PHYTOCHROME-INTERACTING FACTOR 7 and RELATIVE OF EARLY FLOWERING 6 act in shade avoidance memory in Arabidopsis.

Shade avoidance helps plants maximize their access to light for growth under crowding. It is unknown, however, whether a priming shade avoidance mechanism exists that allows plants to respond more effectively to successive shade conditions. Here, we show that the shade-intolerant plant Arabidopsis can remember a first experienced shade event and respond more efficiently to the next event on hypocotyl elongation. The transcriptional regulator PHYTOCHROME-INTERACTING FACTOR 7 (PIF7) and the histone H3K27-demethylase RELATIVE OF EARLY FLOWERING 6 (REF6) are identified as being required for this shade avoidance memory. RNA-sequencing analysis reveals that shade induction of shade-memory-related genes is impaired in the pif7 and ref6 mutants. Based on the analyses of enrichments of H3K27me3, REF6 and PIF7, we find that priming shade treatment induces PIF7 accumulation, which further recruits REF6 to demethylate H3K27me3 on the chromatin of certain shade-memory-related genes, leading to a state poised for their transcription. Upon a second shade treatment, enhanced shade-mediated inductions of these genes result in stronger hypocotyl growth responses. We conclude that the transcriptional memory mediated by epigenetic modification plays a key role in the ability of primed plants to remember previously experienced shade and acquire enhanced responses to recurring shade conditions.

lncRNAs involved in the Shade Avoidance Syndrome (SAS) in Arabidopsis thaliana.

BACKGROUND: Plant long non-coding RNAs (lncRNAs) have important regulatory roles in responses to various biotic and abiotic stresses, including light quality. However, no lncRNAs have been specifically linked to the Shade Avoidance Response (SAS). RESULTS: To better understand the involvement of lncRNAs in shade avoidance, we examined RNA-seq libraries for lncRNAs with the potential to function in the neighbor proximity phenomenon in Arabidopsis thaliana (A. thaliana). Using transcriptomes generated from seedlings exposed to high and low red/far-red (R/FR) light conditions, we identified 13 lncRNA genes differentially expressed in cotyledons and 138 in hypocotyls. To infer possible functions for these lncRNAs, we used a 'guilt-by-association' approach to identify genes co-expressed with lncRNAs in a weighted gene co-expression network. Of 34 co-expression modules, 10 showed biological functions related to differential growth. We identified three potential lncRNAs co-regulated with genes related to SAS. T-DNA insertions in two of these lncRNAs were correlated with morphological differences in seedling responses to increased FR light, supporting our strategy for computational identification of lncRNAs involved in SAS. CONCLUSIONS: Using a computational approach, we identified multiple lncRNAs in Arabidopsis involved in SAS. T-DNA insertions caused altered phenotypes under low R/FR light, suggesting functional roles in shade avoidance. Further experiments are needed to determine the specific mechanisms of these lncRNAs in SAS.

Ubiquitin-specific protease UBP14 stabilizes HY5 by deubiquitination to promote photomorphogenesis in Arabidopsis thaliana.

Transcription factor ELONGATED HYPOCOTYL5 (HY5) is the central hub for seedling photomorphogenesis. E3 ubiquitin (Ub) ligase CONSTITUTIVE PHOTOMORPHOGENIC 1 (COP1) inhibits HY5 protein accumulation through ubiquitination. However, the process of HY5 deubiquitination, which antagonizes E3 ligase-mediated ubiquitination to maintain HY5 homeostasis has never been studied. Here, we identified that Arabidopsis thaliana deubiquitinating enzyme, Ub-SPECIFIC PROTEASE 14 (UBP14) physically interacts with HY5 and enhances its protein stability by deubiquitination. The da3-1 mutant lacking UBP14 function exhibited a long hypocotyl phenotype, and UBP14 deficiency led to the failure of rapid accumulation of HY5 during dark to light. In addition, UBP14 preferred to stabilize nonphosphorylated form of HY5 which is more readily bound to downstream target genes. HY5 promoted the expression and protein accumulation of UBP14 for positive feedback to facilitate photomorphogenesis. Our findings thus established a mechanism by which UBP14 stabilizes HY5 protein by deubiquitination to promote photomorphogenesis in A. thaliana.

SlCPK27 cross-links SlHY5 and SlPIF4 in brassinosteroid-dependent photo- and thermo-morphogenesis in tomato.

ELONGATED HYPOCOTOYL5 (HY5) and PHYTOCHROME INTERACTING FACTORs (PIFs) are two types of important light-related regulators of plant growth, however, their interplay remains elusive. Here, we report that the activated tomato (Solanum lycopersicum) HY5 (SlHY5) triggers the transcription of a Calcium-dependent Protein Kinase SlCPK27. SlCPK27 interacts with and phosphorylates SlPIF4 at Ser-252 and Ser-308 phosphosites to promote its degradation. SlPIF4 promotes hypocotyl elongation mainly by activating the transcription of SlDWF, a key gene in brassinosteroid (BR) biosynthesis. Such a SlHY5-SlCPK27-SlPIF4-BR cascade not only plays a crucial role in photomorphogenesis but also regulates thermomorphogenesis. Our results uncover a previously unidentified mechanism that integrates Ca2+ signaling with the light signaling pathways to regulate plant growth by modulating BR biosynthesis in response to changes in ambient light and temperature.

Arabidopsis thaliana MYC2 and MYC3 Are Involved in Ethylene-Regulated Hypocotyl Growth as Negative Regulators.

The ethylene-regulated hypocotyl elongation of Arabidopsis thaliana involves many transcription factors. The specific role of MYC transcription factors in ethylene signal transduction is not completely understood. The results here revealed that two MYCs, MYC2 and MYC3, act as negative regulators in ethylene-suppressed hypocotyl elongation. Etiolated seedlings of the loss-of-function mutant of MYC2 or MYC3 were significantly longer than wild-type seedlings. Single- or double-null mutants of MYC2 and MYC3 displayed remarkably enhanced response to ACC(1-aminocyclopropane-1-carboxylate), the ethylene precursor, compared to wild-type seedlings. MYC2 and MYC3 directly bind to the promoter zone of ERF1, strongly suppressing its expression. Additionally, EIN3, a key component in ethylene signaling, interacts with MYC2 or MYC3 and significantly suppresses their binding to ERF1's promoter. MYC2 and MYC3 play crucial roles in the ethylene-regulated expression of functional genes. The results revealed the novel role and functional mechanism of these transcription factors in ethylene signal transduction. The findings provide valuable information for deepening our understanding of their role in regulating plant growth and responding to stress.

Atmospheric nitrogen dioxide suppresses the activity of phytochrome interacting factor 4 to suppress hypocotyl elongation.

MAIN CONCLUSION: Ambient concentrations of atmospheric nitrogen dioxide (NO2) inhibit the binding of PIF4 to promoter regions of auxin pathway genes to suppress hypocotyl elongation in Arabidopsis. Ambient concentrations (10-50 ppb) of atmospheric nitrogen dioxide (NO2) positively regulate plant growth to the extent that organ size and shoot biomass can nearly double in various species, including Arabidopsis thaliana (Arabidopsis). However, the precise molecular mechanism underlying NO2-mediated processes in plants, and the involvement of specific molecules in these processes, remain unknown. We measured hypocotyl elongation and the transcript levels of PIF4, encoding a bHLH transcription factor, and its target genes in wild-type (WT) and various pif mutants grown in the presence or absence of 50 ppb NO2. Chromatin immunoprecipitation assays were performed to quantify binding of PIF4 to the promoter regions of its target genes. NO2 suppressed hypocotyl elongation in WT plants, but not in the pifq or pif4 mutants. NO2 suppressed the expression of target genes of PIF4, but did not affect the transcript level of the PIF4 gene itself or the level of PIF4 protein. NO2 inhibited the binding of PIF4 to the promoter regions of two of its target genes, SAUR46 and SAUR67. In conclusion, NO2 inhibits the binding of PIF4 to the promoter regions of genes involved in the auxin pathway to suppress hypocotyl elongation in Arabidopsis. Consequently, PIF4 emerges as a pivotal participant in this regulatory process. This study has further clarified the intricate regulatory mechanisms governing plant responses to environmental pollutants, thereby advancing our understanding of how plants adapt to changing atmospheric conditions.

NHX5/NHX6/SPY22 complex regulates BRI1 and brassinosteroid signaling in Arabidopsis.

NHX5 and NHX6, Arabidopsis endosomal antiporters, play a vital role in facilitating ion and pH homeostasis in endosomal compartments. Studies have found that NHX5 and NHX6 are essential for protein trafficking, auxin homeostasis, and plant growth and development. Here, we report the role of NHX5 and NHX6 in brassinosteroid (BR) signaling. We found that hypocotyl growth was enhanced in nhx5 nhx6 under epibrassinolide (eBR) treatment. nhx5 nhx6 bri1 was insensitive to eBR treatment, indicating that NHX5 and NHX6 are downstream of the BRI1 receptor in BR signaling. Moreover, confocal observation with both hypocotyls and root tips showed that BRI1-YFP localization in the plasma membrane (PM) was reduced in nhx5 nhx6. Interestingly, brefeldin A (BFA) treatment showed that formation of the BFA bodies containing BRI1 and their disassembling were disrupted in nhx5 nhx6. Further genetic analysis showed that NHX5/NHX6 and SYP22 may act coordinately in BR signaling. NHX5 and NHX6 may regulate SYP22 function by modulating cellular K+ and pH homeostasis. Importantly, NHX5 and NHX6 colocalize and interact with SYP22, but do not interact with BRI1. In summary, our findings indicate that NHX5/NHX6/SYP22 complex is essential for the regulation of BRI1 recycling and PM localization. The H+-leak facilitated by NHX5 and NHX6 offers a means of controlling BR signaling in plants.

A Novel 10-Base Pair Deletion in the First Exon of GmHY2a Promotes Hypocotyl Elongation, Induces Early Maturation, and Impairs Photosynthetic Performance in Soybean.

Plants photoreceptors perceive changes in light quality and intensity and thereby regulate plant vegetative growth and reproductive development. By screening a γ irradiation-induced mutant library of the soybean (Glycine max) cultivar "Dongsheng 7", we identified Gmeny, a mutant with elongated nodes, yellowed leaves, decreased chlorophyll contents, altered photosynthetic performance, and early maturation. An analysis of bulked DNA and RNA data sampled from a population segregating for Gmeny, using the BVF-IGV pipeline established in our laboratory, identified a 10 bp deletion in the first exon of the candidate gene Glyma.02G304700. The causative mutation was verified by a variation analysis of over 500 genes in the candidate gene region and an association analysis, performed using two populations segregating for Gmeny. Glyma.02G304700 (GmHY2a) is a homolog of AtHY2a in Arabidopsis thaliana, which encodes a PΦB synthase involved in the biosynthesis of phytochrome. A transcriptome analysis of Gmeny using the Kyoto Encyclopedia of Genes and Genomes (KEGG) revealed changes in multiple functional pathways, including photosynthesis, gibberellic acid (GA) signaling, and flowering time, which may explain the observed mutant phenotypes. Further studies on the function of GmHY2a and its homologs will help us to understand its profound regulatory effects on photosynthesis, photomorphogenesis, and flowering time.

A cytosol-tethered YHB variant of phytochrome B retains photomorphogenic signaling activity.

The red and far-red light photoreceptor phytochrome B (phyB) transmits light signals following cytosol-to-nuclear translocation to regulate transcriptional networks therein. This necessitates changes in protein-protein interactions of phyB in the cytosol, about which little is presently known. Via introduction of a nucleus-excluding G767R mutation into the dominant, constitutively active phyBY276H (YHB) allele, we explore the functional consequences of expressing a cytosol-localized YHBG767R variant in transgenic Arabidopsis seedlings. We show that YHBG767R elicits selective constitutive photomorphogenic phenotypes in dark-grown phyABCDE null mutants, wild type and other phy-deficient genotypes. These responses include light-independent apical hook opening, cotyledon unfolding, seed germination and agravitropic hypocotyl growth with minimal suppression of hypocotyl elongation. Such phenotypes correlate with reduced PIF3 levels, which implicates cytosolic targeting of PIF3 turnover or PIF3 translational inhibition by YHBG767R. However, as expected for a cytoplasm-tethered phyB, YHBG767R elicits reduced light-mediated signaling activity compared with similarly expressed wild-type phyB in phyABCDE mutant backgrounds. YHBG767R also interferes with wild-type phyB light signaling, presumably by formation of cytosol-retained and/or otherwise inactivated heterodimers. Our results suggest that cytosolic interactions with PIFs play an important role in phyB signaling even under physiological conditions.

Establishment of an efficient regeneration system of 'ZiKui' tea with hypocotyl as explants.

Zikui (Camellia sinensis cv. Zikui) is a recently discovered cultivar of local purple tea in Guizhou, China. It is a purple leaf bud mutation material of Meitan Taicha (Camellia sinensis cv. 'Meitan-taicha') 'N61' strain, which is an important local germplasm resource in Guizhou. It is also a model plant for the study of anthocyanins, but the limited germplasm resources and the limitation of traditional reproduction hinder its application. Here, an efficient regeneration system is established by using hypocotyl as explants for the first time. Different plant growth regulators (PGRs) are evaluated during different regeneration processes including callus and root induction. According to our findings, using the optimal disinfection conditions, the seed embryo contamination rate is 17.58%. Additionally, the mortality rate is 9.69%, while the survival rate is measured as 72.73%. Moreover, the highest germination rate of 93.64% is observed under MS + 2.40 mg/L GA3 medium conditions. The optimal callus induction rate is 95.19%, while the optimal adventitious bud differentiation rate is 20.74%, Medium with 1.6 mg/L IBA achieved 68.6% rooting of the adventitious shoots. The survival rate is more than 65% after 6 days growth in the cultivated matrix. In summary, our research aims to establish a regeneration system for Zikui tea plants and design a transformation system for tea plant tissue seedlings. This will enable transfer of the target gene and ultimately facilitate the cultivation of new tea varieties with unique characteristics.

ADA2b acts to positively regulate blue light-mediated photomorphogenesis in Arabidopsis.

Cryptochromes (CRYs) act as blue light photoreceptors to regulate various plant physiological processes including photomorphogenesis and repair of DNA double strand breaks (DSBs). ADA2b is a conserved transcription co-activator that is involved in multiple plant developmental processes. It is known that ADA2b interacts with CRYs to mediate blue light-promoted DSBs repair. Whether ADA2b may participate in CRYs-mediated photomorphogenesis is unknown. Here we show that ADA2b acts to inhibit hypocotyl elongation and hypocotyl cell elongation in blue light. We found that the SWIRM domain-containing C-terminus mediates the blue light-dependent interaction of ADA2b with CRYs in blue light. Moreover, ADA2b and CRYs act to co-regulate the expression of hypocotyl elongation-related genes in blue light. Based on previous studies and these results, we propose that ADA2b plays dual functions in blue light-mediated DNA damage repair and photomorphogenesis.

Non-transcriptional regulatory activity of SMAX1 and SMXL2 mediates karrikin-regulated seedling response to red light in Arabidopsis.

Karrikins and strigolactones govern plant development and environmental responses through closely related signaling pathways. The transcriptional repressor proteins SUPPRESSOR OF MAX2 1 (SMAX1), SMAX1-like2 (SMXL2), and D53-like SMXLs mediate karrikin and strigolactone signaling by directly binding downstream genes or by inhibiting the activities of transcription factors. In this study, we characterized the non-transcriptional regulatory activities of SMXL proteins in Arabidopsis. We discovered that SMAX1 and SMXL2 with mutations in their ethylene-response factor-associated amphiphilic repression (EAR) motif had undetectable or weak transcriptional repression activities but still partially rescued the hypocotyl elongation defects and fully reversed the cotyledon epinasty defects of the smax1 smxl2 mutant. SMAX1 and SMXL2 directly interact with PHYTOCHROME INTERACTION FACTOR 4 (PIF4) and PIF5 to enhance their protein stability by interacting with phytochrome B (phyB) and suppressing the association of phyB with PIF4 and PIF5. The karrikin-responsive genes were then identified by treatment with GR24ent-5DS, a GR24 analog showing karrikin activity. Interestingly, INDOLE-3-ACETIC ACID INDUCIBLE 29 (IAA29) expression was repressed by GR24ent-5DS treatment in a PIF4- and PIF5-dependent and EAR-independent manner, whereas KARRIKIN UPREGULATED F-BOX 1 (KUF1) expression was induced in a PIF4- and PIF5-independent and EAR-dependent manner. Furthermore, the non-transcriptional regulatory activity of SMAX1, which is independent of the EAR motif, had a global effect on gene expression. Taken together, these results indicate that non-transcriptional regulatory activities of SMAX1 and SMXL2 mediate karrikin-regulated seedling response to red light.

miR394 modulates brassinosteroid signaling to regulate hypocotyl elongation in Arabidopsis.

The interplay between microRNAs (miRNAs) and phytohormones allows plants to integrate multiple internal and external signals to optimize their survival of different environmental conditions. Here, we report that miR394 and its target gene LEAF CURLING RESPONSIVENESS (LCR), which are transcriptionally responsive to BR, participate in BR signaling to regulate hypocotyl elongation in Arabidopsis thaliana. Phenotypic analysis of various transgenic and mutant lines revealed that miR394 negatively regulates BR signaling during hypocotyl elongation, whereas LCR positively regulates this process. Genetically, miR394 functions upstream of BRASSINOSTEROID INSENSITIVE2 (BIN2), BRASSINAZOLEs RESISTANT1 (BZR1), and BRI1-EMS-SUPPRESSOR1 (BES1), but interacts with BRASSINOSTEROID INSENSITIVE1 (BRI1) and BRI1 SUPRESSOR PROTEIN (BSU1). RNA-sequencing analysis suggested that miR394 inhibits BR signaling through BIN2, as miR394 regulates a significant number of genes in common with BIN2. Additionally, miR394 increases the accumulation of BIN2 but decreases the accumulation of BZR1 and BES1, which are phosphorylated by BIN2. MiR394 also represses the transcription of PACLOBUTRAZOL RESISTANCE1/5/6 and EXPANSIN8, key genes that regulate hypocotyl elongation and are targets of BZR1/BES1. These findings reveal a new role for a miRNA in BR signaling in Arabidopsis.

Light control of three-dimensional chromatin organization in soybean.

Higher-order chromatin structure is critical for regulation of gene expression. In plants, light profoundly affects the morphogenesis of emerging seedlings as well as global gene expression to ensure optimal adaptation to environmental conditions. However, the changes and functional significance of chromatin organization in response to light during seedling development are not well documented. We constructed Hi-C contact maps for the cotyledon, apical hook and hypocotyl of soybean subjected to dark and light conditions. The resulting high-resolution Hi-C contact maps identified chromosome territories, A/B compartments, A/B sub-compartments, TADs (Topologically Associated Domains) and chromatin loops in each organ. We observed increased chromatin compaction under light and we found that domains that switched from B sub-compartments in darkness to A sub-compartments under light contained genes that were activated during photomorphogenesis. At the local scale, we identified a group of TADs constructed by gene clusters consisting of different numbers of Small Auxin-Upregulated RNAs (SAURs), which exhibited strict co-expression in the hook and hypocotyl in response to light stimulation. In the hypocotyl, RNA polymerase II (RNAPII) regulated the transcription of a SAURs cluster under light via TAD condensation. Our results suggest that the 3D genome is involved in the regulation of light-related gene expression in a tissue-specific manner.

NAC transcription factor ATAF1 negatively modulates the PIF-regulated hypocotyl elongation under a short-day photoperiod.

Day length modulates hypocotyl elongation in seedlings to optimize their overall fitness. Variations in cell growth-associated genes are regulated by several transcription factors. However, the specific transcription factors through which the plant clock increases plant fitness are still being elucidated. In this study, we identified the no apical meristem, Arabidopsis thaliana-activating factor (ATAF-1/2), and cup-shaped cotyledon (NAC) family transcription factor ATAF1 as a novel repressor of hypocotyl elongation under a short-day (SD) photoperiod. Variations in day length profoundly affected the transcriptional and protein levels of ATAF1. ATAF1-deficient mutant exhibited increased hypocotyl length and cell growth-promoting gene expression under SD conditions. Moreover, ATAF1 directly targeted and repressed the expression of the cycling Dof factor 1/5 (CDF1/5), two key transcription factors involved in hypocotyl elongation under SD conditions. Additionally, ATAF1 interacted with and negatively modulated the effects of phytochrome-interacting factor (PIF), thus inhibiting PIF-promoted gene expression and hypocotyl elongation. Taken together, our results revealed ATAF1-PIF as a crucial pair modulating the expression of key transcription factors to facilitate plant growth during day/night cycles under fluctuating light conditions.

The PRR-EC complex and SWR1 chromatin remodeling complex function cooperatively to repress nighttime hypocotyl elongation by modulating PIF4 expression in Arabidopsis.

The circadian clock entrained by environmental light-dark cycles enables plants to fine-tune diurnal growth and developmental responses. Here, we show that physical interactions among evening clock components, including PSEUDO-RESPONSE REGULATOR 5 (PRR5), TIMING OF CAB EXPRESSION 1 (TOC1), and the Evening Complex (EC) component EARLY FLOWERING 3 (ELF3), define a diurnal repressive chromatin structure specifically at the PHYTOCHROME-INTERACTING FACTOR 4 (PIF4) locus in Arabidopsis. These three clock components act interdependently as well as independently to repress nighttime hypocotyl elongation, as hypocotyl elongation rate dramatically increased specifically at nighttime in the prr5-1 toc1-21 elf3-1 mutant, concomitantly with a substantial increase in PIF4 expression. Transcriptional repression of PIF4 by ELF3, PRR5, and TOC1 is mediated by the SWI2/SNF2-RELATED (SWR1) chromatin remodeling complex, which incorporates histone H2A.Z at the PIF4 locus, facilitating robust epigenetic suppression of PIF4 during the evening. Overall, these findings demonstrate that the PRR-EC-SWR1 complex represses hypocotyl elongation at night through a distinctive chromatin domain covering PIF4 chromatin.

RACK1A promotes hypocotyl elongation by scaffolding light signaling components in Arabidopsis.

Plants deploy versatile scaffold proteins to intricately modulate complex cell signaling. Among these, RACK1A (Receptors for Activated C Kinase 1A) stands out as a multifaceted scaffold protein functioning as a central integrative hub for diverse signaling pathways. However, the precise mechanisms by which RACK1A orchestrates signal transduction to optimize seedling development remain largely unclear. Here, we demonstrate that RACK1A facilitates hypocotyl elongation by functioning as a flexible platform that connects multiple key components of light signaling pathways. RACK1A interacts with PHYTOCHROME INTERACTING FACTOR (PIF)3, enhances PIF3 binding to the promoter of BBX11 and down-regulates its transcription. Furthermore, RACK1A associates with ELONGATED HYPOCOTYL 5 (HY5) to repress HY5 biochemical activity toward target genes, ultimately contributing to hypocotyl elongation. In darkness, RACK1A is targeted by CONSTITUTIVELY PHOTOMORPHOGENIC (COP)1 upon phosphorylation and subjected to COP1-mediated degradation via the 26 S proteasome system. Our findings provide new insights into how plants utilize scaffold proteins to regulate hypocotyl elongation, ensuring proper skoto- and photo-morphogenic development.

B-Box transcription factor BBX28 requires CONSTITUTIVE PHOTOMORPHOGENESIS1 to induce shade-avoidance response in Arabidopsis thaliana.

Shade avoidance syndrome is an important adaptive strategy. Under shade, major transcriptional rearrangements underlie the reallocation of resources to elongate vegetative structures and redefine the plant architecture to compete for photosynthesis. BBX28 is a B-box transcription factor involved in seedling de-etiolation and flowering in Arabidopsis (Arabidopsis thaliana), but its function in shade-avoidance response is completely unknown. Here, we studied the function of BBX28 using two mutant and two transgenic lines of Arabidopsis exposed to white light and simulated shade conditions. We found that BBX28 promotes hypocotyl growth under shade through the phytochrome system by perceiving the reduction of red photons but not the reduction of photosynthetically active radiation or blue photons. We demonstrated that hypocotyl growth under shade is sustained by the protein accumulation of BBX28 in the nuclei in a CONSTITUTIVE PHOTOMORPHOGENESIS1 (COP1)-dependent manner at the end of the photoperiod. BBX28 up-regulates the expression of transcription factor- and auxin-related genes, thereby promoting hypocotyl growth under prolonged shade. Overall, our results suggest the role of BBX28 in COP1 signaling to sustain the shade-avoidance response and extend the well-known participation of other members of BBX transcription factors for fine-tuning plant growth under shade.

The phytochrome-interacting factor genes PIF1 and PIF4 are functionally diversified due to divergence of promoters and proteins.

Phytochrome-interacting factors (PIFs) are basic helix-loop-helix transcription factors that regulate light responses downstream of phytochromes. In Arabidopsis (Arabidopsis thaliana), 8 PIFs (PIF1-8) regulate light responses, either redundantly or distinctively. Distinctive roles of PIFs may be attributed to differences in mRNA expression patterns governed by promoters or variations in molecular activities of proteins. However, elements responsible for the functional diversification of PIFs have yet to be determined. Here, we investigated the role of promoters and proteins in the functional diversification of PIF1 and PIF4 by analyzing transgenic lines expressing promoter-swapped PIF1 and PIF4, as well as chimeric PIF1 and PIF4 proteins. For seed germination, PIF1 promoter played a major role, conferring dominance to PIF1 gene with a minor contribution from PIF1 protein. Conversely, for hypocotyl elongation under red light, PIF4 protein was the major element conferring dominance to PIF4 gene with the minor contribution from PIF4 promoter. In contrast, both PIF4 promoter and PIF4 protein were required for the dominant role of PIF4 in promoting hypocotyl elongation at high ambient temperatures. Together, our results support that the functional diversification of PIF1 and PIF4 genes resulted from contributions of both promoters and proteins, with their relative importance varying depending on specific light responses.

A nanopore-based cucumber genome assembly reveals structural variations at two QTLs controlling hypocotyl elongation.

The Xishuangbanna (XIS) cucumber (Cucumis sativus var. xishuangbannanesis) is a semiwild variety that has many distinct agronomic traits. Here, long reads generated by Nanopore sequencing technology helped assembling a high-quality genome (contig N50 = 8.7 Mb) of landrace XIS49. A total of 10,036 structural/sequence variations (SVs) were identified when comparing with Chinese Long (CL), and known SVs controlling spines, tubercles, and carpel number were confirmed in XIS49 genome. Two QTLs of hypocotyl elongation under low light, SH3.1 and SH6.1, were fine-mapped using introgression lines (donor parent, XIS49; recurrent parent, CL). SH3.1 encodes a red-light receptor Phytochrome B (PhyB, CsaV3_3G015190). A ∼4 kb region with large deletion and highly divergent regions (HDRs) were identified in the promoter of the PhyB gene in XIS49. Loss of function of this PhyB caused a super-long hypocotyl phenotype. SH6.1 encodes a CCCH-type zinc finger protein FRIGIDA-ESSENTIAL LIKE (FEL, CsaV3_6G050300). FEL negatively regulated hypocotyl elongation but it was transcriptionally suppressed by long terminal repeats retrotransposon insertion in CL cucumber. Mechanistically, FEL physically binds to the promoter of CONSTITUTIVE PHOTOMORPHOGENIC 1a (COP1a), regulating the expression of COP1a and the downstream hypocotyl elongation. These above results demonstrate the genetic mechanism of cucumber hypocotyl elongation under low light.