Control of Seed Germination by Abscisic Acid

Abstract

The germination process of mustard seeds (Simapis alba L.) has been characterized by the time courses of water uptake, rupturing of the seed coat (12 hours after sowing), onset of axis growth (18 hours after sowing), and the point of no return, where the seeds lose the ability to survive redesiccation (12 to 24 hours after sowing, depending on embryo part). Abscisic acid (ABA) reversibly arrests embryo development at the brink of radicle growth initiation, inhibiting the water uptake which accompanies embryo growth. Seeds which have been kept dormant by ABA for several days will, after removal of the hormone, rapidly take up water and continue the germination process. Seeds which have been preincubated in water lose the sensitivity to be arrested by ABA after about 12 hours after sowing. This escape from ABA-mediated dormancy is not due to an inactivation of the hormone but to a loss of competence to respond to ABA during the course of germination. The sensitivity to ABA can be restored in these seeds by redrying. It is concluded that a primary action of ABA in inhibiting seed germination is the control of water uptake of the embryo tissues rather than the control of DNA, RNA, or protein syntheses. When a quiescent, nondormant seed (13) is supplied with water and 02 at favorable temperatures the embryo rapidly takes up water and continues its temporarily suspended development. After building up a certain threshold hydrostatic pressure the seed coat is ruptured and visible protrusion of the elongating radicle indicates the onset of elongation of the embryonic axis. Some time later the release from quiescence becomes irreversible, ie. the embryo will no longer survive redesiccation. The developmental period from the increase of metabolic activity after imbibition up to this point of no return, logically separating the embryo from the seedling stage, can be referred to as germination. At 25 C germination lasts not more than about 1 day in many seeds. The dormancy hormone ABA can inhibit continuation of embryo development during germination and related developmental processes (e.g. the growth of Lemna turions or tree buds, 19, 20). Exogenously applied ABA is rapidly taken up by the embryo even through the intact seed coat (3, 16). The investigation of dormancy imposed by supplying germinable seeds with ABA provides an opportunity to study a physiologically relevant phenomenon (5) which, in contrast to most cases of natural dormancy, can be precisely controlled. This approach may provide information not only on the regulation of seed dormancy but also on the molecular mechanism of ABA action in plants. It has been shown that fundamental processes such as cell division and the synthesis of DNA, RNA, and protein become inhibited in plant systems treated with ABA (6, 10, 11, 19, 21). In barley aleurone tissue and cotton embryos exogenous ABA prevents the formation of enzymes which are involved in mobilizing storage materials during germination (5, 12). Although the molecular nature of the physiological block, by which ABA prevents the completion of germination, is still unknown, it is generally believed that ABA functions by interfering with mRNA synthesis, processing, or translation (e.g. 5-7, 10, 11, 24). In a previous study with mustard (1) we have observed that seedling development is surprisingly insensitive to ABA if the hormone is applied after germination. In particular it has been established that seedling photomorphogenesis including the induction of several enzyme activities is essentially unaffected by ABA concentrations which prevent the completion of germination. Here, we characterize the physiological action of ABA and the disappearance of sensitivity toward this hormone during the germination of mustard seeds. MATERIALS AND METHODS ± Abscisic acid (puriss.) was a product of Fluka AG (Buchs, Switzerland). Seeds of Sinapis alba L. (harvest 1972, obtained from Asgrow Company, Freiburg-Ebnet) were selected for uniformity of color, size, shape, and sown using standard conditions: 50 seeds were rinsed briefly with distilled H20 (or ABA solution) and placed on five layers of Schleicher and Schull chromatography paper (No. 2043 b Mgl, presoaked with the appropriate solution for 12 h) in covered plastic boxes (10 x 10 x 8 cm). After adding 5 ml excess water (or ABA solution) a small meniscus formed around the seed which was covered by a thin film of liquid due to the mucilaginous testa surface. The boxes were kept in darkness at 25.0 ± 0.3 C if not indicated otherwise. Before transferring to a different medium the seeds were thoroughly washed by incubation for 10 min in an abundance of new medium. All manipulations were done under green safelight. For redrying, pregermi-nated seeds were transferred to Petri dishes containing silica drying beads (Merck, Darmstadt). The seeds lost 50%o of their moisture within 1 h and reached their original dry weight within 3 h. These seeds were left on the silica beads for 24 to 72 h and were then resown on water. Their further development was observed in white fluorescent light (600 lx) at 20 C. The percentage of seeds with broken testa (tip of radicle just visible) and of seeds with growing radicle (radicle protruding for more than 2 mm) was determined at suitable intervals under safelight. Data of the figures represent the means of at least four independent experiments (200 seeds). RESULTS Time Course of Germination Parameters. The conventional criterion for discriminating germinated from nongerminated seeds is the protrusion of the radicle tip through the seed coat (2, 9). This purely operational criterion for germination appears to be 822