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Role of ABA and ABI3 in Desiccation Tolerance

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Science  29 Jan 2010:
Vol. 327, Issue 5965, pp. 546
DOI: 10.1126/science.1183672

Abstract

We show in bryophytes that abscisic acid (ABA) pretreatment of moss (Physcomitrella patens) cells confers desiccation tolerance. In angiosperms, both ABA and the transcriptional regulator ABSCISIC ACID INSENSITIVE 3 (ABI3) are required to protect the seed during desiccation. ABA was not able to protect moss cells in stable deletion lines of ABI3 (ΔPpabi3). Hence, moss has the same functional link between ABA, ABI3, and the desiccation tolerance phenotype that is found in angiosperms. Furthermore, we identified 22 genes that were induced during ABA pretreatment in wild-type lines. When their expression was compared with that of ΔPpabi3 during ABA pretreatment and immediately after desiccation, a new target of ABI3 action appears to be in the recovery period.

To survive on land, the earliest land plants had to develop mechanisms to tolerate desiccation. Modern vascular plants possess an array of morphological features to retain water (such as conductive tissues, cuticle, and stomata) and have retained desiccation tolerance in only a few specialized structures (e.g., seeds). Present-day bryophytes (mosses), in contrast, lack water transport and retention tissues, presumably like early land plants. As a result, their vegetative state is at equilibrium with the surrounding air, creating a water-deficit condition that most angiosperms could not tolerate (1). Phylogenetic analyses suggest that desiccation tolerance in vegetative tissue of bryophytes was lost in the first vascular plants (2). Here, we evaluate whether desiccation tolerance in angiosperm seeds and in vegetative tissues of the moss Physcomitrella patens use similar regulatory pathways.

The phytohormone abscisic acid (ABA) protects seeds during water stress by activating genes through transcription factors such as ABSCISIC ACID INSENSITIVE 3 (ABI3) (3).

ABA is also found in nonseed plants such as algae and P. patens (4) and uses similar signaling pathways. For example, a wheat ABA-responsive promoter can be activated by ABA in cells of P. patens (5), and one of three homologs of ABI3 found in P. patens partially complements the Arabidopsis abi3-6 mutant (6).

Untreated wild-type (WT) filaments of P. patens can survive up to 92% water loss (7) but cannot recover from complete desiccation (Fig. 1A). We generated two independent lines (∆abi3-1 and ∆abi3-2) in which all three P. patens ABI3 genes (A, B, and C) were deleted by using sequential gene targeting (fig. S1) (8, 9). WT lines survived if incubated with ABA (10 to 100 μM) for 24 hours before desiccation, whereas two ∆abi3 lines did not survive, even at 100 μM ABA (Fig. 1A). The ∆abi3 lines were also not responsive to an ABA-responsive promoter from moss (PpLEA1a-GUS), whereas WT exhibited an increase (fig. S2). Expression of 22 ABA up-regulated genes from WT P. patens (that are presumably required for tolerance) were compared with those of ∆abi3 at 24 hours after ABA treatment, 24 hours after drying, and 5 min and 15 min after rehydration (Fig. 1B). Without PpABI3, only a few transcripts had reduced expression after ABA treatment and drying, whereas the others maintained their expression. The loss of PpABI3 had little effect on this subset of ABA up-regulated genes before rehydration. However, all 22 genes assayed at 5 and 15 min after rehydration showed drastically reduced transcripts or none at all in the ∆abi3-1 line when compared with WT (Fig. 1B). For successful recovery from desiccation, PpABI3 appears to be essential for the maintenance, either by synthesis or stabilization, of those transcripts induced during the ABA pretreatment that are critical for tolerance.

Fig. 1

(A) ABA and PpABI3 are required for desiccation tolerance. Tissues from 6-day-old WT, ∆abi3-1, and ∆abi3-2 were treated with ABA (0, 10, 50, and 100 μM) for 24 hours. Tissues were dried for 24 hours, rehydrated with sterile distilled water, and incubated for 2 weeks. (B) Reverse transcription polymerase chain reaction (RT-PCR) analysis of ABA-induced transcripts in WT and ∆abi3-1 during ABA treatment, drying, and rehydration. RNA was extracted from 6-day-old tissues 24 hours after ABA treatment, 24 hours after drying, and 5 and 15 min after rehydration in basal medium. cDNA was synthesized with use of 2 μg of RNA, and PCR was performed with use of gene-specific primers (table S1).

We conclude that both ABA and ABI3 are required for P. patens vegetative tissue to survive desiccation. Because the P. patens genome lacks the transcription factors FUS3 and LEC2 (10) that are required for seed maturation like ABI3 (3), the role of ABI3 in this nonseed plant appears to be directly in desiccation tolerance, primarily in the recovery stage. Our working hypothesis is that gene regulatory pathways that include both ABA and ABI3 originally evolved for cellular protection from water deficits but independently have been used to provide desiccation tolerance in vegetative tissues of bryophytes and in angiosperm seeds.

Supporting Online Material

www.sciencemag.org/cgi/content/full/327/5965/546/DC1

Materials and Methods

Figs. S1 and S2

Table S1

References

References and Notes

  1. Materials and methods are available as supporting material on Science Online.
  2. We thank L. Gunther and L. Maines for technical support and D. Cove and S. McDaniel for many helpful discussions. Supported by funds from NSF (EF-0425749) and Washington University.
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