SWAP1-SFPS-RRC1 splicing factor complex modulates pre-mRNA splicing to promote photomorphogenesis in Arabidopsis.

Light signals perceived by a group of photoreceptors have profound effects on the physiology, growth, and development of plants. The red/far-red light-absorbing phytochromes (phys) modulate these aspects by intricately regulating gene expression at multiple levels. Here, we report the identification and functional characterization of an RNA-binding splicing factor, SWAP1 (SUPPRESSOR-OF-WHITE-APRICOT/SURP RNA-BINDING DOMAIN-CONTAINING PROTEIN1). Loss-of-function swap1-1 mutant is hyposensitive to red light and exhibits a day length-independent early flowering phenotype. SWAP1 physically interacts with two other splicing factors, (SFPS) SPLICING FACTOR FOR PHYTOCHROME SIGNALING and (RRC1) REDUCED RED LIGHT RESPONSES IN CRY1CRY2 BACKGROUND 1 in a light-independent manner and forms a ternary complex. In addition, SWAP1 physically interacts with photoactivated phyB and colocalizes with nuclear phyB photobodies. Phenotypic analyses show that the swap1sfps, swap1rrc1, and sfpsrrc1 double mutants display hypocotyl lengths similar to that of the respective single mutants under red light, suggesting that they function in the same genetic pathway. The swap1sfps double and swap1sfpsrrc1 triple mutants display pleiotropic phenotypes, including sterility at the adult stage. Deep RNA sequencing (RNA-seq) analyses show that SWAP1 regulates the gene expression and pre-messenger RNA (mRNA) alternative splicing of a large number of genes, including those involved in plant responses to light signaling. A comparative analysis of alternative splicing among single, double, and triple mutants showed that all three splicing factors coordinately regulate the alternative splicing of a subset of genes. Our study uncovered the function of a splicing factor that modulates light-regulated alternative splicing by interacting with photoactivated phyB and other splicing factors.

Phytochrome B triggers light-dependent chromatin remodelling through the PRC2-associated PHD finger protein VIL1.

To compensate for a sessile nature, plants have developed sophisticated mechanisms to sense varying environmental conditions. Phytochromes (phys) are light and temperature sensors that regulate downstream genes to render plants responsive to environmental stimuli1-4. Here, we show that phyB directly triggers the formation of a repressive chromatin loop by physically interacting with VERNALIZATION INSENSITIVE 3-LIKE1/VERNALIZATION 5 (VIL1/VRN5), a component of Polycomb Repressive Complex 2 (PRC2)5,6, in a light-dependent manner. VIL1 and phyB cooperatively contribute to the repression of growth-promoting genes through the enrichment of Histone H3 Lys27 trimethylation (H3K27me3), a repressive histone modification. In addition, phyB and VIL1 mediate the formation of a chromatin loop to facilitate the repression of ATHB2. Our findings show that phyB directly utilizes chromatin remodelling to regulate the expression of target genes in a light-dependent manner.

Light-regulated pre-mRNA splicing in plants.

Light signal perceived by the red/far-red absorbing phytochrome (phy) family of photoreceptors regulates plant growth and development throughout the life cycle. Phytochromes regulate the light-triggered physiological responses by controlling gene expression both at the transcriptional and post-transcriptional levels. Recent large-scale RNA-seq studies have demonstrated the roles of phys in altering the global transcript diversity by modulating the pre-mRNA splicing in response to light. Moreover, several phy-interacting splicing factors/regulators from different species have been identified using forward genetics and protein-protein interaction studies, which modulate the light-regulated pre-mRNA splicing. In this article, we summarize our current understanding of the role of phys in the light-mediated pre-mRNA splicing and how that contributes to the regulation of gene expression to promote photomorphogenesis.

Phytochrome Signaling Networks.

The perception of light signals by the phytochrome family of photoreceptors has a crucial influence on almost all aspects of growth and development throughout a plant's life cycle. The holistic regulatory networks orchestrated by phytochromes, including conformational switching, subcellular localization, direct protein-protein interactions, transcriptional and posttranscriptional regulations, and translational and posttranslational controls to promote photomorphogenesis, are highly coordinated and regulated at multiple levels. During the past decade, advances using innovative approaches have substantially broadened our understanding of the sophisticated mechanisms underlying the phytochrome-mediated light signaling pathways. This review discusses and summarizes these discoveries of the role of the modular structure of phytochromes, phytochrome-interacting proteins, and their functions; the reciprocal modulation of both positive and negative regulators in phytochrome signaling; the regulatory roles of phytochromes in transcriptional activities, alternative splicing, and translational regulation; and the kinases and E3 ligases that modulate PHYTOCHROME INTERACTING FACTORs to optimize photomorphogenesis.

PCH1 and PCHL Directly Interact with PIF1, Promote Its Degradation, and Inhibit Its Transcriptional Function during Photomorphogenesis.

PHOTOPERIODIC CONTROL OF HYPOCOTYL 1 (PCH1) and PCH1-LIKE (PCHL) were shown to directly bind to phytochrome B (phyB) and suppress phyB thermal reversion, resulting in plants with dramatically enhanced light sensitivity. Here, we show that PCH1 and PCHL also positively regulate various light responses, including seed germination, hypocotyl gravitropism, and chlorophyll biosynthesis, by physically interacting with PHYTOCHROME INTERACTING FACTOR 1 (PIF1) and CONSTITUTIVE PHOTOMORPHOGENIC 1 (COP1). PCH1 and PCHL interact with PIF1 both in the dark and light, and regulate PIF1 abundance. Moreover, PCH1 and PCHL facilitate the physical interaction between phyB and PIF1 in vivo to promote the light-induced degradation of PIF1. PCH1 and PCHL also inhibit the DNA-binding ability of PIF1 to negatively regulate the expressions of PIF1 target genes. In addition, PCH1 and PCHL interact with COP1 and undergo degradation through the 26S proteasome pathway in the dark. Consistently, pch1 suppresses cop1 phenotype in darkness. Collectively, our study reveals a novel mechanism by which PCH1 and PCHL regulate diverse light responses not only by stabilizing phyB Pfr form but also by directly interacting with PIF1 and COP1, providing a molecular understanding of the control of hypocotyl growth by these proteins.

Coordinated Regulation of Pre-mRNA Splicing by the SFPS-RRC1 Complex to Promote Photomorphogenesis.

Light signals perceived by the phytochrome (phy) family of photoreceptors control gene expression at both transcriptional and posttranscriptional levels to promote photomorphogenesis. Recently, we identified a factor called SPLICING FACTOR FOR PHYTOCHROME SIGNALING (SFPS) that directly interacts with the photoreceptor phyB and regulates pre-mRNA splicing in Arabidopsis (Arabidopsis thaliana). To identify SFPS-interacting proteins, we performed an immunoprecipitation followed by a mass spectrometry and identified the Ser/Arg-like protein REDUCED RED-LIGHT RESPONSES IN CRY1CRY2 BACKGROUND1 (RRC1). Genetic analyses revealed that the sfps-2 rrc1-3 phenotypes are similar to those of the single mutants, suggesting that RRC1 and SFPS might function together. RNA sequence analyses of rrc1-3 identified a large number of genes whose pre-mRNA splicing is altered under dark and light conditions. Comparison of the sequence data revealed a subset of common genes coregulated by SFPS and RRC1 under dark and light conditions. Similar to SFPS, RRC1 also interacts with phyB, colocalizes in nuclear photobodies, and regulates light-dependent pre-mRNA splicing of a subset of genes. Taken together, these data suggest that although SFPS and RRC1 can regulate distinct subsets of genes, they also form a complex and coordinately control pre-mRNA splicing of a subset of genes involved in light signaling and circadian clock pathways to promote photomorphogenesis.

SAUR53 regulates organ elongation and apical hook development in Arabidopsis.

SAUR53 is a member of SAUR (Small Auxin-Up RNA) gene family of primary auxin responsive genes. In Arabidopsis, SAUR gene family is represented by 81 genes including two pseudogenes; however, the functions of most of these genes are not fully characterized yet. In the present study, we show that SAUR53 expresses throughout the plant and localizes to both plasma membrane and the nucleus. Unlike most other SAUR genes, expression of SAUR53 is not induced in response to auxin. Ectopic expression of SAUR53 results in the elongation of cells and organs, and also interferes with normal apical hook development by accelerating the hook maintenance phase. Moreover, root growth of SAUR53 overexpression seedlings is significantly insensitive to IAA and 2,4-D, while showing wild-type sensitivity to NAA, suggesting that elevated level of SAUR53 may interfere with normal auxin transport. Collectively, this study indicates that while SAUR53 positively regulates cell and organ elongation, it probably negatively regulates auxin transport in Arabidopsis.

Dynamic regulation of PIF5 by COP1-SPA complex to optimize photomorphogenesis in Arabidopsis.

Light signal provides the spatial and temporal information for plants to adapt to the prevailing environmental conditions. Alterations in light quality and quantity can trigger robust changes in global gene expression. In Arabidopsis thaliana, two groups of key factors regulating those changes in gene expression are CONSTITUTIVE PHOTOMORPHOGENESIS/DEETIOLATED/FUSCA (COP/DET/FUS) and a subset of basic helix-loop-helix transcription factors called PHYTOCHROME-INTERACTING FACTORS (PIFs). Recently, rapid progress has been made in characterizing the E3 ubiquitin ligases for the light-induced degradation of PIF1, PIF3 and PIF4; however, the E3 ligase(s) for PIF5 remains unknown. Here, we show that the CUL4COP1-SPA complex is necessary for the red light-induced degradation of PIF5. Furthermore, COP1 and SPA proteins stabilize PIF5 in the dark, but promote the ubiquitination and degradation of PIF5 in response to red light through the 26S proteasome pathway. Genetic analysis illustrates that overexpression of PIF5 can partially suppress both cop1-4 and spaQ seedling de-etiolation phenotypes under dark and red-light conditions. In addition, the PIF5 protein level cycles under both diurnal and constant light conditions, which is also defective in the cop1-4 and spaQ backgrounds. Both cop1-4 and spaQ show defects in diurnal growth pattern. Overexpression of PIF5 partially restores growth defects in cop1-4 and spaQ under diurnal conditions, suggesting that the COP1-SPA complex plays an essential role in photoperiodic hypocotyl growth, partly through regulating the PIF5 level. Taken together, our data illustrate how the CUL4COP1-SPA E3 ligase dynamically controls the PIF5 level to regulate plant development.

KELCH F-BOX protein positively influences Arabidopsis seed germination by targeting PHYTOCHROME-INTERACTING FACTOR1.

Seeds employ sensory systems that assess various environmental cues over time to maximize the successful transition from embryo to seedling. Here we show that the Arabidopsis F-BOX protein COLD TEMPERATURE-GERMINATING (CTG)-10, identified by activation tagging, is a positive regulator of this process. When overexpressed (OE), CTG10 hastens aspects of seed germination. CTG10 is expressed predominantly in the hypocotyl, and the protein is localized to the nucleus. CTG10 interacts with PHYTOCHROME-INTERACTING FACTOR 1 (PIF1) and helps regulate its abundance in plantaCTG10-OE accelerates the loss of PIF1 in light, increasing germination efficiency, while PIF1-OE lines fail to complete germination in darkness, which is reversed by concurrent CTG10-OE Double-mutant (pif1 ctg10) lines demonstrated that PIF1 is epistatic to CTG10. Both CTG10 and PIF1 amounts decline during seed germination in the light but reaccumulate in the dark. PIF1 in turn down-regulates CTG10 transcription, suggesting a feedback loop of CTG10/PIF1 control. The genetic, physiological, and biochemical evidence, when taken together, leads us to propose that PIF1 and CTG10 coexist, and even accumulate, in the nucleus in darkness, but that, following illumination, CTG10 assists in reducing PIF1 amounts, thus promoting the completion of seed germination and subsequent seedling development.

Phytochromes and Phytochrome Interacting Factors.

The basic helix-loop-helix domain-containing transcription factors that interact physically with the red and far-red light photoreceptors, phytochromes, are called PHYTOCHROME INTERACTING FACTORS (PIFs). In the last two decades, the phytochrome-PIF signaling module has been shown to be conserved from Physcomitrella patens to higher plants. Exciting recent studies highlight the discovery of at least four distinct kinases (PPKs, CK2, BIN2, and phytochrome itself) and four families of ubiquitin ligases (SCFEBF1/2, CUL3LRB, CUL3BOP, and CUL4COP1-SPA) that regulate PIF abundance both in dark and light conditions. This review discusses these recent discoveries with a focus on the central phytochrome signaling mechanisms that have a profound impact on plant growth and development in response to light.

PCH1 and PCHL promote photomorphogenesis in plants by controlling phytochrome B dark reversion.

Phytochrome B (phyB) is the primary red light photoreceptor in plants, and regulates both growth and development. The relative levels of phyB in the active state are determined by the light conditions, such as direct sunlight or shade, but are also affected by light-independent dark reversion. Dark reversion is a temperature-dependent thermal relaxation process, by which phyB reverts from the active to the inactive state. Here, we show that the homologous phyB-binding proteins PCH1 and PCHL suppress phyB dark reversion, resulting in plants with dramatically enhanced light sensitivity. Moreover, far-red and blue light upregulate the expression of PCH1 and PCHL in a phyB independent manner, thereby increasing the response to red light perceived by phyB. PCH1 and PCHL therefore provide a node for the molecular integration of different light qualities by regulation of phyB dark reversion, allowing plants to adapt growth and development to the ambient environment.

Expanding Roles of PIFs in Signal Integration from Multiple Processes.

PHYTOCHROME-INTERACTING FACTORs (PIFs) are members of the basic helix-loop-helix (bHLH) family of transcription factors in Arabidopsis. Since their discovery in phytochrome-mediated light signaling pathways, recent studies have unraveled new functions of PIFs in integrating multiple signaling pathways not only through their role as transcription factors directly targeting gene expression but also by interacting with diverse groups of factors to optimize plant growth and development. These include endogenous (e.g., hormonal) as well as abiotic (light, circadian, and elevated temperature) and biotic (defense responses) pathways. PIFs interact with key factors in each of these pathways and tailor the outcome of the signal integration among these pathways. This review discusses the roles of PIFs as pivotal signal integrators in regulating plant growth and development.

Reciprocal proteasome-mediated degradation of PIFs and HFR1 underlies photomorphogenic development in Arabidopsis.

The phytochrome-mediated regulation of photomorphogenesis under red and far-red light conditions involves both positively and negatively acting factors. The positively acting factors (e.g. HY5/HFR1/LAF1 and others) are degraded in the dark to prevent photomorphogenesis. By contrast, the negatively acting factors (e.g. phytochrome-interacting factors or PIFs) are degraded in response to light to promote photomorphogenesis. Here, we show that the negatively acting factor PIF1 is also degraded in the dark by direct heterodimerization with the positively acting factor HFR1. Conversely, PIF1 also promotes the degradation of HFR1 in darkness. PIF1 enhances the poly-ubiquitylation of HFR1 by COP1 in vivo and in vitro In addition, the reciprocal co-degradation of PIF1 and HFR1 is dependent on the 26S proteasome pathway in vivo Genetic evidence shows that the hfr1 mutant partially suppresses the constitutive photomorphogenic phenotypes of cop1-6 pif1 and of the quadruple mutant pifq both in the dark and in far-red light conditions. Taken together, these data uncover a co-degradation mechanism between PIFs and HFR1 that underlies photomorphogenic development in Arabidopsis thaliana.