Arabidopsis TERMINAL FLOWER1 is involved in the regulation of flowering time and inflorescence development through transcriptional repression

Abstract

TERMINAL FLOWER1 (TFL1) is a key regulator of flowering time and the development of the inflorescence meristem in Arabidopsis thaliana. TFL1 and FLOWERING LOCUS T (FT) have highly conserved amino acid sequences but opposite functions. For example, FT promotes flowering and TFL1 represses it; FT-overexpressing plants and TFL1 loss-of-function mutants have a similar phenotype production of terminal flowers in the shoot apex. FT is believed to function in a transcriptional activator complex by interacting with FD. Here, we demonstrate that TFL1 is involved in the transcriptional repression of genes that are activated by FT. We analyzed transgenic plants overexpressing TFL1 fused to a transcriptional repressor domain (TFL1-SRDX) or an activator domain (TFL1-VP16). Plants carrying 35S:TFL1-SRDX showed delayed flowering similar to 35S:TFL1 plants, and plants carrying 35S:TFL1-VP16 showed an early flowering phenotype and produced terminal flowers. Furthermore, the tfl1 and 35S:TFL1-VP16 plant phenotypes were strongly suppressed by the fd mutation, and TFL1 interacted with FD in the cell nucleus, as shown by bimolecular fluorescence complementation experiments. We conclude that TFL1 negatively modulates the FD-dependent transcription of target genes to fine-tune flowering time and the development of the inflorescence meristem.

Figure 1.

The Flowering and Inflorescence Phenotypes of the tfl1 Mutant Alleles. (A) and (B) Flowering time of tfl1 alleles in the Col background (tfl1-1, tfl1-11, tfl1-13, tfl1-14, and tfl1-17) under LD (16 h light/8 h dark; white bars) and SD (10 h light/14 h dark; gray bars) at 22°C (A) and under LD conditions at 16°C (B). The vertical axis indicates the number of rosette leaves at flowering. (C) As an indicator of the terminal flower phenotype, we counted the number of flower buds between the last lateral shoot and the terminal flower in the primary inflorescence of tfl1 mutants. Terminal flowers were not included. More than 20 flower buds were produced without terminal flower in wild-type plants (Col). Error bars represent ± se. The difference between Col and tfl1-11, tfl1-13, and tfl1-14 under SD is not statistically significant. Other differences between Col and tfl1 alleles are significant (P < 0.05, n = 6 to 16), as indicated by statistical analysis of multiple independent replicates using the two-tailed multiple t test with Bonferroni correction following analysis of variance (ANOVA).

Figure 2.

35S:TFL1-VP16 and 35S:TFL1-SRDX Phenotypes in the Col and tfl1-17 Background. (A) From left to right, 6-week-old plants of Col, tfl1-17, 35S:TFL1-VP16/Col, and 35S:TFL1-VP16/tfl1-17. Inset shows top view of the inflorescence of 35S:TFL1-VP16/Col with a terminal flower (TF). (B) Two-month-old 35S:TFL1-SRDX/Col plant. (C) Two-month-old 35S:TFL1-SRDX/tfl1-17 plant. (D) Flower of 35S:TFL1-SRDX showing bract-like sepals and proliferating inner flowers. (E) 35S:TFL1-SRDX inflorescence with elongated peduncle of the secondary flower resulting in a lateral shoot-like organ. (F) Two-week-old 35S:FT-VP16/ft-101 plant. Bars = 1 cm in (A), (B), (C), (E), and (F) and 1 mm in the inset of (A) and (D). [See online article for color version of this figure.]

Figure 3.

Expression of Meristem Identity Genes in 35S:TFL1-SRDX and 35S:TFL1-VP16 Plants. Left panels: Gene expression in 35S:TFL1-SRDX plants at 22°C. Plants were transferred from SD to LD at day 0. For RNA isolation, the shoot apical region of 10 to 15 plants were collected at 7 d before and at 0, 2, 6, 10, and 14 d after the transfer. Right panels: Gene expression in 35S:TFL1-VP16 plants under LD at 16°C. RNAs were isolated from the shoot apical regions of 10 to 15 plants that were 4, 5, 6, and 7 weeks old. Expression levels were determined by real-time quantitative PCR; the ratio of each gene expression level relative to CLV3 expression is shown. Results from three biological replicates are shown as means ± se. Since harvested plants were not completely synchronized regarding phase transition, both between replicates and within each series, some data sets show high variances (large error bars).

Figure 4.

Phenotypes of the fd-1 tfl1-2 Double Mutant and fd-1 Plants Carrying 35S:TFL1-VP16. Three-week-old tfl1-2 (A), 9-week-old fd-1 (B), 9-week-old fd-1 tfl1-2 (C), and 9-week-old 35S:TFL1-VP16/fd-1 (D) plants are shown. Top views of the inflorescence of 11-week-old fd-1 (E) and its higher magnification ([E] inset), 11-week-old fd-1 tfl1-2 (F), and 11-week-old 35S:TFL1-VP16/fd-1 (G) plants are also shown. Inflorescences of ap1-7 (H) and ap1-7 tfl1-17 double mutants (I). Bars 1 cm in (A) to (D), 1 mm in (E) to (I), and 0.1 mm in the inset of (E). TF, terminal flower. [See online article for color version of this figure.]

Figure 5.

BiFC Analysis of Protein Interactions between TFL1 or FT and FD, and TFL1 Localization in N. benthamiana Leaf Epidermis Cells. G, green channel image showing fluorescence of YFP (BiFC) or GFP; BF, bright-field image; Merged, merged image of G and BF. TFL1 and FD, coexpression of 35S:TFL1-cYFP and 35S:nYFP-FD; TFL1-GFP, expression of 35S:TFL1-GFP; FT and FD, coexpression of 35S:FT-cYFP and 35S:nYFP-FD. Bars = 50 μm. [See online article for color version of this figure.]