BBX21 (also known as SALT TOLERANCE HOMOLOG 2), a B-box (BBX)-containing protein, has been previously identified as a positive regulator of light signaling; however, the precise role of BBX21 in regulating seedling photomorphogenesis remains largely unclear. In this study, we report that CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1) interacts with BBX21 in vivo and is able to ubiquitinate BBX21 in vitro. Thus, BBX21 is targeted for 26S proteasome-mediated degradation in dark-grown Arabidopsis seedlings in a COP1-dependent manner. Moreover, we show that BBX21 binds to the T/G-box in the ELONGATED HYPOCOTYL 5 (HY5) promoter and directly activates HY5 expression in the light. Transgenic seedlings overexpressing BBX21 exhibit dramatically shortened hypocotyls in the light, and this phenotype is dependent on a functional HY5. Taken together, our data suggest a molecular base underlying BBX21-mediated seedling photomorphogenesis, indicating that BBX21 is a pivotal component involved in the COP1-HY5 regulatory hub.P lant seedlings undergo two distinct developmental processes dependent on the presence and absence of light, termed photomorphogenesis and skotomorphogenesis, respectively (1). CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1) is a central repressor of seedling photomorphogenesis, and mutants impaired in COP1 display constitutively photomorphogenic phenotypes in darkness (2, 3). COP1 acts as an E3 ubiquitin ligase targeting a subset of substrates for degradation in darkness, including ELONGATED HYPOCOTYL 5 (HY5), HY5 HOMOLOG (HYH), LONG AFTER FAR-RED LIGHT 1 (LAF1), LONG HYPOCOTYL IN FAR RED (HFR1), B-BOX PROTEIN 22/ SALT TOLERANCE HOMOLOG 3 (BBX22/STH3), and PHYTOCHROME INTERACTING FACTOR 3-LIKE1 (PIL1) (4-11). Of these, HY5 is considered a key signal integration point from dark to light transition (10, 12, 13). Its abundance is directly correlated with the extent of seedling photomorphogenic growth, but inversely correlated with the nuclear abundance of COP1 (4, 10, 14). As a b-ZIP type transcription factor, HY5 specifically interacts with the ACGT-containing elements (ACEs) and might directly bind to approximately one-third of the Arabidopsis genes (12, 13). Thus, HY5 ensures proper expression of a large number of downstream regulatory genes, which in turn eventually promotes photomorphogenesis in the light. Recent studies have shown that HY5 expression is regulated by HY5 itself, as well as by HYH and CALMODULIN7 (CAM7) (15,16).A total of 32 B-box (BBX)-containing proteins have been identified in the Arabidopsis genome (17), among which a number of BBX proteins have been shown to be involved in COP1-and HY5-mediated seedling photomorphogenesis (18). BBX4, BBX20, BBX22, BBX24, and BBX25 genetically and physically interact with COP1 and undergo COP1-mediated degradation in the dark (6,(19)(20)(21)(22)(23)(24). In addition, BBX22, BBX24, and BBX25 physically interact with HY5, and, interestingly, BBX22 acts as a coactivator of HY5 action (6), whereas BBX24 and BBX25 repress the transcriptional activating activity of HY5 ...

BackgroundHigh invasion and metastasis are the primary factors causing poor prognosis of patients with hepatocellular carcinoma (HCC). However, the molecular mechanisms underlying these biological behaviors have not been completely elucidated. In this study, we investigate the molecular mechanism by which hypoxia promotes HCC invasion and metastasis through inducing epithelial-mesenchymal transition (EMT).MethodsThe expression of EMT markers was analyzed by immunohistochemistry. Effect of hypoxia on induction of EMT and ability of cell migration and invasion were performed. Luciferase reporter system was used for evaluation of Snail regulation by hypoxia-inducible factor -1α (HIF-1α).ResultsWe found that overexpression of HIF-1α was observed in HCC liver tissues and was related to poor prognosis of HCC patients. HIF-1α expression profile was correlated with the expression levels of SNAI1, E-cadherin, N-cadherin and Vimentin. Hypoxia was able to induce EMT and enhance ability of invasion and migration in HCC cells. The same phenomena were also observed in CoCl2-treated cells. The shRNA-mediated HIF-1α suppression abrogated CoCl2-induced EMT and reduced ability of migration and invasion in HCC cells. Luciferase assay showed that HIF-1α transcriptional regulated the expression of SNAI1 based on two hypoxia response elements (HREs) in SNAI1 promoter.ConclusionsWe demonstrated that hypoxia-stabilized HIF1α promoted EMT through increasing SNAI1 transcription in HCC cells. This data provided a potential therapeutic target for HCC treatment.

Light is one of the most important environmental cues regulating multiple aspects of plant growth and development, and abscisic acid (ABA) is a plant hormone that plays important roles during many phases of the plant life cycle and in plants' responses to various environmental stresses. How plants integrate the external light signal with endogenous ABA pathway for better adaptation and survival remains poorly understood. Here, we show that BBX21 (also known as SALT TOLERANCE HOMOLOG 2), a B-box (BBX) protein previously shown to positively regulate seedling photomorphogenesis, is also involved in ABA signaling. Our genetic data show that BBX21 may act upstream of several ABA INSENSITIVE (ABI) genes and ELONGATED HYPOCOTYL 5 (HY5) in ABA control of seed germination. Previous studies showed that HY5 acts as a direct activator of ABI5 expression, and that BBX21 interacts with HY5. We further demonstrate that BBX21 negatively regulates ABI5 expression by interfering with HY5 binding to the ABI5 promoter. In addition, ABI5 was shown to directly activate its own expression, whereas BBX21 negatively regulates this activity by directly interacting with ABI5. Together, our study indicates that BBX21 coordinates with HY5 and ABI5 on the ABI5 promoter and that these transcriptional regulators work in concert to integrate light and ABA signaling in Arabidopsis thaliana.

Plants have evolved a delicate molecular system to fine-tune their growth and development in response to dynamically changing light environments. In this study, we found that BBX28, a B-box domain protein, negatively regulates photomorphogenic development in a dose-dependent manner in Arabidopsis thaliana. BBX28 interferes with the binding of transcription factor HY5 to the promoters of its target genes through physical interactions, thereby repressing its activity and negatively affecting HY5-regulated gene expression. In darkness, BBX28 associates with CONSTITUTIVELY PHOTOMORPHOGENIC1 (COP1) and undergoes COP1-mediated degradation via the 26S proteasome system. Collectively, these results demonstrate that BBX28 acts as a key factor in the COP1-HY5 regulatory hub by maintaining proper HY5 activity to ensure normal photomorphogenic development in plants.

Several different species of Pseudomonas produce N-acylhomoserine lactones (AHLs), quorum-sensing signal molecules which are involved in the cell-densitydependent control of secondary metabolite and virulence gene expression. When Pseudomonas fluorescens F113 was cross-streaked against AHL biosensors capable of sensitively detecting either short (C 4 -C 8 ) or long (C 10 -C 14 ) acyl chain AHLs, no activity was detectable. However, by extracting cell-free stationary-phase culture supernatants with dichloromethane followed by reverse-phase HPLC, three distinct fractions were obtained capable of activating the AHL biosensors. Three AHLs were subsequently characterized using high-resolution MS and chemical synthesis. These were (i) N-(3-hydroxy-7-cis-tetradecenoyl)homoserine lactone (3OH,C 14 :1 -HSL), a molecule previously known as the Rhizobium leguminosarum small bacteriocin as a consequence of its growth inhibitory properties, (ii) N-decanoylhomoserine lactone (C 10 -HSL) and (iii) N-hexanoylhomoserine lactone (C 6 -HSL). A gene (hdtS) capable of directing synthesis of all three P. fluorescens AHLs in Escherichia coli was cloned and sequenced. In vitro transcription/translation of hdtS yielded a protein of approximately 33 kDa capable of directing the synthesis of 3OH,C 14 :1 -HSL, C 10 -HSL and C 6 -HSL in E. coli. HdtS does not belong to either of the known AHL synthase families (LuxI or LuxM) and is related to the lysophosphatidic acid acyltransferase family. HdtS may therefore constitute a member of a third protein family capable of AHL biosynthesis.

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Light-mediated seedling development is coordinately controlled by a variety of key regulators. Here, we identified two B-box (BBX)-containing proteins, BBX30 and BBX31, as repressors of photomorphogenesis. ELONGATED HYPOCOTYL5, a central regulator of light signaling, directly binds to the G-box cis-element present in the promoters of BBX30 and BBX31 and negatively controls their transcription levels in the light. Seedlings with mutations in BBX30 or BBX31 are hypersensitive to light, whereas the overexpression of BBX30 or BBX31 leads to hypo-photomorphogenic growth in the light. Furthermore, transgenic and phenotypic analysis revealed that the B-box domain of BBX30 or BBX31 is essential for their respective functioning in the regulation of photomorphogenic development in plants. In conclusion, BBX30 and BBX31 act as key negative regulators of light signaling, and their transcription is repressed by ELONGATED HYPOCOTYL5 through directly associating with their promoters.