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Identification of BBX proteins as rate-limiting co-factors of HY5

ABSTRACT: As a source of both energy and environmental information, the importance for plants to monitor the incoming light is evidenced by the highly integrated nature of the light signal transduction pathway with numerous other pathways throughout plant development. One of these signal integrators is the bZIP transcription factor HY5 which holds a key role as a positive regulator of light signalling in plants. Although HY5 is described to act as a DNA-binding transcriptional regulator, the lack of any apparent transactivation domain makes it unclear how HY5 is able to accomplish its many functions. We describe the identification of three B-box containing proteins (BBX20, 21 and 22) as essential partners for HY5 dependent modulation of hypocotyl elongation, anthocyanin accumulation and transcriptional regulation. The bbx202122 triple mutant mimics the phenotypes of hy5 in the light and its strong suppression of the cop1 phenotype in darkness. Furthermore, RNA-seq experiments show that 84% of genes differentially regulated in bbx202122 are also HY5 regulated, consistent with the B-box proteins and HY5 acting interdependently.

ORGANISM(S): Arabidopsis thaliana

PROVIDER: GSE137147 | GEO | 2020/07/01

REPOSITORIES: GEO

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DET1-mediated COP1 regulation avoids HY5 activity over second-site targets to tune plant photomorphogenesis

Project description:DE-ETIOLATED1 (DET1) is a negative regulator of plant photomorphogenesis acting as a component of the C3D complex, which can further associate to CULLIN4 to form a CRL4C3D E3 ubiquitin ligase. CRL4C3D is thought to act together with CRL4COP1SPA ubiquitin ligase, to promote the ubiquitin-mediated degradation of the master regulatory transcription factor ELONGATED HYPOCOTYL5 (HY5), thereby controlling photomorphogenic gene regulatory networks. Yet, functional links between COP1 and DET1 have long remained elusive. Here, upon mass spectrometry identification of DET1 and COP1-associated proteins, we provide in vivo evidence that DET1 associates with COP1 to promote its destabilization, a process necessary to dampen HY5 protein abundance. By regulating HY5 over-accumulation, DET1 is critical to avoid its association to second-site loci, including many PIF3 target genes. Accordingly, excessive HY5 levels result in an increased HY5 repressive activity and are sufficient to trigger fusca-like phenotypes otherwise observed typically in COP1 and COP9 signalosome mutant seedlings. This study therefore identifies that DET1-mediated regulation of COP1 stability tunes down HY5 cistrome and avoids hyper-photomorphogenic responses that might compromise plant viability.

2021-09-09 | GSE155147 | GEO

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2024-02-06 | GSE254813 | GEO

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Chimeric activators and repressors define HY5 activity and a feedback control mechanism in Arabidopsis (HY5_ChIPseq)

Project description:Transcription regulation requires many protein interactions on chromatin, and only a subset of transcription factors have well-defined activation or repression domains. The Arabidopsis transcription factor HY5 controls critical growth-related gene expression programs during plant development, but it’s primary activity in regulating transcription remains unclear. To address this question, we generated constitutive repressor and activator HY5 fusion proteins to direct the expression of HY5 target genes. We used RNA-seq, ChIP-seq, and multiple phenotypes to demonstrate that HY5 depends on accessory factors to promote transcription and identify high confidence direct targets of HY5. We suggest that this strategy can be used broadly to define the transcription regulation activity and direct targets of transcription factors. Interestingly, this approach also revealed a mechanism by which HY5 promotes the accumulation of its own negative regulators. We show that HY5 directly regulates components of the COP1 E3-ubiquitin ligase complex, and by uncoupling this feedback loop we can induce partial de-etiolation in the dark. This provides a system by which plants can quickly repress growth upon light exposure. Lastly, we show that modulating this system can generate significant phenotypic diversity and provide proof of concept that these fusion proteins can modulate growth in tomato, opening a novel path toward selecting desirable traits in crop species.

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Chimeric activators and repressors define HY5 activity and a feedback control mechanism in Arabidopsis (HY5_RNAseq)

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Project description:The core regulatory mechanisms affecting the function of ELONGATED HYPOCOTYL 5 (HY5) during plant photomorphogenesis include dynamics in the transcriptional activity and protein stability of HY5 in response to changes in ambient light; however, little is known about the mediators of these processes. Here, we identified PHOTOREGULATORY PROTEIN KINASE 1 (PPK1), which interacts directly with and phosphorylates HY5, as one such mediator. PPK1 and its homologs play redundant roles in the negative regulation of photomorphogenesis. PPK1 phosphorylates HY5 at Thr55, Ser56, and Ser60. The phosphorylation of these residues is essential to establish high-affinity binding with B-BOX PROTEIN 24 and CONSTITUTIVE PHOTOMORPHOGENIC 1, which inhibit the transcriptional activity and promote the ubiquitination and degradation of HY5, respectively. As such, PPKs regulate not only the binding of HY5 to its target genes under light conditions,but also HY5 degradation when plants are transferred from light to dark conditions. Our data reveal a PPK-mediated phospho-code on HY5 that integrates the molecular mechanisms underlying the regulation of HY5 transcriptional activity and protein stability to precisely control plant photomorphogenesis.

2024-02-07 | PXD049266 |

Analysis of transcription factor HY5 genomic binding sites in Arabidopsis

Project description:The transcription factor HY5 acts downstream of multiple families of the photoreceptors and promotes photomorphogenesis. Although it is well accepted that HY5 acts to regulate target gene expression, in vivo binding of HY5 to any of its target gene promoters has yet to be demonstrated. Here we used a chromatin immunoprecipitation procedure to verify suspected in vivo HY5 binding sites. We demonstrated that in vivo association of HY5 with promoter targets is not altered under distinct light qualities or during light-to-dark transition. Coupled with DNA chip hybridization using high density 60-nucleotide oligomer microarray that contains one probe for every 500 nucleotides over the entire Arabidopsis genome, we mapped genome wide in vivo HY5 binding sites. This analysis showed that HY5 binds preferentially to promoter regions in vivo and revealed over 3 thousand chromosomal sites as putative HY5 binding targets. HY5 binding targets tend to be enriched in the early light responsive genes and transcription factor genes. Our data thus supports a model in which HY5 is a high hierarchical regulator of the transcriptional cascades for photomorphogenesis. Keywords: ChIP-chip

2007-03-01 | GSE6510 | GEO

expression profiling of wild type and the hy5 mutant in arabidopsis

Project description:To determine the functional relevance of the HY5 binding sites to the transcriptional regulation, we performed genome wide expression analysis using WT and hy5 grown in continuous white light for 4 days. Keywords: mutant/widetype comparative

2007-03-01 | GSE6517 | GEO

ChIP-chip analysis of GBF1 in the presence and absence of its heterodimerizing partners, HY5 and HYH

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2014-03-24 | GSE36965 | GEO

A missense mutation in COP1 SUPPRESSOR 5 causes reversed action in mediating photomorphogenic development

Project description:CONSTITUTIVELY PHOTOMORPHOGENIC 1 (COP1) mediates various cellular and physiological processes in plants by targeting a large number of substrates for ubiquitination and degradation. In this study, we reveal that a substitution of Pro for Leu at the site of 409 in COP1 SUPPRESSOR 5 (CSU5, also called WRKY32) largely suppresses the short hypocotyls and expanded cotyledon phenotypes of cop1-6. CSU5P409L promotes the hypocotyl growth and inhibits the opening of cotyledons in plants. Loss of CSU5 function mutant seedlings display elongated hypocotyls, whereas overexpression of CSU5 leads to shortened hypocotyls. CSU5 directly associates with the promoter regions of HY5 to activate its transcription. Although COP1 interacts with both CSU5 and CSU5P409L, it negatively controls the stability of CSU5, but not CSU5P409L. CSU5P409L exhibits enhanced DNA binding ability and affects the expression of more genes compared with CSU5. Our results not only reveal the molecular role for CSU5 in promoting photomorphogenesis, but also provide insights into manipulating plant growth by engineering key components of light signaling.

2022-08-23 | GSE183463 | GEO

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