Inhibit 4-Coumaric Acid: Coenzyme A Ligases and Modulate Metabolic Flux for 3-Hydroxylation in Monolignol Biosynthesis of Populus trichocarpa.
abstract: In a number of plant species, downregulation of 4-coumaric acid: coenzyme A ligase (4CL) can reduce lignin content. In lignin precursor (monolignol) biosynthesis during stem wood formation in Populus trichocarpa, two enzymes Ptr4CL3 and Ptr4CL5 catalyze the CoA ligation of 4-coumaric acid to 4-coumaroyl-CoA and caffeic acid to caffeoyl-CoA. CoA ligation of 4-coumaric acid is essential for the 3-hydroxylation of 4-coumaroyl shikimic acid. This hydroxylation results from sequential reactions of 4-hydroxycinnamoyl-CoA: shikimic acid hydroxycinnamoyl transferases (PtrHCT1 and PtrHCT6) and 4-coumaric acid 3-hydroxylase 3 (PtrC3H3). Alternatively, 3-hydroxylation of 4-coumaric acid to caffeic acid may occur through an enzyme complex of cinnamic acid 4-hydroxylase 1 and 2 (PtrC4H1 and PtrC4H2) and PtrC3H3. We found that 4-coumaroyl and caffeoyl shikimic acids are inhibitors of Ptr4CL3 and Ptr4CL5. 4-Coumaroyl shikimic acid strongly inhibits formation of 4-coumaroyl-CoA and caffeoyl-CoA. Caffeoyl shikimic acid inhibits only formation of 4-coumaroyl-CoA. 4-coumaroyl and caffeoyl shikimic acids both act as competitive and uncompetitive inhibitors. Estimates of metabolic flux in wildtype and PtrC3H3 downregulated P. trichocarpa transgenics have been made using LC-MS/MS based absolute protein and metabolite quantification, mass action kinetics and inhibition equations. Inhibition by 4-coumaroyl and caffeoyl shikimic acids may play significant regulatory roles when these inhibitors accumulate.
authors.affiliation: State Key Laboratory of Tree Genetics and Breeding, Northeast Forestry University, Harbin 150040, China Forest Biotechnology Group, Department of Forestry and Environmental Resources, North Carolina State University, Raleigh, North Carolina 27695, U.S.A.
authors.forename: Chien-Yuan
authors.initials: CY
authors.lastname: Lin
authors.affiliation: State Key Laboratory of Tree Genetics and Breeding, Northeast Forestry University, Harbin 150040, China Forest Biotechnology Group, Department of Forestry and Environmental Resources, North Carolina State University, Raleigh, North Carolina 27695, U.S.A.
authors.forename: Jack P
authors.initials: JP
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Tree Genetics and Breeding, Northeast Forestry University, Harbin 150040, China State Key Laboratory of Tree Genetics and Breeding, Chinese Academy of Forestry, Beijing, 100091, China.
authors.forename: Quanzi
authors.initials: Q
authors.lastname: Li
authors.affiliation: Forest Biotechnology Group, Department of Forestry and Environmental Resources, North Carolina State University, Raleigh, North Carolina 27695, U.S.A.
authors.forename: Hsi-Chuan
authors.initials: HC
authors.lastname: Chen
authors.affiliation: Forest Biotechnology Group, Department of Forestry and Environmental Resources, North Carolina State University, Raleigh, North Carolina 27695, U.S.A.
authors.forename: Jie
authors.initials: J
authors.lastname: Liu
authors.affiliation: W.M. Keck FT-ICR Mass Spectrometry Laboratory, Department of Chemistry, North Carolina State University, Raleigh, North Carolina 27695, U.S.A.
authors.forename: Philip
authors.initials: P
authors.lastname: Loziuk
authors.affiliation: Department of Electrical and Computer Engineering, North Carolina State University, Raleigh North Carolina, 27695, U.S.A.
authors.forename: Jina
authors.initials: J
authors.lastname: Song
authors.affiliation: Department of Electrical and Computer Engineering, North Carolina State University, Raleigh North Carolina, 27695, U.S.A.
authors.forename: Cranos
authors.initials: C
authors.lastname: Williams
authors.affiliation: W.M. Keck FT-ICR Mass Spectrometry Laboratory, Department of Chemistry, North Carolina State University, Raleigh, North Carolina 27695, U.S.A.
authors.forename: David C
authors.initials: DC
authors.lastname: Muddiman
authors.affiliation: Forest Biotechnology Group, Department of Forestry and Environmental Resources, North Carolina State University, Raleigh, North Carolina 27695, U.S.A ron_sederoff@ncsu.edu.
authors.forename: Ronald R
authors.initials: RR
authors.lastname: Sederoff
authors.affiliation: State Key Laboratory of Tree Genetics and Breeding, Northeast Forestry University, Harbin 150040, China Forest Biotechnology Group, Department of Forestry and Environmental Resources, North Carolina State University, Raleigh, North Carolina 27695, U.S.A vchiang@ncsu.edu.
authors.forename: Vincent L
authors.initials: VL
authors.lastname: Chiang
keywords: 4-coumaroyl shikimic acid; LC-MS/MS; Populus trichocarpa; caffeoyl shikimic acid; metabolic flux; monolignol biosynthesis; reaction and inhibition kinetics