Enhancer‐mediated reporter gene expression in <i>Arabidopsis thaliana</i>: a forward genetic screen

Lin, Yuan; Zhao, Hainan; Kotlarz, Magdalena; Jiang, Jiming

doi:10.1111/tpj.15189

Cited by 6 publications

(3 citation statements)

References 78 publications

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“…In addition to the aforementioned perspectives, the application of the proposed data integration strategy to other complex organisms is a promising lead. In this work, TFBM influencing gene expression were assumed to be located in the promoter regions of their target genes because very few distal regulations have been reported in Arabidopsis, and are still poorly understood [35]. In organisms where regulation by distant enhancers is well documented and responsible for tissue-specificity [4, 28], delineating enhancer regions may be achieved through the use of additional molecular layers such as chromatin accessibility, chromatin contacts, or eQTLs.…”

Section: Discussionmentioning

confidence: 99%

Optimizing data integration improves Gene Regulatory Network inference in Arabidopsis thaliana

Cassan,

Lecellier,

Martin

et al. 2023

Preprint

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MotivationsGene Regulatory Networks (GRN) are traditionnally inferred from gene expression profiles monitoring a specific condition or treatment. In the last decade, integrative strategies have successfully emerged to guide GRN inference from gene expression with complementary prior data. However, datasets used as prior information and validation gold standards are often related and limited to a subset of genes. This lack of complete and independent evaluation calls for new criteria to robustly estimate the optimal intensity of prior data integration in the inference process.ResultsWe address this issue for two common regression-based GRN inference models, an integrative Random Forest (weigthedRF) and a generalized linear model with stability selection estimated under a weighted LASSO penalty (weightedLASSO). These approaches are applied to data from the root response to nitrate induction inArabidopsis thaliana. For each gene, we measure how the integration of transcription factor binding motifs influences model prediction. We propose a new approach, DIOgene, that uses model prediction error and a simulated null hypothesis for optimizing data integration strength in a hypothesis-driven, gene-specific manner. The resulting integration scheme reveals a strong diversity of optimal integration intensities between genes. In addition, it provides a good trade-off between prediction error minimization and validation on experimental interactions, while master regulators of nitrate induction can be accurately retrieved.Availability and implementationThe R code and notebooks demonstrating the use of the proposed approaches are available in the repositoryhttps://github.com/OceaneCsn/integrative_GRN_N_induction.

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Section: Discussionmentioning

confidence: 99%

Optimizing data integration improves Gene Regulatory Network inference in Arabidopsis thaliana

Cassan,

Lecellier,

Martin

et al. 2023

Preprint

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“…Various factors, such as Agrobacterium strains, plant species or genotypes, and cocultural conditions, have been reported to affect AMTE efficiency [27,35]. The optimization of these factors has been the general focus for further improvement of gene expression.…”

Section: Discussionmentioning

confidence: 99%

An Agrobacterium-Mediated Transient Expression Method for Functional Assay of Genes Promoting Disease in Monocots

Chang

Huang

et al. 2023

IJMS

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Agrobacterium-mediated transient expression (AMTE) has been widely used for high-throughput assays of gene function in diverse plant species. However, its application in monocots is still limited due to low expression efficiency. Here, by using histochemical staining and a quantitative fluorescence assay of β-glucuronidase (GUS) gene expression, we investigated factors affecting the efficiency of AMTE on intact barley plants. We found prominent variation in GUS expression levels across diverse vectors commonly used for stable transformation and that the vector pCBEP produced the highest expression. Additionally, concurrent treatments of plants with one day of high humidity and two days of darkness following agro-infiltration also significantly increased GUS expression efficiency. We thus established an optimized method for efficient AMTE on barley and further demonstrated its efficiency on wheat and rice plants. We showed that this approach could produce enough proteins suitable for split-luciferase assays of protein-protein interactions on barley leaves. Moreover, we incorporated the AMTE protocol into the functional dissection of a complex biological process such as plant disease. Based on our previous research, we used the pCBEP vector to construct a full-length cDNA library of genes upregulated during the early stage of rice blast disease. A subsequent screen of the library by AMTE identified 15 candidate genes (out of ~2000 clones) promoting blast disease on barley plants. Four identified genes encode chloroplast-related proteins: OsNYC3, OsNUDX21, OsMRS2-9, and OsAk2. These genes were induced during rice blast disease; however, constitutive overexpression of these genes conferred enhanced disease susceptibility to Colletotrichum higginsianum in Arabidopsis. These observations highlight the power of the optimized AMTE approach on monocots as an effective tool for facilitating functional assays of genes mediating complex processes such as plant-microbe interactions.

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“…1a-d). This finding was surprising because distal regulatory elements that control transcription from such a long distance are commonly found in animals but are less known in plants [22][23][24][25] . Histone H3K9 acetylation was significantly increased at this site upon WOX5 induction (Extended Data Fig.…”

Section: Wox5 Upregulates Han Transcription During Ectopic Stem Cell ...mentioning

confidence: 99%

A coherent feed-forward-loop in the Arabidopsis root stem cell organizer regulates auxin biosynthesis and columella stem cell maintenance

Sharma,

Friedrich,

Peruzzo

et al. 2023

Preprint

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Stem cells in the plant meristems are kept undifferentiated by signals from surrounding cells and provide the basis for continuous organ formation. In the stem cell organizer of the Arabidopsis thaliana root, the Quiescent Center (QC), the WOX5 transcription factor functions as a central hub in regulating columella stem cell (CSC) homeostasis. However, the processes mediating WOX5 function have yet to be discovered. Here, we identify the transcription factor HAN as a central mediator of WOX5-regulated stem cell maintenance. HAN is required and sufficient to maintain CSCs undifferentiated and to induce ectopic stem cells. WOX5 and HAN repress transcription of the differentiation factor gene CDF4 in a coherent Feed Forward Loop (cFFL), one output of which is the expression of the auxin biosynthesis gene TAA1 and maintaining auxin response maxima in the organizer. Mathematical WOX5/HAN/CDF4cFFL modeling suggests a mechanism to buffer columella stem cell maintenance against input noise.

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Enhancer‐mediated reporter gene expression in Arabidopsis thaliana: a forward genetic screen

Cited by 6 publications

References 78 publications

Optimizing data integration improves Gene Regulatory Network inference in Arabidopsis thaliana

Optimizing data integration improves Gene Regulatory Network inference in Arabidopsis thaliana

An Agrobacterium-Mediated Transient Expression Method for Functional Assay of Genes Promoting Disease in Monocots

A coherent feed-forward-loop in the Arabidopsis root stem cell organizer regulates auxin biosynthesis and columella stem cell maintenance

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