FUSCA3 from barley unveils a common transcriptional regulation of seed-specific genes between cereals and Arabidopsis.

Accumulation of storage compounds in the embryo and endosperm of developing seeds is a highly regulated process that allows seedling growth upon germination until photosynthetic capacity is acquired. A critical regulatory element in the promoters of seed storage protein (SSP) genes from dicotyledonous species is the RY box, a target of B3-type transcription factors. However, the functionality of this motif in the transcriptional regulation of SSP genes from cereals has not been fully established. We report here the identification and molecular characterization of barley FUSCA3, a B3-type transcription factor as yet uncharacterized in monocotyledonous plants. Our results show that both the barley and Arabidopsis FUS3 genes maintain a conserved functionality for the regulation of SSP genes and anthocyanin biosynthesis in these two distantly related phylogenetic groups. Complementation of the loss-of-function mutant fus3 in Arabidopsis by the barley HvFus3 gene resulted in restored transcription from the At2S3 gene promoter and normal accumulation of anthocyanins in the seed. In barley, HvFUS3 participates in transcriptional activation of the endosperm-specific genes Hor2 and Itr1. HvFUS3, which specifically binds to RY boxes in EMSA experiments, trans-activates Hor2 and Itr1 promoters containing intact RY boxes in transient expression assays in developing endosperms. Mutations in the RY boxes abolished the HvFUS3-mediated trans-activation. HvFus3 transcripts accumulate in the endosperm and in the embryo of developing seeds, peaking at mid maturation phase. Remarkably, HvFUS3 interacts with the Opaque2-like bZIP factor BLZ2 in yeast, and this interaction is essential for full trans-activation of the seed-specific genes in plant

Identification of Novel Components of the Unfolded Protein Response in Arabidopsis

Unfavorable environmental and developmental conditions may cause disturbances in protein folding in the endoplasmic reticulum (ER) that are recognized and counteracted by components of the Unfolded Protein Response (UPR) signaling pathways. The early cellular responses include transcriptional changes to increase the folding and processing capacity of the ER. In this study, we systematically screened a collection of inducible transgenic Arabidopsis plants expressing a library of transcription factors for resistance toward UPR-inducing chemicals. We identified 23 candidate genes that may function as novel regulators of the UPR and of which only three genes (bZIP10, TBF1, and NF-YB3) were previously associated with the UPR. The putative role of identified candidate genes in the UPR signaling is supported by favorable expression patterns in both developmental and stress transcriptional analyses. We demonstrated that WRKY75 is a genuine regulator of the ER-stress cellular responses as its expression was found to be directly responding to ER stress-inducing chemicals. In addition, transgenic Arabidopsis plants expressing WRKY75 showed resistance toward salt stress, connecting abiotic and ER-stress responses

A Developmental Switch of Gene Expression in the Barley Seed Mediated by HvVP1 (Viviparous-1) and HvGAMYB Interactions

The accumulation of storage compounds in the starchy endosperm of developing cereal seeds is highly regulated at the transcriptional level. These compounds, mainly starch and proteins, are hydrolyzed upon germination to allow seedling growth. The transcription factor HvGAMYB is a master activator both in the maturation phase of seed development and upon germination, acting in combination with other transcription factors. However, the precise mechanism controlling the switch from maturation to germination programs remains unclear. We report here the identification and molecular characterization of Hordeum vulgare VIVIPAROUS1 (HvVP1), orthologous to ABA-INSENSITIVE3 from Arabidopsis thaliana. HvVP1 transcripts accumulate in the endosperm and the embryo of developing seeds at early stages and in the embryo and aleurone of germinating seeds up to 24 h of imbibition. In transient expression assays, HvVP1 controls the activation of Hor2 and Amy6.4 promoters exerted by HvGAMYB. HvVP1 interacts with HvGAMYB in Saccharomyces cerevisiae and in the plant nuclei, hindering its interaction with other transcription factors involved in seed gene expression programs, like BPBF. Similarly, this interaction leads to a decrease in the DNA binding of HvGAMYB and the Barley Prolamine-Box binding Factor (BPBF) to their target sequences. Our results indicate that the HvVP1 expression pattern controls the full Hor2 expression activated by GAMYB and BPBF in the developing endosperm and the Amy6.4 activation in postgerminative reserve mobilization mediated by GAMYB. All these data demonstrate the participation of HvVP1 in antagonistic gene expression programs and support its central role as a gene expression switch during seed maturation and germination

Transcriptional Control of Glutaredoxin GRXC9 Expression by a Salicylic Acid-Dependent and NPR1-Independent Pathway in Arabidopsis

Salicylic acid (SA) is a key hormone that mediates gene transcriptional reprogramming in the context of the defense response to stress. GRXC9, coding for a CC-type glutaredoxin from Arabidopsis, is an SA-responsive gene induced early and transiently by an NPR1-independent pathway. Here, we address the mechanism involved in this SA-dependent pathway, using GRXC9 as a model gene. We first established that GRXC9 expression is induced by UVB exposure through this pathway, validating its activation in a physiological stress condition. GRXC9 promoter analyses indicate that SA controls gene transcription through two activating sequence-1 (as-1)-like elements located in its proximal region. TGA2 and TGA3, but not TGA1, are constitutively bound to this promoter region. Accordingly, the transient recruitment of RNA polymerase II to the GRXC9 promoter, as well as the transient accumulation of gene transcripts detected in SA-treated WT plants, was abolished in a knockout mutant for the TGA class II factors. We conclude that constitutive binding of TGA2 is essential for controlling GRXC9 expression, while binding of TGA3 in a lesser extent contributes to this regulation. Finally, overexpression of GRXC9 indicates that the GRXC9 protein negatively controls its own gene expression, forming part of the complex bound to the as-1-containing promoter region. These findings are integrated in a model that explains how SA controls transcription of GRXC9 in the context of the defense response to stress

Expression patterns within the Arabidopsis C/S1 bZIP transcription factor network: availability of heterodimerization partners controls gene expression during stress response and development

Members of the Arabidopsis group C/S1 basic leucine zipper (bZIP) transcription factor (TF) network are proposed to implement transcriptional reprogramming of plant growth in response to energy deprivation and environmental stresses. The four group C and five group S1 members form specific heterodimers and are, therefore, considered to cooperate functionally. For example, the interplay of C/S1 bZIP TFs in regulating seed maturation genes was analyzed by expression studies and target gene regulation in both protoplasts and transgenic plants. The abundance of the heterodimerization partners significantly affects target gene transcription. Therefore, a detailed analysis of the developmental and stress related expression patterns was performed by comparing promoter: GUS and transcription data. The idea that the C/S1 network plays a role in the allocation of nutrients is supported by the defined and partially overlapping expression patterns in sink leaves, seeds and anthers. Accordingly, metabolic signals strongly affect bZIP expression on the transcriptional and/or post-transcriptional level. Sucrose induced repression of translation (SIRT) was demonstrated for all group S1 bZIPs. In particular, transcription of group S1 genes strongly responds to various abiotic stresses, such as salt (AtbZIP1) or cold (AtbZIP44). In summary, heterodimerization and expression data provide a basic framework to further determine the functional impact of the C/S1 network in regulating the plant energy balance and nutrient allocation

YUCCA8 and YUCCA9 overexpression reveals a link between auxin signaling and lignification through the induction of ethylene biosynthesis.

Auxin is associated with the regulation of virtually every aspect of plant growth and development. Many previous genetic and biochemical studies revealed that, among the proposed routes for the production of auxin, the so-called indole-3-pyruvic acid (IPA) pathway is the main source for indole-3-acetic acid (IAA) in plants. The IPA pathway involves the action of 2 classes of enzymes, tryptophan-pyruvate aminotransferases (TRYPTOPHAN AMINOTRANSFERASE OF ARABIDOPSIS 1(TAA1)/TRYPTOPHAN AMINOTRANSFERASE RELATED (TAR)) and flavin monooxygenases (YUCCA). Both enzyme classes appear to be encoded by small gene families in Arabidopsis consisting of 5 and 11 members, respectively. We recently showed that it is possible to induce transcript accumulation of 2 YUCCA genes, YUC8 and YUC9, by methyl jasmonate treatment. Both gene products were demonstrated to contribute to auxin biosynthesis in planta.1 Here we report that the overexpression of YUC8 as well as YUC9 led to strong lignification of plant aerial tissues. Furthermore, new evidence indicates that this abnormally strong secondary growth is linked to increased levels of ethylene production

SnRK1-triggered switch of bZIP63 dimerization mediates the low-energy response in plants

Metabolic adjustment to changing environmental conditions, particularly balancing of growth and defense responses, is crucial for all organisms to survive. The evolutionary conserved AMPK/Snf1/SnRK1 kinases are well-known metabolic master regulators in the low-energy response in animals, yeast and plants. They act at two different levels: by modulating the activity of key metabolic enzymes, and by massive transcriptional reprogramming. While the first part is well established, the latter function is only partially understood in animals and not at all in plants. Here we identified the Arabidopsis transcription factor bZIP63 as key regulator of the starvation response and direct target of the SnRK1 kinase. Phosphorylation of bZIP63 by SnRK1 changed its dimerization preference, thereby affecting target gene expression and ultimately primary metabolism. A bzip63 knock-out mutant exhibited starvation-related phenotypes, which could be functionally complemented by wild type bZIP63, but not by a version harboring point mutations in the identified SnRK1 target sites