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Figure 1. 

ROP1 overexpression disrupts dormancy of pollen inside anther under moist environments. (a), (b) Images and germination ratio of Col-0 and ROP1-OE pollen 1 after imbibition. Scale bars = 20 μm. Error bars represent SD. Statistical significance was determined by Student's t-test (*** p < 0.001, n = 3). (c) Fluorescence images of Col-0 and ROP1-OE anther with or without pollen tubes stained with aniline blue. Scale bars = 100 μm. (d) Image of Col-0 and ROP1-OE anther and stigma under high humidity conditions. The red arrow points to the pollen tube. Scale bars = 500 μm in the left images. Scale bars = 100 μm in the images of the middle and right.

Figure 2. 

Active ROP1 localization in the dormant pollen and germinated pollen of Arabidopsis. (a) CRIB4- GFP distribution in the dormancy mature pollen grain. (b)−(d) CRIB4-GFP polar distribution in the germinated site of pollen during germination. (e), (f) CRIB4-GFP polar distribution in the germinated and elongated pollen tube. Scale bars = 10 μm.

Figure 3. 

Pollen dormancy was interrupted in the ren1 mutant under moist environments. (a), (b) Images and germination ratio of Col-0 and ren1-3 pollen 1 h after imbibition. Scale bars = 20 μm. Error bars represent SD. Statistical significance was determined by Student's t-test (*** p < 0.001, n = 3). (c) Fluorescence images of Col-0 and ren1-3 anther with or without pollen tubes stained with aniline blue. Scale bars = 100 μm. (d) Image of Col-0 and ren1-3 anther and stigma under high humidity conditions. The red arrow points to the pollen tube. Scale bars = 500 μm in the left images. Scale bars = 100 μm in the images of the middle and right.

Figure 4. 

Pollen dormancy was interrupted by the knockdown of GDI2 under moist environments. (a), (b) Images and germination ratio of Col-0, gdi2-ami2, and gdi2-ami8 pollen 1 hour after imbibition. Scale bars = 20 μm. Error bars represent SD. Statistical significance was determined by Student's t-test (*** p < 0.001, n = 3). (c) Fluorescence images of Col-0 and gdi2-ami anther with or without pollen tubes stained with aniline blue. Scale bars = 100 μm. (d) Image of Col-0, gdi2-ami2 and gdi2-ami8 anther and stigma under high humidity conditions. The red arrow points to the pollen tube. Scale bars = 500 μm in the left images. Scale bars = 100 μm in the images of middle and right.

Figure 5. 

Inhibition of pollen germination by REN1 overexpression. (a), (b) In vitro pollen germination of Col-0 and three REN1-OE transgenic lines (pREN1: GFP-REN1 CDS) after 3 h cultivation on PGM. Scale bars = 50 μm. (c) Fluorescence intensity assay of GFP-REN1 expressed pollens from three REN1-OE. (d) Germination ratio of Col-0 and REN1-OE pollens after 3 h cultivation on PGM. Statistical significance was determined by Student's t-test (** p < 0.01; *** p < 0.001), error bars represent SD, n = 3.

Figure 6. 

Inhibition of pollen germination by GDI2 overexpression. (a), (b) In vitro pollen germination of Col-0 and three GDI2 OE transgenic lines (pGDI2:GDI2-RFP) after 3 h cultivation on PGM. Scale bars = 50 μm. (c) Fluorescence intensity assay of GDI2-RFP expressed pollens from three GDI2 OE transgenic lines. (d) Germination ratio of Col-0 and GDI2-OE pollens after 3 h cultivation on PGM. Statistical significance was determined by Student's t-test (* p < 0.05; ** p < 0.01; *** p < 0.001), error bars represent SD, n = 3.

Figure 7. 

Pollen germination on stigma and silique of REN1 or GDI2 overexpression lines. (a) Germination of GFP labeled REN1-overexpressed pollens on Arabidopsis stigma. Scale bars = 25 μm. (b) Germination of GFP labeled GDI2-overexpressed pollens on Arabidopsis stigma. Scale bars = 25 μm. (c) The developed silique of self-fertilized wild type and REN1-overexpressed lines. Scale bars = 2 mm. (d) The developed silique of self fertilized wild type and GDI2-overexpressed lines. Scale bars = 2 mm.