Seed dormancy and germination : light and nitrate

Research output: Thesisinternal PhD, WU


<p><TT>One of the most important aspects of the life cycle of seed plants is the formation and development of seeds on the motherplant and the subsequent dispersal. An equally important element of the survival strategy is the ability of seeds to prevent germination in unfavorable conditions, such as the wrong season, low light irradiance, or an unfavorable soil composition. Seeds of many species may remain in the soil, either in a dry state or fully imbibed, for hundreds of years without losing their viability. Earlier investigations have shown that this state of dormancy may be broken and induced in a seasonal cycle. Evidently, temperature is an important regulating factor in these cycles. When dormancy is broken, seeds may germinate, providing the conditions are adequate. Hence, seeds must be able to 'sense' their environment. Again, temperature is important, but also two naturally occurring environmental factors, light and nitrate, are known to stimulate the germination of many (wild) species.</TT><p><TT>Seeds of <em>Sisymbrium officinale</em> (hedge mustard) only germinate in the presence of light and nitrate (either endogenous or exogenous). Irradiation with far-red light (720 run) antagonized the stimulating action of red light (660 nm). Both wavelengths are part of the daylight spectrum. This is proof that the light-induced germination is mediated by the plant pigment phytochrome. The induction of germination by light and nitrate could also be inhibited by application of tetcyclacis, an inhibitor of the biosynthesis of gibberellins. Application of gibberellins 4 and 7 antagonized the inhibition. It was concluded that synthesis of gibberellins is part of the transduction chain that leads to germination. Comparison of the escape times for the antagonizing action of far-red light and the inhibition of tetcyclacis led to the conclusion that induction of germination occurred during the first 8 hours after irradiation with red light and application of nitrate, while the synthesis of gibberellins was completed for all seeds after 16 hours.</TT><p><TT>Seeds only germinated in the dark when exogenous gibberellins were present. In the absence of nitrate, red light reduced the requirement for gibberellins. In other words, besides a promotive effect on gibberellin synthesis (light-effect I), light also enhanced the sensitivity to gibberellins (light-effect II). Light-effect I disappeared after prolonged treatment at elevated temperatures while the seeds remained responsive to exogenous gibberellins and light-effect II (Chapter 2).</TT><p><TT>Both light-effects were studied in detail (Chapter 3). The influence of several concentrations of nitrate and gibberellins on the shape and position of fluence-response curves was examined. The results obtained with seeds of <em>Sisymbrium officinale</em> were compared with those of the gibberellin-deficient mutant of</TT><em>Arabidopsis t<TT>haliana</TT></em><TT>, a related species. Remarkable similarities were observed. In both cases nitrate steepened the fluence-response curves. In other words, the fluence-range over which the total seed population responded was reduced. Application of gibberellins 4 and 7 to seeds, in the absence of exogenous nitrate, resulted for both species in a parallel shift to lower fluence values. Application of cofactor analysis led to the conclusion that the interaction between the effects of nitrate and phytochrome was multiplicative. The germination response was a function of the product of nitrate concentration and fluence value. This is an indication that the factors nitrate and phytochrome act on the same pathway. The interaction between the effects of phytochrome and gibberellins, however, was additive. Both factors acted independently in different pathways leading to the same response. Phytochrome might enhance the sensitivity of the gibberellin- receptors (light-effect II). However, this effect could only be brought to expression when sufficient gibberellins were present. The use of the gibberellindeficient</TT><em>Arabidopsis mutant</em><TT>supported the hypothesis that in Sisymbrium no active gibberellins are present in the absence of nitrate.</TT><p><TT>The hypothesis that nitrate is active in the induction of germination because its reduction would lead to production of NADP, stimulator of the pentose phosphate pathway, was tested by studying the light- and nitrateinduced germination in the presence of inhibitors of nitrate reductase (Chapter 4). Furthermore, a method was developed to measure the nitrate content of seeds. The inhibitors of nitrate reductase, sodium chlorate and sodium tungstate had no influence on the light-induced germination in a range of nitrate concentrations. At several intervals after a pretreatment during which the seeds had taken up nitrate from the medium, the nitrate contents of the seeds and the medium were measured. It was found that during the induction of germination, the 8-hour period after irradiation, the nitrate content of the seeds decreased. However, this decrease could be fully explained by leakage into the medium. Thus, no nitrate was reduced during this period. Nitrate reduction did not occur until actual growth (protrusion of the radicle) had started. Apparently, at that time the nitrate assimilation began. In the presence of the inhibitors this reduction was almost completely inhibited. The growth of the seedling was abnormally slow in the presence of the inhibitors. These results led to the conclusion that nitrate is active in the unreduced state at induction of germination.</TT><p><TT>To gain more insight in the process of dormancy induction and the decreasing sensitivity to light and nitrate during this process, doseresponse experiments were carried out for these factors during induction of dormancy at constant temperature (Chapter 5). Fluence - response curves of seeds in supra-optimal nitrate concentrations shifted to higher fluence values upon increasing duration of the pretreatment. After approximately 120 hours at 15°C the maximum germination decreased. The slopes of the curves did not change. The observed fluence-response curves could be simulated by formulations from the general receptor -occupancy theory. This enabled us to interpret calculated curve parameters as interaction parameters of the binding of phytochrome to its receptor. The shifts of the curves with unchanged maximal response could in this way be explained by assuming that more phytochrome -receptors were present than required for maximal germination. The induction of secondary dormancy could then be the result of a temperature dependent decrease of the number of receptors. Once below a critical value this decrease could lead to reduction of the maximum response.</TT><p><TT>The role of nitrate was studied in a similar way (Chapter 6). The response to nitrate was monitored in optimal light conditions. The results of the nitrate-response experiments showed similarities with those of the fluence-response experiments. Upon increasing pretreatment duration a shift to higher nitrate concentrations was observed, followed by a decrease of the maximum germination. The nitrate-response curves could also be simulated with an equation for a simple bimolecular reaction. Furthermore, it was shown that the presence of nitrate was an absolute requirement for the light-induced germination. It was concluded that nitrate may act as an activator of the phytochrome-receptors. In the absence of exogenous nitrate, the response was limited by the amount of endogenous nitrate. Since nitrate leached out during incubation in water the germination in water was correlated with the nitrate leaching. A remarkable aspect of the nitrate -responses was the occurrence of biphasic response curves. The very low nitrate response disappeared after 48-72 hours of pretreatment. The occurrence of these two phases was related to the uptake of nitrate from a range of nitrate concentrations. For the first phase the uptake appeared to be the limiting factor; all the nitrate that was taken up was bound. For the second phase relatively high endogenous nitrate levels were required. A considerable amount of this nitrate was not bound. The possibility of the existence of two different nitrate receptors was discarded. By analogy with an existing model for biphasic fluence-response curves it was suggested that the requirement of the nitrate -receptor for nitrate for a certain fraction of the seed population could differ from the rest of the population. Seemingly, induction of dormancy could influence the ratio of these fractions.</TT><p><TT>The results were summarized in a model in which the phytochrome-receptor is located in a membrane (Chapter 7). Breakage of dormancy, regulated by temperature, would induce synthesis of this receptor. Changing the temperature to an optimal germination temperature would induce a phase transition of the membrane, thus enabling the receptor to move laterally in the membrane. In this way the receptor may become accessible to nitrate. Nitrate would bind to the receptor and alter its conformation in such a way that phytochrome can bind. The transduction chain, leading to germination, may be initiated via a secondary messenger like calcium. Induction of dormancy makes this process impossible because the rate of degradation of phytochrome-receptors will exceed the synthesis.</TT>
Original languageEnglish
QualificationDoctor of Philosophy
Awarding Institution
  • Karssen, C.M., Promotor
Award date21 Mar 1990
Place of PublicationS.l.
Publication statusPublished - 1990


  • germination
  • seed germination
  • seed dormancy
  • light
  • photoperiodism
  • nitrates

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