RESUMEN
Nitrogen (N) is often a limiting nutrient whose availability determines plant growth and productivity. Because its availability is often low and/or not uniform over time and space in nature, plants respond to variations in N availability by altering uptake and recycling mechanisms, but the molecular mechanisms underlying how these responses are regulated are poorly understood. Here, we show that a group of GARP G2-like transcription factors, Arabidopsis thaliana NITRATE-INDUCIBLE, GARP-TYPE TRANSCRIPTIONAL REPRESSOR1/HYPERSENSITIVE TO LOW Pi-ELICITED PRIMARY ROOT SHORTENING1 proteins (NIGT1/HRS1s), are factors that bind to the promoter of the N starvation marker NRT2.4 and repress an array of N starvation-responsive genes under conditions of high N availability. Transient assays and expression analysis demonstrated that NIGT1/HRS1s are transcriptional repressors whose expression is regulated by N availability. We identified target genes of the NIGT1/HRS1s by genome-wide transcriptome analyses and found that they are significantly enriched in N starvation response-related genes, including N acquisition, recycling, remobilization, and signaling genes. Loss of NIGT1/HRS1s resulted in deregulation of N acquisition and accumulation. We propose that NIGT1/HRS1s are major regulators of N starvation responses that play an important role in optimizing N acquisition and utilization under fluctuating N conditions.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Regulación de la Expresión Génica de las Plantas , Nitrógeno/metabolismo , Factores de Transcripción/metabolismo , Arabidopsis/fisiología , Proteínas de Arabidopsis/genética , Transporte Biológico , Biomarcadores/metabolismo , Perfilación de la Expresión Génica , Regiones Promotoras Genéticas/genética , Factores de Transcripción/genéticaRESUMEN
Arabidopsis thaliana DEHYDRATION-RESPONSIVE ELEMENT BINDING PROTEIN2A (DREB2A) functions as a transcriptional activator that increases tolerance to osmotic and heat stresses; however, its expression also leads to growth retardation and reduced reproduction. To avoid these adverse effects, the expression of DREB2A is predicted to be tightly regulated. We identified a short promoter region of DREB2A that represses its expression under nonstress conditions. Yeast one-hybrid screening for interacting factors identified GROWTH-REGULATING FACTOR7 (GRF7). GRF7 bound to the DREB2A promoter and repressed its expression. In both artificial miRNA-silenced lines and a T-DNA insertion line of GRF7, DREB2A transcription was increased compared with the wild type under nonstress conditions. A previously undiscovered cis-element, GRF7-targeting cis-element (TGTCAGG), was identified as a target sequence of GRF7 in the short promoter region of DREB2A via electrophoretic mobility shift assays. Microarray analysis of GRF7 knockout plants showed that a large number of the upregulated genes in the mutant plants were also responsive to osmotic stress and/or abscisic acid. These results suggest that GRF7 functions as a repressor of a broad range of osmotic stress-responsive genes to prevent growth inhibition under normal conditions.
Asunto(s)
Ácido Abscísico/farmacología , Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Proteínas Represoras/metabolismo , Estrés Fisiológico , Factores de Transcripción/metabolismo , Arabidopsis/efectos de los fármacos , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Secuencia de Bases , Ensayo de Cambio de Movilidad Electroforética , Regulación de la Expresión Génica de las Plantas , Técnicas de Inactivación de Genes/métodos , Genes de Plantas , Vectores Genéticos/genética , Vectores Genéticos/metabolismo , Datos de Secuencia Molecular , Ósmosis , Regiones Promotoras Genéticas , Unión Proteica , Estructura Terciaria de Proteína , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Proteínas Represoras/genética , Transactivadores/genética , Transactivadores/metabolismo , Factores de Transcripción/genética , Transcripción Genética , Activación Transcripcional , Técnicas del Sistema de Dos Híbridos , Regulación hacia ArribaRESUMEN
Yeast one-hybrid screening is widely used for the identification of transcription factors (TFs) that interact with specific DNA sequences. However, screening a whole cDNA library is not efficient for the identification of TFs because TF genes represent only a small percentage of clones in a cDNA library. Here, we present the development of an efficient yeast one-hybrid screening system using a prey library composed only of approximately 1,500 TF cDNAs of Arabidopsis thaliana. This library enabled us to isolate a TF that binds to a specific promoter sequence with high efficiency, even when the promoter region of the gene of interest was directly employed as bait. Furthermore, this library was also successfully applied as a yeast two-hybrid library to find TFs that interact with specific proteins. This efficient system will contribute to the elucidation of gene regulatory networks in plants.