Mutant analyses define multiple roles for phytochrome C in Arabidopsis photomorphogenesis

Abstract

The analysis of Arabidopsis mutants deficient in the A, B, D, and E phytochromes has revealed that each of these phytochrome isoforms has both distinct and overlapping roles throughout plant photomorphogenesis. Although overexpression studies of phytochrome C (phyC) have suggested photomorphogenic roles for this receptor, conclusive evidence of function has been lacking as a result of the absence of mutants in the PHYC locus. Here, we describe the isolation of a T-DNA insertion mutant of phyC (phyC-1), the subsequent creation of mutant lines deficient in multiple phytochrome combinations, and the physiological characterization of these lines. In addition to operating as a weak red light sensor, phyC may perform a significant role in the modulation of other photoreceptors. phyA and phyC appear to act redundantly to modulate the phyB-mediated inhibition of hypocotyl elongation in red light and to function together to regulate rosette leaf morphology. In addition, phyC performs a significant role in the modulation of blue light sensing. Several of these phenotypes are supported by the parallel analysis of a quadruple mutant deficient in phytochromes A, B, D, and E, which thus contains only active phyC. Together, these data suggest that phyC has multiple functions throughout plant development that may include working as a coactivator with other phytochromes and the cryptochrome blue light receptors.

Figure 1.

Mapping and Analysis of phyC-1 Mutation. (A) The phyC-1 mutation results from a T-DNA insertion at the 5′ end of the third exon of the PHYC gene. The position of the insertion point was mapped to 3292 bp. Boxes represent exons, and lines represent introns. Arrows 1 and 2 represent the positions of primers used for RT-PCR analysis. (B) PCR analyses of PHYC using genomic DNA and cDNA from light-grown seedlings of phyD and phyCD plants. Intron-spanning primers produced a 510-bp product from all genomic samples and a 374-bp product from phyD RT samples only.

Figure 2.

Rosette Leaf Phenotypes of phy Mutant Combinations at 28 Days. (A) Mature plant phenotypes at 28 days of phyD and phyCD growth under 8-h-light/16-h-dark cycles of white light (100 μmol·m⁻²·s⁻¹) at 22°C. (B) Mature plant phenotypes of phy mutant combinations grown under similar conditions with 16-h-light/8-h-dark cycles. WT, wild type. (C) Length of leaf 8 in plants grown under 16-h-light/8-h-dark cycles (mean ± se; n = 20).

Figure 3.

Primary Leaf Areas of phy Mutant Combinations at 18 Days. Seedlings were grown under continuous white light (100 μmol·m⁻²·s⁻¹) at 22°C (mean ± se; n = 20). WT, wild type.

Figure 4.

Hypocotyl Lengths of phy Mutant Combinations Grown under Different Fluence Rates of R. (A) Seedling phenotypes of phy mutant combinations grown for 3 days under Rc at 0.1 μmol·m⁻²·s⁻¹ at 22°C. WT, wild type. (B) to (F) Hypocotyl lengths of phy mutant combinations grown under different fluence rates of R. Seedlings were grown under continuous irradiation at 22°C for 3 days (mean ± se; n = 40).

Figure 5.

Cotyledon Angles in phy Mutant Combinations at 3 Days. Seedlings were grown at 22°C under continuous R at 3 μmol·m⁻²·s⁻¹ (mean ± se; n = 20). WT, wild type.

Figure 6.

Hypocotyl Lengths of phy Mutant Combinations Grown under Different Fluence Rates of B. Seedlings were grown under continuous irradiation at 22°C for 3 days (mean ± se; n = 40). WT, wild type.