UV-B Perceived by the UVR8 Photoreceptor Inhibits Plant Thermomorphogenesis

Abstract

Small increases in ambient temperature can elicit striking effects on plant architecture, collectively termed thermomorphogenesis [1]. In Arabidopsis thaliana, these include marked stem elongation and leaf elevation, responses that have been predicted to enhance leaf cooling [2-5]. Thermomorphogenesis requires increased auxin biosynthesis, mediated by the bHLH transcription factor PHYTOCHROME-INTERACTING FACTOR 4 (PIF4) [6-8], and enhanced stability of the auxin co-receptor TIR1, involving HEAT SHOCK PROTEIN 90 (HSP90) [9]. High-temperature-mediated hypocotyl elongation additionally involves localized changes in auxin metabolism, mediated by the indole-3-acetic acid (IAA)-amido synthetase Gretchen Hagen 3 (GH3).17 [10]. Here we show that ultraviolet-B light (UV-B) perceived by the photoreceptor UV RESISTANCE LOCUS 8 (UVR8) [11] strongly attenuates thermomorphogenesis via multiple mechanisms inhibiting PIF4 activity. Suppression of thermomorphogenesis involves UVR8 and CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1)-mediated repression of PIF4 transcript accumulation, reducing PIF4 abundance. UV-B also stabilizes the bHLH protein LONG HYPOCOTYL IN FAR RED (HFR1), which can bind to and inhibit PIF4 function. Collectively, our results demonstrate complex crosstalk between UV-B and high-temperature signaling. As plants grown in sunlight would most likely experience concomitant elevations in UV-B and ambient temperature, elucidating how these pathways are integrated is of key importance to the understanding of plant development in natural environments.

Keywords: Arabidopsis; HFR1; PIF4; UV-B; UVR8; auxin; high temperature; hypocotyl.

Figure 1

UV-B Perceived by UVR8 Inhibits High-Temperature-Induced Architectural Adaptations in Arabidopsis (A) Hypocotyl lengths of Ler and uvr8-1 seedlings grown in continuous light for 3 days at 20°C, before transfer to 20°C, 28°C, 20°C + UV-B, or 28°C + UV-B for a further 4 days. Data represent mean length (n = 40) ± SE. (B) Petiole length of leaf 4 of Ler and uvr8-1 plants grown for 10 days in 16 hr light/8 hr dark cycles at 20°C before transfer to 20°C, 28°C, 20°C + UV-B, or 28°C + UV-B for a further 9 days. Data represent mean length (n ≥ 23) ± SE. Different letters indicate statistically significant means (p < 0.05). Two-way ANOVA confirmed that there was a significant interaction between genotype and condition on petiole length (p < 0.001). (C) Representative rosettes of plants grown as in (B). See also Figure S1.

Figure 2

UV-B Perceived by UVR8 Inhibits PIF4 Activity and Auxin Signaling at High Temperature (A) Hypocotyl lengths of Col-0, pif4-101, and 35S:PIF4-HA seedlings grown in continuous light for 3 days at 20°C, before transfer to 20°C, 28°C, 20°C + UV-B, or 28°C + UV-B for a further 4 days (n ≤ 27; ±SE). Different letters indicate statistically significant means (p < 0.05). (B) Relative transcript abundance of YUC8 and IAA29 in Ler and uvr8-1 seedlings grown for 10 days in 16 hr light/8 hr dark cycles at 20°C, before transfer at dawn to the indicated conditions for 4 hr (n = 3; ±SE; ^∗significant UV-B-mediated decrease in transcript abundance when compared to 20°C, p < 0.05; ^∗∗significant UV-B-mediated decrease in transcript abundance when compared to 28°C, p < 0.05). See also Figure S1.

Figure 3

UV-B Inhibits PIF4 Transcript Accumulation in a UVR8-Dependent Manner and Promotes PIF4 Degradation in a Temperature-Conditional Manner (A) PIF4-HA abundance in 35S:PIF4-HA seedlings grown for 10 days in 16 hr light/8 hr dark cycles at 20°C, harvested before dawn and 2 hr after dawn following transfer to the stated conditions. Col-0 serves as a negative control. Ponceau stain of Rubisco large subunit (rbcL) serves as a loading control. (B) Time course of plants grown and treated as in (A). Relative protein abundance was normalized to Ponceau staining of the Rubisco large subunit, then expressed as a value relative to pre-dawn levels (n = 3; ±SE). Asterisks denote a significant difference between UV-B- and white light (WL)-treated controls at their respective temperatures. (C) PIF4 transcript abundance in Ler and uvr8-1 seedlings grown as in (A) and harvested at 4 hr (^∗significant UV-B-mediated decrease in transcript abundance when compared to 20°C, p < 0.05; ^∗∗significant UV-B-mediated decrease in transcript abundance when compared to 28°C, p < 0.05). (D) Representative blot showing PIF4 abundance in Ler grown as in (A) at the 2 hr time point using anti-PIF4 antibody. Ponceau stain of Rubisco large subunit (rbcL) serves as a loading control. (E and F) Time course of PIF4 transcript abundance. Seedlings were grown for 10 days in 8 hr light/16 hr dark cycles at 20°C. On day 11, plants were transferred to either (E) 20°C or (F) 28°C ± UV-B. UV-B treatment was maintained for the duration of the photoperiod and plants harvested at the times shown. All values are normalized to time 0. The mean of two biological repeats are shown ± SD. See also Figures S2 and S3.

Figure 4

UV-B-Mediated Stabilization of HFR1 Suppresses PIF4 Activity at 28°C (A–C) Hypocotyl lengths of (A) Col-0 and elf3-1; (B) Ws, hy5, hyh, and hy5/hyh; and (C) Col-7 and hfr1 seedlings grown in continuous light for 3 days at 20°C, before transfer to 20°C, 28°C, 20°C + UV-B, or 28°C + UV-B for a further 4 days. Data represent mean values (n = 40) ± SE. Different letters indicate statistically significant means (p < 0.05). Two-way ANOVA confirmed an interaction between genotype^∗condition on hypocotyl length between Col-7 and hfr1 plants (p < 0.001). (D) Representative blot showing HFR1-HA abundance in pHFR1:HFR1-HA seedlings grown for 10 days in 16 hr light/8 hr dark cycles at 20°C, following 2 hr transfer to the stated conditions using an anti-HA antibody. (E) Hypothetical model depicting UV-B-mediated inhibition of hypocotyl elongation at different temperatures. At 20°C, UV-B perceived by UVR8 inhibits PIF4 transcript accumulation in a response requiring COP1. This reduces PIF4 protein abundance. Simultaneously, UV-B drives degradation of PIF4 protein and stabilizes HFR1. At 28°C, UV-B perceived by UVR8 inhibits PIF4 transcript abundance, in a response requiring COP1. This reduces PIF4 protein accumulation. PIF4 is protected from UV-B-induced degradation at elevated temperature, but its transcriptional activity is inhibited by high HFR1 levels. The abundance of HFR1 increases at 28°C. In UV-B, UVR8 sequesters COP1, inhibiting COP1-mediated HFR1 degradation. A role for HYH in the UV-B-mediated inhibition of hypocotyl elongation was additionally observed at high temperature, although no known mechanism exists for HYH regulation of PIF4 activity. Collectively, UV-B inhibits hypocotyl elongation by reducing PIF4 abundance and activity, thereby limiting auxin biosynthesis. The relative contributions of different regulatory mechanisms to this overall response are dependent on ambient temperature. See also Figures S2–S4.