Physiological and Genetic Characterization of End-of-Day Far-Red Light Response in Maize Seedlings

Developmental responses associated with end-of-day far-red light (EOD-FR) signaling were investigated in maize (Zea mays subspecies mays) seedlings. A survey of genetically diverse inbreds of temperate and tropical/semitropical origins, together with teosinte (Zea mays subspecies parviglumis) and a modern hybrid, revealed distinct elongation responses. A mesocotyl elongation response to the EOD-FR treatment was largely absent in the tropical/semitropical lines, but both hybrid and temperate inbred responses were of the same magnitude as in teosinte, suggesting that EOD-FR-mediated mesocotyl responses were not lost during the domestication or breeding process. The genetic architecture underlying seedling responses to EOD-FR was investigated using the intermated B73 × Mo17 mapping population. Among the different quantitative trait loci identified, two were consistently detected for elongation and responsiveness under EOD-FR, but none were associated with known light signaling loci. The central role of phytochromes in mediating EOD-FR responses was shown using a phytochromeB1 phytochromeB2 (phyB1 phyB2) mutant series. Unlike the coleoptile and first leaf sheath, EOD-FR-mediated elongation of the mesocotyl appears predominantly controlled by gibberellin. EOD-FR also reduced abscisic acid (ABA) levels in the mesocotyl for both the wild type and phyB1 phyB2 double mutants, suggesting a FR-mediated but PHYB-independent control of ABA accumulation. EOD-FR elongation responses were attenuated in both the wild type and phyB1 phyB2 double mutants when a chilling stress was applied during the dark period, concomitant with an increase in ABA levels. We present a model for the EOD-FR response that integrates light and hormonal control of seedling elongation.Plants utilize a complex network of photoreceptors to monitor the spectral quality, fluence, direction, and duration of light (Smith, 1995). These photosensory pigments include phytochromes that sense red light (R; 580–690 nm) and far-red light (FR; 690–800 nm) and the cryptochromes, phototropins, and zeitlupes for blue light (380–495 nm) and UV-A light (320–380 nm). The light reflected and transmitted by the vegetation creates a canopy characterized by reductions in both the R-to-FR ratio (R:FR) and the photosynthetically active radiation (400–700 nm). This light environment induces adaptive biochemical and morphological responses known as the shade avoidance syndrome (Smith and Whitelam, 1997). These responses can be induced early in development, before canopy closure, through FR reflected from adjacent neighbor plants (Ballare et al., 1990) or from low-lying weeds (Rajcan and Swanton, 2001), which can negatively impact yields in maize (Zea mays subspecies mays; Rajcan et al., 2004), even if only present early in the growing season (Liu et al., 2009).R:FR signals are transduced by the phytochrome family of photoreceptors (Franklin and Whitelam, 2007b). In rice (Oryza sativa) and sorghum (Sorghum bicolor), three genes constitute the phytochrome family: PhytochromeA (PhyA), PhytochromeB (PhyB), and PhytochromeC (PhyC). In maize, an ancient alloploidization has doubled the family size to six: PhyA1, PhyA2, PhyB1, PhyB2, PhyC1, and PhyC2 (Sheehan et al., 2004). Although many similarities are apparent between maize and Arabidopsis (Arabidopsis thaliana) light response, there are significant differences between members of the phytochrome gene family in copy number and selection pressures that have resulted in the divergence of phytochrome signaling networks (Sawers et al., 2005; Sheehan et al., 2007). Thus far, only three phytochrome mutants have been characterized in maize: elongated mesocotyl1 (elm1), phyB1, and phyB2. The elm1 mutant carries a mutation in phytochromobilin synthase, necessary for the biosynthesis of the chromophore common to all phytochromes (Sawers et al., 2004). The mutation severely reduces the total phytochrome pool, but the weak nature of the allele enables a partial responsiveness to R and FR (Markelz et al., 2003). At maturity, elm1 mutants have elongated internodes, pale green leaves, and flower early (Sawers et al., 2002). Mutations at phyB1 and phyB2 also impair light signal transduction. At maturity, both PHYB1 and PHYB2 contribute to plant height, stem diameter, and sheath-internode length, but PHYB2 predominates in the control of flowering (Sheehan et al., 2007). Like the sorghum and rice phyB mutants (Childs et al., 1997; Takano et al., 2005; Kebrom et al., 2010), both elm1 and phyB1 phyB2 double mutants constitutively display several traits associated with low R:FR response (Sawers et al., 2002; Markelz et al., 2003; Sheehan et al., 2007).In Arabidopsis, R/FR-mediated responses are developmentally complex and involve the PIF proteins (Duek and Fankhauser, 2005) and many hormones including auxins (Tao et al., 2008), ethylene (Khanna et al., 2007), jasmonate (Moreno et al., 2009), and GA (Djakovic-Petrovic et al., 2007). In particular, there is a direct interaction between PIF and DELLA proteins that govern phytochrome-mediated elongation (de Lucas et al., 2008; Feng et al., 2008; Lorrain et al., 2008). DELLA proteins also regulate FR inhibition of germination by abscisic acid (ABA; Piskurewicz et al., 2009), suggesting an interaction between the PIFs and ABA signaling. Complex cross talk between light and temperature has also been reported (Franklin, 2009). In Arabidopsis, colder temperatures can repress the early-flowering phenotype of the phyB mutant (Halliday et al., 2003). These studies suggest a complex interplay between light, hormone, and temperature to fine-tune the elongation response.The end-of-day far-red light (EOD-FR) treatment consists of a pulse of FR given at subjective dusk (Kasperbauer, 1971) and triggers a circadian clock-gated response (Salter et al., 2003). EOD-FR treatments result in a minimal pool of active Pfr during the dark period (Fankhauser and Casal, 2004), and plants submitted to daily treatments display similar developmental responses to those elicited by a continuous photoperiod with low R:FR (Smith, 1994). One of the key features that contributed to the discovery of the phytochromes is the photoreversibility of the response (Borthwick et al., 1952). These low-fluence responses (LFRs) are induced or repressed by alternating R and FR treatments (Mancinelli, 1994). The LFR nature of EOD-FR in maize was previously demonstrated in 5-d-old mesocotyl and coleoptile tissues (Gorton and Briggs, 1980). The EOD-FR treatment offers several advantages over growing plants in continuous low R:FR, including exposing plants to relatively brief treatment periods, thus potentially reducing genotype × environment effects. It also facilitates kinetic assays of phytochrome response, as treatments are limited to a single point in the diurnal cycle and can be delivered at any stage in plant development. Finally, as relatively low fluences of light are needed to saturate EOD-FR responses, large populations of seedlings can be screened without the need for large numbers of FR light-emitting diodes (LEDs) or sophisticated light chambers.Here, we have examined EOD-FR-mediated responses in maize and its closest relative, teosinte (Zea mays subspecies parviglumis). A survey of genetically diverse maize and teosinte accessions revealed extensive tissue-specific variations in mesocotyl, coleoptile, and first leaf sheath elongation. EOD-FR responses were greatly attenuated in tropical/semitropical (TS) accessions but present in teosinte, temperate inbreds, and a modern commercial hybrid, suggesting that the EOD-FR response is plastic in Z. mays. To investigate the genetic regulation underlying seedling responses to EOD-FR, we performed a quantitative trait locus (QTL) analysis using the intermated B73 × Mo17 (IBM) recombinant inbred population. We identified several QTLs that regulate mesocotyl and first leaf sheath response to EOD-FR and show that these QTLs mediate tissue-specific responses. The phyB1 phyB2 mutant series was also evaluated, indicating that the two PhyB paralogs are largely redundant in mediating the EOD-FR response. Pharmacological assays revealed a major role for GA in promoting mesocotyl, but not coleoptile or first leaf sheath, elongation in response to EOD-FR treatments. In contrast, EOD-FR reduced mesocotyl ABA levels. A chill treatment (10°C) applied during dark breaks attenuated EOD-FR elongation responses. Based on these observations, we discuss a model that integrates temperature, light, and hormonal inputs in the regulation of mesocotyl elongation.