Targeted down-regulation of cytochrome P450 enzymes for forage quality improvement in alfalfa (Medicago sativa L.)

^*Plant Biology Division and ^‡Agriculture Division, Samuel Roberts Noble Foundation, 2510 Sam Noble Parkway, Ardmore, OK 73401

Edited by Rodney B. Croteau, Washington State University, Pullman, WA, and approved September 27, 2005 (received for review July 8, 2005)

Abstract

Improving the digestibility of forages provides a means to enhance animal performance and protect the environment against excessive animal waste. Increased lignin content during maturity, and corresponding changes in lignin composition, correlate with decreased digestibility of forages. These relationships have yet to be investigated in isogenic systems. By targeting three specific cytochrome P450 enzymes of the lignin pathway for antisense down-regulation, we generated transgenic alfalfa lines with a range of differences in lignin content and composition. There was a strong negative relationship between lignin content and rumen digestibility, but no relationship between lignin composition and digestibility, in these transgenic lines. Models for genetic manipulation of forage digestibility based on the changes in lignin composition that increase paper-pulping efficiency in trees are therefore invalid. Down-regulation of 4-coumarate 3-hydroxylase provided the largest improvements in digestibility yet seen in a forage crop.

Footnotes

↵ § To whom correspondence should be addressed. E-mail: radixon{at}noble.org.
↵ † M.S.S.R. and F.C. contributed equally to this work.
Author contributions: M.S.S.R. and R.A.D. designed research; M.S.S.R., F.C., G.S., L.J., and H.A. performed research; F.C. and H.A. contributed new reagents/analytic tools; M.S.S.R., F.C., and R.A.D. analyzed data; and R.A.D. wrote the paper.
Conflict of interest statement: No conflicts declared.
This paper was submitted directly (Track II) to the PNAS office.
Abbreviations: ADL, acid detergent lignin; C3H, coumarate 3-hydroxylase; C4H, cinnamate 4-hydroxylase; COMT, caffeic acid 3-O-methyltransferase; CCoAOMT, caffeoyl CoA 3-O-methyltransferase; F5H, ferulate 5-hydroxylase; G, guaiacyl; H, p-hydroxyphenyl; NDF, neutral detergent fiber; S, syringyl.
Data deposition: The alfalfa F5H cDNA sequences reported in this paper have been deposited in the GenBank database [accession nos. DQ222911 (F5H-K1) and DQ222912 (F5H-K10)].