CONSTANS activates SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 through FLOWERING LOCUS T to promote flowering in Arabidopsis

Abstract

CONSTANS (CO) regulates flowering time by positively regulating expression of two floral integrators, FLOWERING LOCUS T (FT) and SUPPRESSOR OF OVEREXPRESSION OF CO 1 (SOC1), in Arabidopsis (Arabidopsis thaliana). FT and SOC1 have been proposed to act in parallel pathways downstream of CO based on genetic analysis using weak ft alleles, since ft soc1 double mutants showed an additive effect in suppressing the early flowering of CO overexpressor plants. However, this genetic analysis was inconsistent with the sequential induction pattern of FT and SOC1 found in inducible CO overexpressor plants. Hence, to identify genetic interactions of CO, FT, and SOC1, we carried out genetic and expression analyses with a newly isolated T-DNA allele of FT, ft-10. We found that ft-10 almost completely suppressed the early flowering phenotype of CO overexpressor plants, whereas soc1-2 partially suppressed the phenotype, suggesting that FT is the major output of CO. Expression of SOC1 was altered in gain- or loss-of-function mutants of FT, whereas expression of FT remained unchanged in gain- or loss-of-function mutants of SOC1, suggesting that FT positively regulates SOC1 to promote flowering. In addition, inactivation of FT caused down-regulation of SOC1 even in plants overexpressing CO, indicating that FT is required for SOC1 induction by CO. Taken together, these data suggest that CO activates SOC1 through FT to promote flowering in Arabidopsis.

Figure 1.

Flowering time of an ft-10 allele and its genetic interaction with 35S∷CO plants. A, Map of a T-DNA insertion 844 bases from the A of the start codon of FT; the allele was named ft-10. Black boxes indicate the four exons of FT. B, PCR genotyping of the ft-10 allele. Genotyping primer sets (JH2295, JH2296, and JH2297) were used to identify a homozygous line. Homozygous ft-10 plants produced a single band of 926 bp in size, whereas wild-type plants produced a band of 1,392 bp in size. M, One-kilobase ladder. C, An RT-PCR experiment showing that ft-10 was a strong knock-down allele. FT mRNA abundance in ft-10 plants was compared to ft-1 (Col), a point mutation allele of FT, and to wild-type Col plants. D and E, Flowering time and phenotypes of 35S∷CO, ft-10, 35S∷CO ft-10, soc1-2, 35S∷CO soc1-2, and wild-type Columbia (WT Col) plants under long-day conditions. ft-10 almost completely suppressed the early flowering phenotype of 35S∷CO plants, whereas soc1-2 partially suppressed the early flowering.

Figure 2.

Time-course expression of FT, SOC1, and AP1 in 35S∷FT, 35S∷SOC1, ft-10, soc1-2, and wild-type plants grown under long-day conditions. soc1-2 is a T-DNA-tagged allele of SOC1 (Lee et al., 2000). Gene expression levels of FT and SOC1 in 6-, 8-, 10-, and 12-d-old seedlings were measured. AP1 and UBQ10 were used as a molecular marker of floral transition and an internal positive control, respectively.

Figure 3.

Histochemical GUS analysis to examine genetic interactions between FT and SOC1. A, Comparison of GUS-staining pattern of 5-d-old seedlings of SOC1∷GUS (left) and 35S∷FT/+ SOC1∷GUS/+ plants (right). B to E, Close-up views of GUS-staining patterns in different tissues of SOC1∷GUS (left) and 35S∷FT/+ SOC1∷GUS/+ plants (right): apical region (B), cotyledon (C), hypocotyl (D), and root (E). An arrowhead indicates faint staining in the shoot apex of a SOC1∷GUS/+ seedling. F, Comparison of the staining pattern of 10-d-old seedlings of FT∷GUS (left) and 35S∷SOC1/+ FT∷GUS/+ plants (right). G to J, Close-up views of staining patterns observed in different tissues of FT∷GUS (left) and 35S∷SOC1/+ FT∷GUS/+ plants (right); apical region (G), cotyledon (H), hypocotyl (I), and root (J).

Figure 4.

Genetic interactions of FT and SOC1. Flowering time (A) and phenotypes (B) of single overexpressors, double overexpressors, and double mutants under long-day conditions are shown. All plants were in the Col background. soc1-101D is an activation-tagged line of SOC1 (Lee et al., 2000). The total number of primary leaves formed was scored in at least 10 plants to measure flowering time. The horizontal T-bars indicate the sd.

Figure 5.

Expression of SOC1 in 6-, 8-, 10-, and 12-d-old 35S∷CO ft-10, ft-10, 35S∷CO, and wild-type Col plants grown under long-day conditions. Semiquantitative RT-PCR was used to measure expression of SOC1. UBQ10 and CO were used for internal positive controls.

Figure 6.

Model for genetic interactions of CO, FT, and SOC1 in determination of flowering time in Arabidopsis. FT primarily mediates floral inductive inputs from CO. Although FT positively regulates SOC1, FT appears to have other downstream mediators in parallel with SOC1. Putative candidates for downstream mediators are FD and AP1.