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The Arabidopsis thaliana F-box protein FBL17 is essential for progression through the second mitosis during pollen development.

Gusti A, Baumberger N, Nowack M, Pusch S, Eisler H, Potuschak T, De Veylder L, Schnittger A, Genschik P - PLoS ONE (2009)

Bottom Line: FBL17 expression is strongly enhanced in plants co-expressing E2Fa and DPa, transcription factors that promote S-phase entry.Seed abortion can, however, be overcome by mutations in FIE, a component of the Polycomb group complex, overall resembling loss-of-function mutations in the A. thaliana cyclin-dependent kinase CDKA;1.Finally we identified ASK11, as an SKP1-like partner protein of FBL17 and discuss a possible mechanism how SCF(FBL17) may regulate cell division during male gametogenesis.

View Article: PubMed Central - PubMed

Affiliation: Institut de Biologie Moléculaire des Plantes du CNRS, Université de Strasbourg, Strasbourg, France.

ABSTRACT
In fungi and metazoans, the SCF-type Ubiquitin protein ligases (E3s) play a critical role in cell cycle regulation by degrading negative regulators, such as cell cycle-dependent kinase inhibitors (CKIs) at the G1-to-S-phase checkpoint. Here we report that FBL17, an Arabidopsis thaliana F-box protein, is involved in cell cycle regulation during male gametogenesis. FBL17 expression is strongly enhanced in plants co-expressing E2Fa and DPa, transcription factors that promote S-phase entry. FBL17 loss-of-function mutants fail to undergo pollen mitosis II, which generates the two sperm cells in mature A. thaliana pollen. Nonetheless, the single sperm cell-like cell in fbl17 mutants is functional but will exclusively fertilize the egg cell of the female gametophyte, giving rise to an embryo that will later abort, most likely due to the lack of functional endosperm. Seed abortion can, however, be overcome by mutations in FIE, a component of the Polycomb group complex, overall resembling loss-of-function mutations in the A. thaliana cyclin-dependent kinase CDKA;1. Finally we identified ASK11, as an SKP1-like partner protein of FBL17 and discuss a possible mechanism how SCF(FBL17) may regulate cell division during male gametogenesis.

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Pollen phenotype of fbl17 mutants and FBL17 expression during male gametogenesis.fbl17 (A) and wild type (B) mature pollen viability test by coloration with Alexander's stain. The purple staining indicates that the grains are viable. Dehiscent pollen of qrt1-1−/− (C) and qrt1-1−/−, fbl17-1+/− (D) stained with DAPI and observed under UV fluorescence. The four pollen grains of the tetrad show two densely stained sperm cell nuclei and one large diffuse vegetative cell nuclei in qrt-1 mutants, whereas two pollen grains of the tetrad show only a single germ cell nuclei in qrt1-1,fbl17-1+/− double mutants. (E, F) transmitted light picture of C and D. (G–L) Expression of the HT10 gene in the fbl17 mutant pollen. Expression of the HTR10-mRFPprotein under the HTR10 promoter in fbl17-1 (H) and wild type (K) pollen, counterstained with DAPI (G, fbl17-1; J, wild type). (I, L) transmitted light pictures of G and J. (A, B) bar = 100 µm; (C–L) bar = 10 µm. (M–U) Promoter-GUS analysis of FBL17 expression in pollen. (M, P, S) DAPI staining is applied to reveal the developmental stage of the pollen grain. (N–U) X-Gluc histochemical staining of pFBL17∶GUS (N, Q, F) and non-transformed Col-0 (O, R, U) pollen grains. Bars = 10 µm (V) DNA content measurement of wild type germinative cell nuclei at prophase (n = 9; DNA = 2C), wild type sperm cell nuclei at telophase (n = 16), WT sperm cell nuclei at anthesis (n = 111) in comparison to the unique germ-cell like nuclei of fbl17 pollen (n = 77). Error bars = standard error mean.
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pone-0004780-g002: Pollen phenotype of fbl17 mutants and FBL17 expression during male gametogenesis.fbl17 (A) and wild type (B) mature pollen viability test by coloration with Alexander's stain. The purple staining indicates that the grains are viable. Dehiscent pollen of qrt1-1−/− (C) and qrt1-1−/−, fbl17-1+/− (D) stained with DAPI and observed under UV fluorescence. The four pollen grains of the tetrad show two densely stained sperm cell nuclei and one large diffuse vegetative cell nuclei in qrt-1 mutants, whereas two pollen grains of the tetrad show only a single germ cell nuclei in qrt1-1,fbl17-1+/− double mutants. (E, F) transmitted light picture of C and D. (G–L) Expression of the HT10 gene in the fbl17 mutant pollen. Expression of the HTR10-mRFPprotein under the HTR10 promoter in fbl17-1 (H) and wild type (K) pollen, counterstained with DAPI (G, fbl17-1; J, wild type). (I, L) transmitted light pictures of G and J. (A, B) bar = 100 µm; (C–L) bar = 10 µm. (M–U) Promoter-GUS analysis of FBL17 expression in pollen. (M, P, S) DAPI staining is applied to reveal the developmental stage of the pollen grain. (N–U) X-Gluc histochemical staining of pFBL17∶GUS (N, Q, F) and non-transformed Col-0 (O, R, U) pollen grains. Bars = 10 µm (V) DNA content measurement of wild type germinative cell nuclei at prophase (n = 9; DNA = 2C), wild type sperm cell nuclei at telophase (n = 16), WT sperm cell nuclei at anthesis (n = 111) in comparison to the unique germ-cell like nuclei of fbl17 pollen (n = 77). Error bars = standard error mean.

Mentions: A failure to transmit mutant alleles through the pollen can be caused by defects in pollen viability and/or development, germination, pollen tube growth or even fertilization. To test for pollen viability, we used Alexander's staining [30]. Similarly to mature pollen grains from wild type plants, pollen from heterozygous fbl17-1 and fbl17-2 mutants appeared full, round and red-stained when treated with Alexander's stain (Figure 2A,B), showing that the mutation does not affect pollen viability.


The Arabidopsis thaliana F-box protein FBL17 is essential for progression through the second mitosis during pollen development.

Gusti A, Baumberger N, Nowack M, Pusch S, Eisler H, Potuschak T, De Veylder L, Schnittger A, Genschik P - PLoS ONE (2009)

Pollen phenotype of fbl17 mutants and FBL17 expression during male gametogenesis.fbl17 (A) and wild type (B) mature pollen viability test by coloration with Alexander's stain. The purple staining indicates that the grains are viable. Dehiscent pollen of qrt1-1−/− (C) and qrt1-1−/−, fbl17-1+/− (D) stained with DAPI and observed under UV fluorescence. The four pollen grains of the tetrad show two densely stained sperm cell nuclei and one large diffuse vegetative cell nuclei in qrt-1 mutants, whereas two pollen grains of the tetrad show only a single germ cell nuclei in qrt1-1,fbl17-1+/− double mutants. (E, F) transmitted light picture of C and D. (G–L) Expression of the HT10 gene in the fbl17 mutant pollen. Expression of the HTR10-mRFPprotein under the HTR10 promoter in fbl17-1 (H) and wild type (K) pollen, counterstained with DAPI (G, fbl17-1; J, wild type). (I, L) transmitted light pictures of G and J. (A, B) bar = 100 µm; (C–L) bar = 10 µm. (M–U) Promoter-GUS analysis of FBL17 expression in pollen. (M, P, S) DAPI staining is applied to reveal the developmental stage of the pollen grain. (N–U) X-Gluc histochemical staining of pFBL17∶GUS (N, Q, F) and non-transformed Col-0 (O, R, U) pollen grains. Bars = 10 µm (V) DNA content measurement of wild type germinative cell nuclei at prophase (n = 9; DNA = 2C), wild type sperm cell nuclei at telophase (n = 16), WT sperm cell nuclei at anthesis (n = 111) in comparison to the unique germ-cell like nuclei of fbl17 pollen (n = 77). Error bars = standard error mean.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2651519&req=5

pone-0004780-g002: Pollen phenotype of fbl17 mutants and FBL17 expression during male gametogenesis.fbl17 (A) and wild type (B) mature pollen viability test by coloration with Alexander's stain. The purple staining indicates that the grains are viable. Dehiscent pollen of qrt1-1−/− (C) and qrt1-1−/−, fbl17-1+/− (D) stained with DAPI and observed under UV fluorescence. The four pollen grains of the tetrad show two densely stained sperm cell nuclei and one large diffuse vegetative cell nuclei in qrt-1 mutants, whereas two pollen grains of the tetrad show only a single germ cell nuclei in qrt1-1,fbl17-1+/− double mutants. (E, F) transmitted light picture of C and D. (G–L) Expression of the HT10 gene in the fbl17 mutant pollen. Expression of the HTR10-mRFPprotein under the HTR10 promoter in fbl17-1 (H) and wild type (K) pollen, counterstained with DAPI (G, fbl17-1; J, wild type). (I, L) transmitted light pictures of G and J. (A, B) bar = 100 µm; (C–L) bar = 10 µm. (M–U) Promoter-GUS analysis of FBL17 expression in pollen. (M, P, S) DAPI staining is applied to reveal the developmental stage of the pollen grain. (N–U) X-Gluc histochemical staining of pFBL17∶GUS (N, Q, F) and non-transformed Col-0 (O, R, U) pollen grains. Bars = 10 µm (V) DNA content measurement of wild type germinative cell nuclei at prophase (n = 9; DNA = 2C), wild type sperm cell nuclei at telophase (n = 16), WT sperm cell nuclei at anthesis (n = 111) in comparison to the unique germ-cell like nuclei of fbl17 pollen (n = 77). Error bars = standard error mean.
Mentions: A failure to transmit mutant alleles through the pollen can be caused by defects in pollen viability and/or development, germination, pollen tube growth or even fertilization. To test for pollen viability, we used Alexander's staining [30]. Similarly to mature pollen grains from wild type plants, pollen from heterozygous fbl17-1 and fbl17-2 mutants appeared full, round and red-stained when treated with Alexander's stain (Figure 2A,B), showing that the mutation does not affect pollen viability.

Bottom Line: FBL17 expression is strongly enhanced in plants co-expressing E2Fa and DPa, transcription factors that promote S-phase entry.Seed abortion can, however, be overcome by mutations in FIE, a component of the Polycomb group complex, overall resembling loss-of-function mutations in the A. thaliana cyclin-dependent kinase CDKA;1.Finally we identified ASK11, as an SKP1-like partner protein of FBL17 and discuss a possible mechanism how SCF(FBL17) may regulate cell division during male gametogenesis.

View Article: PubMed Central - PubMed

Affiliation: Institut de Biologie Moléculaire des Plantes du CNRS, Université de Strasbourg, Strasbourg, France.

ABSTRACT
In fungi and metazoans, the SCF-type Ubiquitin protein ligases (E3s) play a critical role in cell cycle regulation by degrading negative regulators, such as cell cycle-dependent kinase inhibitors (CKIs) at the G1-to-S-phase checkpoint. Here we report that FBL17, an Arabidopsis thaliana F-box protein, is involved in cell cycle regulation during male gametogenesis. FBL17 expression is strongly enhanced in plants co-expressing E2Fa and DPa, transcription factors that promote S-phase entry. FBL17 loss-of-function mutants fail to undergo pollen mitosis II, which generates the two sperm cells in mature A. thaliana pollen. Nonetheless, the single sperm cell-like cell in fbl17 mutants is functional but will exclusively fertilize the egg cell of the female gametophyte, giving rise to an embryo that will later abort, most likely due to the lack of functional endosperm. Seed abortion can, however, be overcome by mutations in FIE, a component of the Polycomb group complex, overall resembling loss-of-function mutations in the A. thaliana cyclin-dependent kinase CDKA;1. Finally we identified ASK11, as an SKP1-like partner protein of FBL17 and discuss a possible mechanism how SCF(FBL17) may regulate cell division during male gametogenesis.

Show MeSH
Related in: MedlinePlus