Adverse environmental conditions produce endoplasmic reticulum (ER) stress in plants. In response to heat or ER stress agents, Arabidopsis seedlings mitigate stress damage by activating ERassociated transcription factors and a RNA splicing factor, IRE1b. IRE1b splices the mRNA-encoding bZIP60, a basic leucine-zipper domain containing transcription factor associated with the unfolded protein response in plants. bZIP60 is required for the upregulation of BINDING PROTEIN3 (BIP3) in response to ER stress, and loss-of-function mutations in IRE1b or point mutations in the splicing site of bZIP60 mRNA are defective in BIP3 induction. These findings demonstrate that bZIP60 in plants is activated by RNA splicing and afford opportunities for monitoring and modulating stress responses in plants.abiotic stress | heat stress | RNA splicing | signal transduction H eat and drought tolerance are some of the most complex and important adaptive traits in plants. These stresses are foremost in placing limits on plant productivity worldwide, and tolerance to these stresses are among the most highly sought after traits in crops (1), particularly in the face of climate change.The unfolded protein response (UPR) in eukaryotes is an ER stress response that activates three different classes of membraneassociated sensor transducers in mammalian cells-activating transcription factor 6 (ATF6), inositol-requiring enzyme-1 (IRE1) and protein kinase RNA (PKR)-like ER kinase. Yeast has only one ER stress transducer, IRE1; nonetheless, this factor sets off a massive UPR by triggering the expression of >5% of genes in the yeast genome. Many of these encode chaperones and ER-associated protein degradation components (2). IRE1 in yeast and mammalian cells acts by splicing a messenger RNA encoding a transcription factor that, then in turn, activates the expression of stress response genes (see recent reviews; refs. 3-5). Yeast cells splice an mRNA encoding a transcription factor called Hac1p (6, 7). The unspliced form of the Hac1 messenger RNA attenuates its own translation, and splicing relieves the translational repression (8).IRE1-mediated splicing is unconventional because mRNA splicing normally occurs in the nucleus, not in the cytoplasm (9). IRE1 is a type I membrane-spanning protein situated in the ER with its N terminus facing the ER lumen and its C terminus, which possesses catalytic functions, facing the cytosol. IRE1 is regarded as a dual functional enzyme possessing both serine/threonine protein kinase and endoribonuclease activity (10). Upon activation, the IRE1 dimer undergoes autotransphosphorylation in which one monomer phosphorylates the other (11). Through the analysis of the structure of the cytosolic domain of IRE1, Lee et al. (12) found that dimerization brings together the kinase domains in a face-to-face manner that would seemingly facilitate autotransphosphorylation.Autotransphosphorylation is then thought to open a nucleotide-binding site that, when occupied, produces a conformational change in the cytosolic domain so as t...

Cytokinins (CKs) are plant hormones that regulate a large number of processes associated with plant growth and development such as induction of stomata opening, delayed senescence, suppression of auxin-induced apical dominance, signaling of nitrogen availability, differentiation of plastids and control of sink strength. In maize, CKs are thought to play an important role in establishing seed size and increasing seed set under normal and unfavorable environmental conditions therefore influencing yield. In recent years, the discovery of isopentenyl transferase (IPT) genes in plants has shed light on the CK biosynthesis pathway in plants. In an effort to increase our understanding of the role played by CKs in maize development and sink-strength, we identified several putative IPT genes using a bioinformatics approach. We focused our attention on one gene in particular, ZmIPT2, because of its strong expression in developing kernels. The expression of the gene and its product overlays the change in CK levels in developing kernels suggesting a major role in CK biosynthesis for kernel development. We demonstrate that at 8-10 days after pollination (DAP) the endosperm and especially the basal transfer cell layer (BETL) is a major site of ZmIPT2 expression, and that this expression persists in the BETL and the developing embryo into later kernel development stages. We show that ectopic expression of ZmIPT2 in calli and in planta created phenotypes consistent with CK overproduction. We also show that ZmIPT2 preferentially uses ADP and ATP over AMP as the substrates for dimethylallyl diphosphate (DMAPP) IPT activity. The expression pattern of ZmIPT2 in the BETL, endosperm and embryo during kernel development will be discussed with an emphasis on the suggested role of CKs in determining sink-strength and grain production in crop plants.

BackgroundAdverse environmental conditions produce ER stress and elicit the unfolded protein response (UPR) in plants. Plants are reported to have two "arms" of the ER stress signaling pathway-one arm involving membrane-bound transcription factors and the other involving a membrane-associated RNA splicing factor, IRE1. IRE1 in yeast to mammals recognizes a conserved twin loop structure in the target RNA.ResultsA segment of the mRNA encoding ZmbZIP60 in maize can be folded into a twin loop structure, and in response to ER stress this mRNA is spliced, excising a 20b intron. Splicing converts the predicted protein from a membrane-associated transcription factor to one that is targeted to the nucleus. Splicing of ZmbZIP60 can be elicited in maize seedlings by ER stress agents such as dithiothreitol (DTT) or tunicamycin (TM) or by heat treatment. Younger, rather than older seedlings display a more robust splicing response as do younger parts of leaf, along a developmental gradient in a leaf. The molecular signature of an ER stress response in plants includes the upregulation of Binding Protein (BIP) genes. Maize has numerous BIP-like genes, and ER stress was found to upregulate one of these, ZmBIPb.ConclusionsThe splicing of ZmbZIP60 mRNA is an indicator of ER stress in maize seedlings resulting from adverse environmental conditions such as heat stress. ZmbZIP60 mRNA splicing in maize leads predictively to the formation of active bZIP transcription factor targeted to the nucleus to upregulate stress response genes. Among the genes upregulated by ER stress in maize is one of 22 BIP-like genes, ZmBIPb.

BackgroundWater and nitrogen are two of the most critical inputs required to achieve the high yield potential of modern corn varieties. Under most agricultural settings however they are often scarce and costly. Fortunately, tremendous progress has been made in the past decades in terms of modeling to assist growers in the decision making process and many tools are now available to achieve more sustainable practices both environmentally and economically. Nevertheless large gaps remain between our empirical knowledge of the physiological changes observed in the field in response to nitrogen and water stresses, and our limited understanding of the molecular processes leading to those changes.ResultsThis work examines in particular the impact of simultaneous stresses on the transcriptome. In a greenhouse setting, corn plants were grown under tightly controlled nitrogen and water conditions, allowing sampling of various tissues and stress combinations. A microarray profiling experiment was performed using this material and showed that the concomitant presence of nitrogen and water limitation affects gene expression to an extent much larger than anticipated. A clustering analysis also revealed how the interaction between the two stresses shapes the patterns of gene expression over various levels of water stresses and recovery.ConclusionsOverall, this study suggests that the molecular signature of a specific combination of stresses on the transcriptome might be as unique as the impact of individual stresses, and hence underlines the difficulty to extrapolate conclusions obtained from the study of individual stress responses to more complex settings.