Gibberellin requirement for Arabidopsis seed germination is determined both by testa characteristics and embryonic abscisic acid

Abstract

The mechanisms imposing a gibberellin (GA) requirement to promote the germination of dormant and non-dormant Arabidopsis seeds were analyzed using the GA-deficient mutant ga1, several seed coat pigmentation and structure mutants, and the abscisic acid (ABA)-deficient mutant aba1. Testa mutants, which exhibit reduced seed dormancy, were not resistant to GA biosynthesis inhibitors such as tetcyclacis and paclobutrazol, contrarily to what was found before for other non-dormant mutants in Arabidopsis. However, testa mutants were more sensitive to exogenous GAs than the wild-types in the presence of the inhibitors or when transferred to a GA-deficient background. The germination capacity of the ga1-1 mutant could be integrally restored, without the help of exogenous GAs, by removing the envelopes or by transferring the mutation to a tt background (tt4 and ttg1). The double mutants still required light and chilling for dormancy breaking, which may indicate that both agents can have an effect independently of GA biosynthesis. The ABA biosynthesis inhibitor norflurazon was partially efficient in releasing the dormancy of wild-type and mutant seeds. These results suggest that GAs are required to overcome the germination constraints imposed both by the seed coat and ABA-related embryo dormancy.

Figure 1

Effect of light and chilling on dormancy breaking and germination of wild types, single mutants, and double mutants. Black bars represent germination of genotypes in Ler background in darkness; the dotted bars (in A and B) represent germination of genotypes in Ws background in darkness. The gray bars (in C and D) represent germination of Ler genotypes in light, and the striped bars (in C and D) represent germination of Ws genotypes in light. Seeds were sown on water 11 d after harvest.

Figure 2

Sensitivity to GA₄₊₇ of wild types, single, and double mutants between tt and ga1 mutants. A cold treatment was applied to seeds before germination. A, ○, Ler; ▴, tt4; ▪, ttg1; ●, ga1-1; ♦, ga1-3; ▵, ga1-1 tt4; □, ga1-1 ttg1. B, ○, Ws; ▴, tt12; ●, ga11-11; ▵, ga1-11 tt12. Dashed lines represent wild types (Ws and Ler).

Figure 3

Sensitivity of wild-type and seed coat mutant seeds to the GA-biosynthesis inhibitors tetcyclacis and paclobutrazol. A cold treatment was applied to seeds before germination. Sowing was done 3 months after seed harvest. A and D, ○, Ler; ●, tt2; ▴, tt4; ▪, tt7; ♦, ttg1. B and E, ○, Ler; ●, ats; ▴, ap2; ▪, gl2. C and F, ○, Ws; ●, tt12. Dashed lines represent the wild types Ler and Ws.

Figure 4

Sensitivity of wild-type and seed coat mutant seeds to GA₄₊₇ in presence of either 100 μm tetcyclacis (A–C) or 100 μm paclobutrazol (D–F). A cold treatment was applied to seeds before germination. Sowing was done 6 months after seed harvest. A and D, ○, Ler; ●, tt2; ▴, tt4; ▪, tt7; ♦, ttg1. B and E, ○, Ler; ●, ats; ▴, ap2; ▪, gl2. C and F, ○, Ws; ●, tt12. The dashed lines represent the wild types Ler and Ws.

Figure 5

Genetic determinism of the increased sensitivity to GA₄₊₇ exhibited by the tt12 mutant in the presence of 100 μm tetcyclacis. The parent mentioned first was used as the female parent and the second as the pollen parent. A cold treatment was applied to seeds before germination. ○, Ws; ●, tt12; ▵, Ws × tt12; ▴, tt12 × Ws.

Figure 6

Sensitivity to the ABA biosynthesis inhibitor norflurazon of wild types, single mutants, and double mutants between tt and ga1 mutants. Seeds were sowed 10 d after harvest. A, ○, Ler; ▴, tt4; ▪, ttg1; ●, ga1-1; ▵, ga1-1 tt4; □, ga1-1 ttg1. B, ○, Ws; ▴, tt12; ●, ga1-11; ▵, ga1-11 tt12.

Figure 7

Aleurone layer of the wild-type Ler before (a) and after (b) seed germination. The aleurone layer (arrowheads) is still associated with pieces of the seed coat. Cell walls are stained with ruthenium red. Bar = 20 μm.

Figure 8

Schematic presentation of the interactions between the envelopes and the embryo affecting dormancy breaking and germination of Arabidopsis seeds. Germination occurs when the growth potential of the embryo is sufficient to overcome the restraint to radicle protrusion imposed by the seed envelopes. When the restraint of the testa is weakened by mutations, the growth potential threshold required for germination is lowered. The sharp arrows and the blunt arrows stand for a promotive and an inhibitory action, respectively. Dashed arrow indicates leaching of ABA through the testa.