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Plcell v9 7 1197
Plcell v9 7 1197
Plcell v9 7 1197
It is 60 years since Went and Thimann (1937) published their two aspects are not considered in the following discussion
classic book Phytohormones. At that time, the term phyto- of how the endogenous pool sizes of the five classical hor-
hormone was synonymous with auxin, although the exis- mones are regulated. Rather, we focus on the biosynthetic
tente of other phytohormones, such as cell division factors, pathways, $he deactivation reactions, and the regulatory
was anticipated on the basis of physiological experiments. It mechanisms involved in these processes.
is impressive that aside from some confusion about the During the past 25 years, the standards set for natural-
structure of auxin, many of the basic phenomena of auxin product chemistry have also been applied to plant hormone
physiology were already known at that time. It is equally im- research. lnstead of “measuring” hormones by bioassays,
pressive that much auxin biology, including the Cholodny- unambiguous physical-chemical methods for the identifica-
Went hypothesis (Went and Thimann, 1937) regarding the tion and measurement of hormones have been developed
role of auxin in mediating gravi- and phototropism, the path- (reviewed in Hedden, 1993). The accuracy and facility of
way of auxin biosynthesis, and the mechanism by which quantitative measurements have been improved by the
auxin causes cell wall loosening, remains controversial. availability of isotopically labeled versions (with 2H, 13C, or
Since 1937, gibberellin (GA), ethylene, cytokinin, and ab- I5N) of the hormones for use as interna1 standards.
scisic acid (ABA) have joined auxin as phytohormones, Molecular genetics is another discipline that has made it
and together, they are regarded as the “classical five” (Fig- possible to solve problems in hormone physiology that were
ure 1). This group is expected to grow as the hormonal hitherto intractable. Hormones are present in plants in very
functions of other compounds are recognized and as new small amounts. Moreover, their biosynthetic and catabolic
hormones are discovered (see Creelman and Mullet, 1997, enzymes are low-abundance proteins, which, in most cases,
in this issue). As is evident from this short review, recent cannot be isolated and purified by classical biochemical
progress on hormone biosynthesis and on hormonal trans- methods. However, when the identification of a mutant leads
duction pathways has been impressive. Also evident is that to the cloning of a gene, that gene can be expressed as a
there are many blanks still to be filled in. With the application fusion protein with which the catalytic function can be deter-
of the powerful new techniques of chemical analysis and mined (e.g., Sun and Kamiya, 1994; Xu et al., 1995; Schwartz
molecular genetics, the rate at which new discoveries are et al., 199713).
made will continue to accelerate. It’s a great time to be a
plant hormonologist!
Auxin
CHEMISTRY AND BIOSYNTHESIS OF HORMONES The primary auxin in plants is indole-3-acetic acid (IAA; Fig-
ure 1). Although other compounds with auxin activity, such
as indole-3-butyric acid, phenyl acetic acid, and 4-chloro-
Clearly, the amount of any compound, including hormones, IAA, are also present in plants (reviewed in Normanly et al.,
in an organ of a plant is determined by the combined rates 1995), little is known about their physiological role. For many
of its biosynthesis, breakdown, import, and export. The last years, it has been assumed that tryptophan is the precur-
sor of IAA. This has recently been confirmed in seedlings
of Phaseolus vulgaris with stable isotope labeling studies
This review is dedicated to the memories of James Bonner, Anton (Bialek et al., 1992). Three routes for IAA biosynthesis from
Lang, Kenneth Thimann, and Philip Wareing, pioneers in plant hor-
tryptophan via indole-3-pyruvic acid, tryptamine, or indole-
mone research, who died during the past year.
To whom correspondence should be addressed. E-mail zeevaart@ 3-acetonitrile have been proposed. The latter precursor is
pilot.msu.edu; fax 517-353-9168. found primarily in the Cruciferae and may be derived from
1198 The Plant Cell
HzC=CH2 O
H
Indole-3-acetic acid Ethylene S-(+)-Abscisic acid
N
N
H
Zeatin Gibberellin AI
Figure 1. Structures of Representatives of the Five Classical Plant Hormones.
Shown are indole-3-acetic acid, ethylene, abscisic acid, zeatin, and gibberellin A,.
indoleglucosinolates (Normanly et al., 1995). Four genes en- first enzyme, tryptophan monooxygenase, converts tryp-
coding nitrilase, which converts indole-3-acetonitrile to IAA, tophan to indole-3-acetamide, which in turn is converted to
have been cloned in Arabidopsis. These four genes are dif- IAA by indole-3-acetamide hydrolase. The genes encoding
ferentially regulated (Bartel and Fink, 1994). these enzymes have been used to alter IAA levels in trans-
Work with tryptophan auxotrophic mutants has estab- genic plants (Klee and Romano, 1994).
lished that IAA biosynthesis can also take place via a tryp- IAA occurs not only in the free form but also conjugated to
tophan-independent route. For example, the orange pericarp amino acids, peptides, or carbohydrates. These IAA conju-
mutant in maize does not produce tryptophan but accu- gates are biologically inactive and appear to serve functions
mulates IAA to levels 50-fold higher than in the wild type as IAA storage forms in seeds and hormonal homeostasis.
(Wright et al., 1991). Tryptophan auxotrophs in Arabidopsis The iaglu gene in maize, which encodes an enzyme that es-
also accumulate more IAA than do wild-type plants. On the terifies IAA to glucose, has been cloned (Szerszen et al.,
basis of these data, it was proposed that IAA can be synthe- 1994). In Arabidopsis, a gene family that encodes IAA conju-
sized through a branch point of the tryptophan biosynthetic gate hydrolases has been identified (Bartel, 1997).
pathway at indole or indole-glycerol phosphate (Normanly et Until recently, IAA catabolism was thought to occur via
al., 1995). Supporting this idea is the finding that in a cell- oxidative decarboxylation (i.e., through the action of an IAA
free system from immature maize endosperm, radioactive oxidase). However, the major catabolic route of IAA in vivo
indole is converted to IAA (Rekoslavskaya and Bandurski, now appears to be oxidation to oxindole-3-acetic acid and
1994). subsequent glycosylation through an added 7-hydroxyl (re-
Certain bacteria and plant cells transformed with Agro- viewed in Normanly et al., 1995). Another catabolic pathway
bacterium tumefaciens synthesize IAA via a unique pathway is via IAA-acetylaspartate to dioxindole-3-acetylaspartate-3-
in which tryptophan is converted to IAA in two steps. The O-glucoside.
The Five Classical Plant Hormones 1199
Location:
Enzymes:
P450 monooxygenases
.-
ent-Kaurene ent-Kaurenol ent-Kaurenal ent-Kaurenoic acid
Inhibitors:
N-heterocyclics:
Paclobutrazol
4 4 Tetcyclacis
OH Uniconazole
COOH COOH
ent-Kaurenoic acid ent-7a-hydroxy- GAl 2-aldehyde
Kaurenoic acid
Location:
Cytoplasm
@ Enzymes:
I,... I..,’ ca @
,.... 4 @ I,... Dioxygenases
@
4
COOH
GAi7 GA2Q GAe
step in Agrobacterium is the addition of an isopentenyl group ring of the ABA molecule (Zeevaart et al., 1991). Finally, in eti-
from isopentenyl diphosphate to N6 of AMP, which is cata- olated leaves and roots, which have low levels of carotenoids,
lyzed by AMP-isopentenyl transferase (IPT). A similar en- a 1 : l stoichiometry was found between the disappearance
zyme activity has also been observed in extracts from plant of violaxanthin and neoxanthin and the appearance of ABA
sources (Blackwell and Horgan, 1994; Chen and Ertl, 1994), and its catabolites (Li and Walton, 1990; Parry et al., 1992).
but because of its instability, the enzyme has only been par- The ABA-deficient aba7 mutant of Arabidopsis is blocked
tially purified (Chen and Ertl, 1994). The I f T gene from Agro- in the epoxidation of zeaxanthin to antheraxanthin and vio-
bacterium has been cloned and expressed in transgenic laxanthin, indicating that the epoxycarotenoids violaxanthin
plants (Klee and Romano, 1994). However, there are no re- and neoxanthin are essential for ABA production (Rock and
ports of plant DNA sequences with similarity to the bacterial Zeevaart, 1991). The aba2 mutant of Nicotiana plumbagini-
cleavage enzyme in transgenic plants may not necessarily organs and at different stages of development have been
result in increased ABA levels. To raise ABA levels and to observed (Barry et al., 1996). Moreover, the positive feed-
make plants tolerant to stress conditions, it may be prefera- back loop in which treatment of tissue with ethylene often
ble to suppress the activity of ABA 8'-hydroxylase, the en- stimulates ethylene production by that tissue appears to
zyme that converts ABA to phaseic acid, by using antisense take place through enhanced expression of ACC synthase
technology. However, the experiments will have to wait until and ACC oxidase (reviewed in Kende, 1993).
the corresponding gene has been cloned. Besides being converted to ethylene, ACC can also be ir-
reversibly conjugated to form N-malonyl-ACC (Kionka and
Amrhein, 1984). Malonylation of ACC regulates the level of
Ethylene ACC and thus the production of ethylene. Ethylene can be
act? Answers to the first question are mostly lacking; an- plant hormones are expected to cause loosening of the cell
swers to the second are beginning to emerge. wall, but how is this achieved? The acid-growth theory pos-
In the intercalary meristem of deepwater rice, GA pro- tulates that secretion of hydrogen ions into the cell wall is
motes cell division and cell elongation (Sauter and Kende, stimulated by auxin and that the lowered pH in the apoplast
1992). This leads to internodal growth rates of up to 5 mm/ activates wall-loosening processes (Rayle and Cleland,
hr. Measurements of cell length and determinations of the 1970; Hager et al., 1971). Indeed, there are severa1 lines of
progress of cells through the cell cycle by flow cytometry evidence that support this hypothesis, in particular, the facts
and 3H-thymidine incorporation indicated that GA-induced that auxin causes acidification of the cell wall and that acidic
cell elongation preceded the promotion of cell division. buffers induce growth in auxin-sensitive tissues (Rayle and
Therefore, it has been proposed that the primary action of Cleland, 1992).
stopped growing (Bergfeld et al., 1988). This may explain membrane. Evidence for a receptor function of the plasma
why GA action usually requires the presence of a meristem, membrane-bound ABP comes from experiments showing
where GA promotes cell elongation and, perhaps indirectly, that auxin-induced hyperpolarization is inhibited by antibod-
cell division (Sauter and Kende, 1992), whereas auxin can ies against ABPl (Barbier-Brygoo et al., 1989). It is not known,
promote elongation of older cells in the absence of a mer- however, whether ABPl plays any role in mediating growth.
istem (see Jacobs, 1997, in this issue). A number of “auxin-resistant” mutants of Arabidopsis
Ethylene inhibits the elongation of terrestrial plants and have been isolated for the genetic dissection of the auxin
causes thickening of their stems. This effect has been as- transduction pathway (Walden and Lubenow, 1996). These
cribed to a reorientation of both the cortical microtubules mutants were selected for their ability to grow on high con-
and the newly deposited cellulose microfibrils from mostly centrations of auxin. By using this strategy, Leyser et al.
ence of ethylene, dark-grown seedlings of such mutants main. The ETR2 and ETHYLENE-INSENSITIVE4 (EIN4) genes
either do not exhibit the "triple response" (i.e., stunted encode homologs of ETR1, and mutations in these genes
growth, swelling of the root and hypocotyl, and exaggerated confer dominant ethylene insensitivity onto Arabidopsis seed-
apical hook formation) or show the triple response pheno- lings (Roman et al., 1995; Hua et al., 1997).
type even in the absence of ethylene. We limit our discus- Hua et al. (1995) have cloned an Arabidopsis gene, ETH-
sion of ethylene signal transduction to four components of YLENE RESPONSE SENSOR (ERS), that encodes a second
the pathway whose functions have been derived from se- type of putative ethylene receptor. The sensor domain of the
quence analysis of the corresponding genes and from direct ERS protein shows high similarity to ETR1, but it lacks a re-
ethylene binding experiments. ceiver domain (Figure 3). When, by site-directed mutagene-
The mutation ethylene-resistantl (etrl) is dominant, and sis, the same amino acid change was introduced into ERS,
N-
Input Histidine protein Receiver Output
domain kinase domain domain domain
B
N-
c
N-
Figure 3. Two-Component Signaling Systems.
(A) The bacterial two-component system composed of sensor and response regulator proteins.
(B) The ETR-type two-component system. The ethylene binding site is located in the frans-membrane region of the input domain. The tomato
eTAE1 homolog implicated in ethylene signal transduction (Zhou et al., 1996) and the CKI1 homolog implicated in cytokinin signaling (Kakimoto,
1996) are of this type.
(C) The ERS-type protein. ERS lacks the receiver domain; the tomato NR protein is a homolog of ERS (Wilkinson et al., 1995).
1206 The Plant Cell
kinase-mediated phosphorylation cascade, albeit one linked Foundation (Grant Nos. IBN-9118377 and lBN-9407763), and the
to a receptor related t o those typically associated with prokary- U.S. Department of Agriculture (Competitive Grants Program; Grant
otic sensing systems. NO.94-37304-0956).
A second downstream gene is HOOKLESS7 (HLS7) of
Arabidopsis, which was identified as an ethylene-responsive
gene whose expression is required for the formation of the REFERENCES
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