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Plant Physiol, June 2001, Vol. 126, pp. 524-535
Flavonoids Act as Negative Regulators of Auxin Transport in
Vivo in Arabidopsis1
Dana E.
Brown,
Aaron M.
Rashotte,
Angus S.
Murphy,2
Jennifer
Normanly,
Brian W.
Tague,
Wendy A.
Peer,2
Lincoln
Taiz, and
Gloria
K.
Muday*
Department of Biology, Wake Forest University, Winston-Salem, North
Carolina 27109 (D.E.B., A.M.R., B.W.T., G.K.M.); Biology Department,
University of California, Santa Cruz, California 95064 (A.S.M., W.A.P.,
L.T.); and Department of Biochemistry and Molecular Biology, University
of Massachusetts, Amherst, Massachusetts 01003 (J.N.)
Polar transport of the plant hormone auxin controls many aspects of
plant growth and development. A number of synthetic compounds have been
shown to block the process of auxin transport by inhibition of the
auxin efflux carrier complex. These synthetic auxin transport inhibitors may act by mimicking endogenous molecules. Flavonoids, a
class of secondary plant metabolic compounds, have been suggested to be
auxin transport inhibitors based on their in vitro activity. The
hypothesis that flavonoids regulate auxin transport in vivo was tested
in Arabidopsis by comparing wild-type (WT) and transparent testa (tt4) plants with a mutation in the gene
encoding the first enzyme in flavonoid biosynthesis, chalcone synthase.
In a comparison between tt4 and WT plants, phenotypic
differences were observed, including three times as many secondary
inflorescence stems, reduced plant height, decreased stem diameter, and
increased secondary root development. Growth of WT Arabidopsis plants
on naringenin, a biosynthetic precursor to those flavonoids with auxin
transport inhibitor activity in vitro, leads to a reduction in root
growth and gravitropism, similar to the effects of synthetic auxin
transport inhibitors. Analyses of auxin transport in the inflorescence
and hypocotyl of independent tt4 alleles indicate that
auxin transport is elevated in plants with a tt4
mutation. In hypocotyls of tt4, this elevated transport
is reversed when flavonoids are synthesized by growth of plants on the
flavonoid precursor, naringenin. These results are consistent with a
role for flavonoids as endogenous regulators of auxin transport.
1
This work was supported by Sigma Xi (grant to
D.E.B.), by the National Aeronautical and Space Administration (grant
no. NAG2 1203 to G.K.M.), by the National Aeronautical and Space
Administration Specialized Center for Research and Training at North
Carolina State University (grants to G.K.M. and A.M.R.), by the
U.S. Department of Agriculture (grant no. 94-37100-0755 to A.S.M. and
L.T.), and by the National Science Foundation (grant no. MCB-9870798 to
J.N.). The Wake Forest University Research and Publications Fund
supported the publication costs.
2
Present address: Department of Horticulture and
Landscape Architecture, 1165 Horticulture Building, Purdue University,
West Lafayette, IN 47907-1165.
*
Corresponding author; e-mail muday{at}wfu.edu; fax
336-758-6008.
© 2001 American Society of Plant Physiologists
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|
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|
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|
|
|
|
|
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|
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|
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|
|
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|
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142(1):
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|
|
|
|
|
|
|
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141(4):
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|
|
|
|
|
|
|
N. A. Eckardt
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18(7):
1539 - 1540.
[Full Text]
[PDF]
|
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|
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18(7):
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|
|
|
|
|
|
|
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281(18):
12919 - 12928.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
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140(4):
1384 - 1396.
[Abstract]
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|
|
|
|
|
|
|
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April 1, 2006;
140(4):
1494 - 1506.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
Y. Dai, H. Wang, B. Li, J. Huang, X. Liu, Y. Zhou, Z. Mou, and J. Li
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PLANT CELL,
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18(2):
308 - 320.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
G. Lazar and H. M. Goodman
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PNAS,
January 10, 2006;
103(2):
472 - 476.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. Poupart, A. M. Rashotte, G. K. Muday, and C. S. Waddell
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Plant Physiology,
November 1, 2005;
139(3):
1460 - 1471.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
J. Misson, K. G. Raghothama, A. Jain, J. Jouhet, M. A. Block, R. Bligny, P. Ortet, A. Creff, S. Somerville, N. Rolland, et al.
A genome-wide transcriptional analysis using Arabidopsis thaliana Affymetrix gene chips determined plant responses to phosphate deprivation
PNAS,
August 16, 2005;
102(33):
11934 - 11939.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
R. Sibout, A. Eudes, G. Mouille, B. Pollet, C. Lapierre, L. Jouanin, and A. Seguin
CINNAMYL ALCOHOL DEHYDROGENASE-C and -D Are the Primary Genes Involved in Lignin Biosynthesis in the Floral Stem of Arabidopsis
PLANT CELL,
July 1, 2005;
17(7):
2059 - 2076.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. A. Tomilov, N. B. Tomilova, I. Abdallah, and J. I. Yoder
Localized Hormone Fluxes and Early Haustorium Development in the Hemiparasitic Plant Triphysaria versicolor
Plant Physiology,
July 1, 2005;
138(3):
1469 - 1480.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. W. WOODWARD and B. BARTEL
Auxin: Regulation, Action, and Interaction
Ann. Bot.,
April 1, 2005;
95(5):
707 - 735.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
W. A. Peer, A. Bandyopadhyay, J. J. Blakeslee, S. N. Makam, R. J. Chen, P. H. Masson, and A. S. Murphy
Variation in Expression and Protein Localization of the PIN Family of Auxin Efflux Facilitator Proteins in Flavonoid Mutants with Altered Auxin Transport in Arabidopsis thaliana
PLANT CELL,
July 1, 2004;
16(7):
1898 - 1911.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
M. Brinker, L. van Zyl, W. Liu, D. Craig, R. R. Sederoff, D. H. Clapham, and S. von Arnold
Microarray Analyses of Gene Expression during Adventitious Root Development in Pinus contorta
Plant Physiology,
July 1, 2004;
135(3):
1526 - 1539.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. S. Buer and G. K. Muday
The transparent testa4 Mutation Prevents Flavonoid Synthesis and Alters Auxin Transport and the Response of Arabidopsis Roots to Gravity and Light
PLANT CELL,
May 1, 2004;
16(5):
1191 - 1205.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
D. C. Gitz III, L. Liu-Gitz, J. W. McClure, and A. J. Huerta
Effects of a PAL inhibitor on phenolic accumulation and UV-B tolerance in Spirodela intermedia (Koch.)
J. Exp. Bot.,
April 1, 2004;
55(398):
919 - 927.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. C. Carey, J. T. Strahle, D. A. Selinger, and V. L. Chandler
Mutations in the pale aleurone color1 Regulatory Gene of the Zea mays Anthocyanin Pathway Have Distinct Phenotypes Relative to the Functionally Similar TRANSPARENT TESTA GLABRA1 Gene in Arabidopsis thaliana
PLANT CELL,
February 1, 2004;
16(2):
450 - 464.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
C. Chaban, F. Waller, M. Furuya, and P. Nick
Auxin Responsiveness of a Novel Cytochrome P450 in Rice Coleoptiles
Plant Physiology,
December 1, 2003;
133(4):
2000 - 2009.
[Abstract]
[Full Text]
|
|
|
|
|
|
|
H.-H. Woo, K. F. Faull, A. M. Hirsch, and M. C. Hawes
Altered Life Cycle in Arabidopsis Plants Expressing PsUGT1, a UDP-Glucuronosyltransferase-Encoding Gene from Pea
Plant Physiology,
October 1, 2003;
133(2):
538 - 548.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. M. Rashotte, J. Poupart, C. S. Waddell, and G. K. Muday
Transport of the Two Natural Auxins, Indole-3-Butyric Acid and Indole-3-Acetic Acid, in Arabidopsis
Plant Physiology,
October 1, 2003;
133(2):
761 - 772.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. S.A. Al-Hammadi, Y. Sreelakshmi, S. Negi, I. Siddiqi, and R. Sharma
The polycotyledon Mutant of Tomato Shows Enhanced Polar Auxin Transport
Plant Physiology,
September 1, 2003;
133(1):
113 - 125.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
R. V. Penmetsa, J. A. Frugoli, L. S. Smith, S. R. Long, and D. R. Cook
Dual Genetic Pathways Controlling Nodule Number in Medicago truncatula
Plant Physiology,
March 1, 2003;
131(3):
998 - 1008.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
E. Scarpella, K. J.M. Boot, S. Rueb, and A. H. Meijer
The Procambium Specification Gene Oshox1 Promotes Polar Auxin Transport Capacity and Reduces Its Sensitivity toward Inhibition
Plant Physiology,
November 1, 2002;
130(3):
1349 - 1360.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
G. K. Muday and A. S. Murphy
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PLANT CELL,
February 1, 2002;
14(2):
293 - 299.
[Full Text]
[PDF]
|
|
|
|
|
|
|
B. Winkel-Shirley
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Plant Physiology,
December 1, 2001;
127(4):
1399 - 1404.
[Full Text]
[PDF]
|
|
|
|
|
|
|
B. Noh, A. S. Murphy, and E. P. Spalding
Multidrug Resistance-like Genes of Arabidopsis Required for Auxin Transport and Auxin-Mediated Development
PLANT CELL,
November 1, 2001;
13(11):
2441 - 2454.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
R. E. Cleland
Unlocking the mysteries of leaf primordia formation
PNAS,
September 25, 2001;
98(20):
10981 - 10982.
[Full Text]
[PDF]
|
|
|
|
|
|
|
A. M. Rashotte, A. DeLong, and G. K. Muday
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PLANT CELL,
July 1, 2001;
13(7):
1683 - 1697.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
B. Winkel-Shirley
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Plant Physiology,
June 1, 2001;
126(2):
485 - 493.
[Full Text]
[PDF]
|
|
|
|
|
|
|
W. A. Peer, D. E. Brown, B. W. Tague, G. K. Muday, L. Taiz, and A. S. Murphy
Flavonoid Accumulation Patterns of Transparent Testa Mutants of Arabidopsis
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June 1, 2001;
126(2):
536 - 548.
[Abstract]
[Full Text]
[PDF]
|
|
|
|
|
|
|
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935 - 950.
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|
|
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