pathway, have been used for this purpose. In each of these cases, several genes have been identified which
require ABA for expression [14,15,87].
A. Identification of Genes That Require ABA for Expression
Mutants that are blocked in the ABA biosynthetic pathway have proved to be useful in the identification
of genes that require elevated levels of ABA for expression. mRNAs for specific water deficit–induced
genes do not accumulate in response to a water deficit in ABA-deficient mutants as they do in the wild
type. For example, the ABA-deficient mutant of tomato, flacca, is blocked in the last step of the ABA
biosynthetic pathway [102]. This mutant does not accumulate as much ABA during periods of water
deficit as does the wild type. After stress, the mutant accumulates 6% of the ABA that the wild type ac-
cumulates [103]. There are fewer proteins accumulating in flaccaleaves than in the wild type during
drought stress. Application of ABA to flaccarestores the accumulation of this set of ABA-induced pro-
teins [103]. The accumulation of three mRNAs was shown to be dependent on the accumulation of ABA
during stress [14]. These mRNAs accumulated only in the drought-stressed wild type and were not de-
tected in the drought-stressed ABA-deficient mutant. Mutants in maize and Arabidopsishave been used
similarly to identify additional ABA-requiring genes (Table 1).
B. Genes That Are Responsive to ABA but Do Not Require ABA
The ABA-deficient mutants have been used to define an additional set of genes, those that are responsive
to ABA but do not require ABA for expression [30,104,105]. These genes are induced by ABA applica-
tion, but unlike the ABA-requiring genes, they are induced by low-temperature and water-deficit treat-
ments in the ABA-deficint mutants of Arabidopsis. Therefore, it has been concluded that these genes do
not require elevated levels of endogenous ABA for expression but are capable of responding to ABA and
may be called ABA-responsive genes. These results indicate that there are two pathways that can be fol-
lowed to induce these genes, but it is unknown if the pathways converge or if there are two entirely sep-
arate pathways. ABA applications have also been used to show that there are a number of water deficit–in-
duced genes that do not respond to ABA application [52,76]. These genes may be induced directly by the
drought stress, or they may be controlled by other signaling mechanisms operating during water deficit.
C. DNA Elements That Confer ABA Responsiveness
The conditions under which a gene is induced is controlled by the DNA elements acting within each gene.
Therefore, to understand the mechanism of regulation of a specific gene, the DNA elements that confer re-
sponsiveness and the factors that recognize those elements must be identified and characterized. Studies
have been initiated on genes that are regulated by ABA to understand the basis of ABA-regulated gene ex-
pression during stress and seed development. A region of Em, a member of the group 1 leafamily from
wheat, was identified which confers ABA inducibility upon a minimal 35S CaMV promoter [106,107]. A
chimeric gene was constructed with a 646-bp segment of Emand the reporter gene -glucuronidase (GUS).
When this gene was introduced into rice protoplasts, GUS activity was increased 15- to 30-fold after ABA
ABIOTIC STRESSES AND ABSCISIC ACID 745
TABLE 1 Genes That Have Been Demonstrated to Be Regulated by Elevated Levels of Endogenous ABA
Resulting from Environmental Stress Using ABA-Deficient Mutants of Maize, Arabidopsis,and Tomato
Genotype Gene designation Gene family Stress inductiona Refs.
Maize rab 17 dhn/rab/group 2 D 13
rab 28 D-34/group 6 D 87
Arabidopsis rab 18 dhn/rab/group 2 D, L 15
lti 65 rd29/cor78 D, L 28,29,104
Tomato le 4 dhn/rab/group 2 D, S, L 11,14
le 16 nsLTP D, S, L 14,64
le 20 H1-histone D, S, L 85
le 25 D-113/group 4 D, S, L 11,14
aD, water deficit; L, low-temperature stress; S, salinity.