The enhancement of phototropin-induced phototropic curvature in Arabidopsis occurs via a photoreversible phytochrome A-dependent modulation of auxin responsiveness

Abstract

The induction of phototropism in etiolated (dark-grown) seedlings exposed to an unidirectional pulse or extended irradiation with low fluence rate blue light (BL) requires the action of the phototropin (nph1) BL receptor. Although cryptochromes and phytochromes are not required for phototropic induction, these photoreceptors do modulate the magnitude of curvature resulting from phototropin activation. Modulatory increases in the magnitude of phototropic curvature have been termed "enhancement." Here, we show that phototropic enhancement is primarily a phytochrome A (phyA)-dependent red/far-red-reversible low fluence response. This phyA-dependent response is genetically separable from the basal phototropin-dependent response, as demonstrated by its retention under extended irradiation conditions in the nph4 mutant background, which normally lacks the basal BL-induced response. It is interesting that the nph4 mutants fail to exhibit the basal phototropin-dependent and phyA-dependent enhancement responses under limiting light conditions. Given that NPH4 encodes a transcriptional activator, auxin response factor 7 (ARF7), we hypothesize that the ultimate target(s) of phyA action during the phototropic enhancement response is a rate-limiting ARF-containing transcriptional complex in which the constituent ARFs can vary in identity or activity depending upon the irradiation condition.

Figure 1

RL-dependent enhancement of phototropism induced by long-term irradiation with BL in wild-type (●) and nph4 mutant (○) seedlings. A, Fluence response curves for RL-dependent enhancement of phototropism. Irradiation of 64-h-old etiolated seedlings with BL (unilateral; 0.5 μmol m⁻² s⁻¹) and RL (actinic from above; 1.6 μmol m⁻² s⁻¹) was initiated at the same time. BL was given for a total of 4 h, whereas RL exposure times varied (1–1,000 s) depending upon the desired fluence. At the end of the BL exposure, phototropic curvatures were measured. Data represent the mean response of at least 97 seedlings from three replicate experiments. Vertical bars represent the se values. B, Reciprocity relationships for RL-dependent enhancement of phototropism. Seedlings were handled as described above, except that fluence rates and exposure times of the RL exposure were varied to achieve a single fluence of 160 μmol m⁻². Curvatures are plotted relative to the exposure time of the RL irradiation. Data represent the mean response of at least 54 seedlings from three replicate experiments. Vertical error bars represent the se values.

Figure 2

Fluence response relationships for RL-dependent enhancement of BL-induced phototropism in nph4 phyA (A) and nph4 phyB (B) double mutant seedlings. Seedlings were handled as described in Figure 1A. Data represent the mean response of at least 63 seedlings from three replicate experiments. Vertical error bars represent the se values. ●, Wild type; ○, nph4; ⋄, phyA; ♦, nph4 phyA; □, phyB; ▪, nph4 phyB.

Figure 3

RL-dependent enhancement of phototropism induced by long-term irradiation with BL in nph4 seedlings overexpressing phyA. Seedlings were handled as described in Figure 1A. Data represent the mean response of at least 33 seedlings from two replicate experiments. Vertical error bars represent the se values. ●, Wild type; ○, nph4; ▵, phyA overexpressor (AOX); ▴, nph4 AOX.

Figure 4

Pulse-induced phototropism in wild-type and nph4 mutant seedlings. A, Fluence response curves for BL pulse-induced phototropism. Seventy-two-h-old etiolated seedlings were irradiated with five pulses of BL at the indicated fluence, and 2 h after the final pulse, curvatures were measured as described in “Materials and Methods.” Data represent the mean response of at least 28 seedlings from five replicate experiments. B, Fluence response curves for RL-dependent enhancement of BL pulse-induced phototropism. Experiments were as described for A, except that 2 h prior to BL irradiation (five pulses at 0.03 μmol m⁻² s⁻¹), seedlings were exposed to the indicated fluence of RL. Data represent the mean response from at least 15 seedlings from three replicate experiments. ●, Wild type; ○, nph4.