Ambient temperature sensing by phytochrome B (PHYB) in Arabidopsis is thought to operate mainly at night. Here we show that PHYB plays an equally critical role in temperature sensing during the daytime. In daytime thermosensing, PHYB signals primarily through the temperature-responsive transcriptional regulator PIF4, which requires the transcriptional activator HEMERA (HMR). HMR does not regulate PIF4 transcription, instead, it interacts directly with PIF4, to activate the thermoresponsive growth-relevant genes and promote warm-temperature-dependent PIF4 accumulation. A missense allele hmr-22, which carries a loss-of-function D516N mutation in HMR’s transcriptional activation domain, fails to induce the thermoresponsive genes and PIF4 accumulation. Both defects of hmr-22 could be rescued by expressing a HMR22 mutant protein fused with the transcriptional activation domain of VP16, suggesting a causal relationship between HMR-mediated activation of PIF4 target-genes and PIF4 accumulation. Together, this study reveals a daytime PHYB-mediated thermosensing mechanism, in which HMR acts as a necessary activator for PIF4-dependent induction of temperature-responsive genes and PIF4 accumulation.

Phytochromes (phys) are red and far-red photoreceptors that control plant development and growth by promoting the proteolysis of a family of antagonistically acting basic helix-loop-helix transcription factors, the PHYTOCHROME-INTERACTING FACTORs (PIFs). We have previously shown that the degradation of PIF1 and PIF3 requires HEMERA (HMR). However, the biochemical function of HMR and the mechanism by which it mediates PIF degradation remain unclear. Here, we provide genetic evidence that HMR acts upstream of PIFs in regulating hypocotyl growth. Surprisingly, genomewide analysis of HMR-and PIF-dependent genes reveals that HMR is also required for the transactivation of a subset of PIF direct-target genes. We show that HMR interacts with all PIFs. The HMR-PIF interaction is mediated mainly by HMR's N-terminal half and PIFs' conserved active-phytochrome B binding motif. In addition, HMR possesses an acidic nine-aminoacid transcriptional activation domain (9aaTAD) and a loss-of-function mutation in this 9aaTAD impairs the expression of PIF target genes and the destruction of PIF1 and PIF3. Together, these in vivo results support a regulatory mechanism for PIFs in which HMR is a transcriptional coactivator binding directly to PIFs and the 9aaTAD of HMR couples the degradation of PIF1 and PIF3 with the transactivation of PIF target genes.

Plant genomes are extremely sensitive to, and can be developmentally reprogrammed by environmental light cues. Here using rolling-circle amplification of gene-specific circularizable oligonucleotides coupled with fluorescence in situ hybridization, we demonstrate that light triggers a rapid repositioning of the Arabidopsis light-inducible chlorophyll a/b-binding proteins (CAB) locus from the nuclear interior to the nuclear periphery during its transcriptional activation. CAB repositioning is mediated by the red/far-red photoreceptors phytochromes (PHYs) and is inhibited by repressors of PHY signalling, including COP1, DET1 and PIFs. CAB repositioning appears to be a separate regulatory step occurring before its full transcriptional activation. Moreover, the light-inducible loci RBCS, PC and GUN5 undergo similar repositioning behaviour during their transcriptional activation. Our study supports a light-dependent gene regulatory mechanism in which PHYs activate light-inducible loci by relocating them to the nuclear periphery; it also provides evidence for the biological importance of gene positioning in the plant kingdom.

Plant phytochromes are thought to transduce light signals by mediating the degradation of phytochrome-interacting transcription factors (PIFs) through the N-terminal photosensory module, while the C-terminal module, including a histidine kinase-related domain (HKRD), does not participate in signaling. Here we show that the C-terminal module of Arabidopsis phytochrome B (PHYB) is sufficient to mediate the degradation of PIF3 specifically and to activate photosynthetic genes in the dark. The HKRD is a dimerization domain for PHYB homo and heterodimerization. A D1040V mutation, which disrupts the dimerization of HKRD and the interaction between C-terminal module and PIF3, abrogates PHYB nuclear accumulation, photobody biogenesis, and PIF3 degradation. By contrast, disrupting the interaction between PIF3 and PHYB’s N-terminal module has little effect on PIF3 degradation. Together, this study demonstrates that the dimeric form of the C-terminal module plays important signaling roles by targeting PHYB to subnuclear photobodies and interacting with PIF3 to trigger its degradation.

Warm temperature is postulated to induce plant thermomorphogenesis through a signaling mechanism similar to shade, as both destabilize the active form of the photoreceptor and thermosensor phytochrome B (phyB). At the cellular level, shade antagonizes phyB signaling by triggering phyB disassembly from photobodies. Here we report temperature-dependent photobody localization of fluorescent protein-tagged phyB (phyB-FP) in the epidermal cells of Arabidopsis hypocotyl and cotyledon. Our results demonstrate that warm temperature elicits different photobody dynamics than those by shade. Increases in temperature from 12°C to 27°C incrementally reduce photobody number by stimulating phyB-FP disassembly from selective thermo-unstable photobodies. The thermostability of photobodies relies on phyB's photosensory module. Surprisingly, elevated temperatures inflict opposite effects on phyB's functions in the hypocotyl and cotyledon despite inducing similar photobody dynamics, indicative of tissue/organ-specific temperature signaling circuitry either downstream of photobody dynamics or independent of phyB. Our results thus provide direct cell biology evidence supporting an early temperature signaling mechanism via dynamic assembly/disassembly of individual photobodies possessing distinct thermostabilities.