Isolation and Characterization of phyC Mutants in Arabidopsis Reveals Complex Crosstalk between Phytochrome Signaling Pathways

Abstract

Studies with mutants in four members of the five-membered Arabidopsis phytochrome (phy) family (phyA, phyB, phyD, and phyE) have revealed differential photosensory and/or physiological functions among them, but identification of a phyC mutant has proven elusive. We now report the isolation of multiple phyC mutant alleles using reverse-genetics strategies. Molecular analysis shows that these mutants have undetectable levels of phyC protein, suggesting that they are null for the photoreceptor. phyC mutant seedlings were indistinguishable from wild-type seedlings under constant far-red light (FRc), and phyC deficiency had no effect in the phyA mutant background under FRc, suggesting that phyC does not participate in the control of seedling deetiolation under FRc. However, when grown under constant red light (Rc), phyC seedlings exhibited a partial loss of sensitivity, observable as longer hypocotyls and smaller cotyledons than those seen in the wild type. Although less severe, this phenotype resembles the effect of phyB mutations on photoresponsiveness, indicating that both photoreceptors function in regulating seedling deetiolation in response to Rc. On the other hand, phyB phyC double mutants did not show any apparent decrease in sensitivity to Rc compared with phyB seedlings, indicating that the phyC mutation in the phyB-deficient background does not have an additive effect. These results suggest that phyB is necessary for phyC function. This functional dependence correlates with constitutively lower levels of phyC observed in the phyB mutant compared with the wild type, a decrease that seems to be regulated post-transcriptionally. phyC mutants flowered early when grown in short-day photoperiods, indicating that phyC plays a role in the perception of daylength. phyB phyC double mutant plants flowered similarly to phyB plants, indicating that in the phyB background, phyC deficiency does not further accelerate flowering. Under long-day photoperiods, phyA phyC double mutant plants flowered later than phyA plants, suggesting that phyC is able to promote flowering in the absence of phyA. Together, these results suggest that phyC is involved in photomorphogenesis throughout the life cycle of the plant, with a photosensory specificity similar to that of phyB/D/E and with a complex pattern of differential crosstalk with phyA and phyB in the photoregulation of multiple developmental processes.

Evolutionary Studies Illuminate the Structural-Functional Model of Plant Phytochromes

“…Perhaps consistent with this is the fact that null mutants of these loci in Arabidopsis were not easily identified in genetic screens and the phenotypes of the monogenic mutants are not readily apparent (Aukerman et al, 1997;Devlin et al, 1998;Franklin et al, 2003;Monte et al, 2003). However, PHYC, D, and E have been retained much longer than the expected half-life of duplicate genes of ;3 to 7 million years (Lynch and Conery, 2000).…”

“…Thus, phyB also is the primary mediator of shade avoidance, with additional light-stable forms in Arabidopsis, phyD and phyE, making lesser (Franklin, 2008) though perhaps important fine-tuning (Ballaré , 2009) contributions. The fourth light-stable form is phyC, a weak R sensor that functions in leaf development and perception of photoperiod Monte et al, 2003). As discussed later in the review, recent studies have revealed roles for phyC-E that were not apparent from the initial characterization of the phyC-E null mutants.…”

“…Together, phyD and phyE may be important for fine-tuning the magnitude of responses to the R:FR ratio in complex light environments (Ballaré , 2009). phyC appears to function as a weak R sensor in the control of seedling deetiolation, photoperiod perception, and in the modulation of other photoreceptors Monte et al, 2003).…”

Dynamic Antagonism between Phytochromes and PIF Family Basic Helix-Loop-Helix Factors Induces Selective Reciprocal Responses to Light and Shade in a Rapidly Responsive Transcriptional Network in Arabidopsis

“…Hypocotyl length and cotyledon opening were measured from at least 20 seedlings as described (Monte et al, 2003;Leivar et al, 2008a). Data were analyzed using Tukey's multiple comparison test (GraphPad Prism software) and/or Student's t test as indicated.…”

“…For RNA gel blot analysis, total RNA extraction, loading, and hybridization conditions were as described (Monte et al, 2003). Specific probe for PIF3 was prepared as described (Monte et al, 2004).…”