An Arabidopsis flavonoid transporter is required for anther dehiscence and pollen development

Abstract

FLOWER FLAVONOID TRANSPORTER (FFT) encodes a multidrug and toxin efflux family transporter in Arabidopsis thaliana. FFT (AtDTX35) is highly transcribed in floral tissues, the transcript being localized to epidermal guard cells, including those of the anthers, stigma, siliques and nectaries. Mutant analysis demonstrates that the absence of FFT transcript affects flavonoid levels in the plant and that the altered flavonoid metabolism has wide-ranging consequences. Root growth, seed development and germination, and pollen development, release and viability are all affected. Spectrometry of mutant versus wild-type flowers shows altered levels of a glycosylated flavonol whereas anthocyanin seems unlikely to be the substrate as previously speculated. Thus, as well as adding FFT to the incompletely described flavonoid transport network, it is found that correct reproductive development in Arabidopsis is perturbed when this particular transporter is missing.

Fig. 1.

Flavonoid biosynthesis pathways and transport steps, in outline. Enzymes and mutants are marked if mentioned in the text. PAL, phenylalanine ammonia lyase; CHS, chalcone synthase; CHI, chalcone isomerase; F3'H, flavonoid 3’ hydroxylase; DFR, dihydroflavonol 4-reductase; tt, transparent testa.

Fig. 2.

X-gluc staining of GUS–FFT–promoter-transformed plants (each row, left–right). (a) Seedling roots: day 4 post-stratification; day 7 root tip, elongation zone, and developing lateral root; day 12; day 17. (b) Aerial tissues: (row 1) cotyledon guard cells (GC); mature leaf hydathode GC; nectaries; (row 2) inflorescence apex; silique apex; close-up of papillae and stigma GC; (row 3) anther GC; developing seed in siliques; developing silique.

Fig. 3.

(a) Germination of the fft-1 (- -) mutant versus Col0 (—). (b) Seed size in Col0 and the mutant. (c) Ruthenium red staining of imbibing seed mucilage. (d) Proportion of imbibing Col0, fft-1 and complemented fft-1 seeds showing complete mucilage when stained with ruthenium red. (This figure is available in colour at JXB online.)

Fig. 4.

Mutant fft-1 (- -) roots grow more quickly than Col0 (—) up to ∼2 weeks. (a) No sucrose: day 6–12, P=0.002–0.003 (n=70–81 for each time point). Inset: Col0 and fft-1 seedlings at day 4 post-stratification. (b) 1% sucrose: day 6–12, P <0.0001–0.001 (n=60–97 for each time point). (c) The fft-1 mutant complemented with 35S-At4g25640 returns to wild-type growth characteristics. (This figure is available in colour at JXB online.)

Fig. 5.

(a) Alexander's staining showing (left–right) lack of dehiscence, no pollen on the stigma, and reduced proportion of viable pollen in anthers in fft-1 (second row) versus Col0 (top row). Top right: pollen production and dehiscence in complemented fft-1. (b) Variable success of silique production in fft-1 versus Col0 and complemented fft-1.

Fig. 6.

(a) Col0 versus fft-1 mean ratio of flavonoid (580 nm) and chlorophyll fluorescence (690 nm; Bolhar-Nordenkampf et al., 1989). Inset: example of the anther guard cell fluorescence scanned for analysis as viewed with the confocal microscope. (b) Examples of replicate fluorescence emission spectra from guard cells in Col0 and fft-1 anthers (background subtracted; normalized to 690 nm). (This figure is available in colour at JXB online.)