cyclic-gmp and gibberellic-acid

An ever increasing amount of transcriptomic data and analysis tools provide novel insight into complex responses of biological systems. Given these resources we have undertaken to review aspects of transcriptional regulation in response to the plant hormone gibberellic acid (GA) and its second messenger guanosine 3',5'-cyclic monophosphate (cGMP) in Arabidopsis thaliana, both wild type and selected mutants. Evidence suggests enrichment of GA-responsive (GARE) elements in promoters of genes that are transcriptionally upregulated in response to cGMP but downregulated in a GA insensitive mutant (ga1-3). In contrast, in the genes upregulated in the mutant, no enrichment in the GARE is observed suggesting that GARE motifs are diagnostic for GA-induced and cGMP-dependent transcriptional upregulation. Further, we review how expression studies of GA-dependent transcription factors and transcriptional networks based on common promoter signatures derived from ab initio analyses can contribute to our understanding of plant responses at the systems level.

Topics: Arabidopsis; Cyclic GMP; Gene Expression Regulation, Plant; Gene Regulatory Networks; Gibberellins; Mutation; Plant Growth Regulators; Transcription Factors; Transcription, Genetic

Cyclic AMP (cAMP) and cyclic GMP (cGMP) have roles in relaying external signals and modifying gene expression within cells in all phyla. Currently there are no reporter systems suitable for bacteria and plant cells that measure alterations in downstream gene expression following changes in intracellular levels of cyclic nucleotides. As the plant protein OLIGOPEPTIDE TRANSPORTER X (OPTX) is upregulated by cGMP, we fused the OPTX promoter to a luciferase reporter gene (OPTX:LUC) to develop a plant cell reporter of cGMP-induced gene expression. We prepared a second construct augmented with three mammalian cGMP response elements (OPTXcGMPRE:LUC) and a third construct containing five gibberellic acid response elements (OPTXGARE:LUC). All three constructs were tested in bacteria and isolated plant protoplasts.. Membrane permeable cGMP enhanced luciferase activity of OPTX:LUC and OPTXGARE:LUC in protoplasts. Treatment with the plant hormone gibberellic acid which acts via cGMP also generated downstream luciferase activity. However, membrane permeable cAMP induced similar responses to cGMP in protoplasts. Significantly increased luciferase activity occurred in bacteria transformed with either OPTXcGMPRE:LUC or OPTXGARE:LUC in response to membrane permeable cAMP and cGMP. Bacteria co-transformed with OPTXcGMPRE:LUC or OPTXGARE:LUC and the soluble cytoplasmic domain of phytosulfokine receptor1 (PSKR1; a novel guanylate cyclase) had enhanced luciferase activity following induction of PSKR1 expression.. We have developed promoter reporter systems based on the plant OPTX promoter that can be employed in bacteria and isolated plant cells. We have shown that it can be used in bacteria to screen recombinant proteins for guanylate cyclase activity as increases in intracellular cGMP levels result in altered gene transcription and luciferase activity.

Topics: Bacteria; Cyclic AMP; Cyclic GMP; Escherichia coli; Gene Expression; Genes, Reporter; Gibberellins; Guanylate Cyclase; Luciferases; Membrane Transport Proteins; Nitric Oxide; Plant Cells; Plant Proteins; Promoter Regions, Genetic; Protoplasts; Recombinant Proteins; Response Elements; Transcription, Genetic; Up-Regulation

Here, α-Amy2/54 gene expression was used as a molecular probe to investigate the interrelationship among nitric oxide (NO), cyclic GMP (cGMP), and heme oxygenase-1 (HO-1) in GA-treated wheat aleurone layers. The inducible expressions of α-Amy2/54 and α-amylase activity were respectively amplified by two NO-releasing compounds, sodium nitroprusside (SNP) and spermine NONOate, in a GA-dependent fashion. Similar responses were observed when an inducer of HO-1, hemin-or one of its catalytic products, carbon monoxide (CO) in aqueous solution-was respectively added. The SNP-induced responses, mimicked by 8-bromoguanosine 3',5'-cyclic monophosphate (8-Br-cGMP), a cGMP derivative, were NO-dependent. This conclusion was supported by the fact that endogenous NO overproduction was rapidly induced by SNP, and thereafter induction of α-Amy2/54 gene expression and increased α-amylase activity were sensitive to the NO scavenger. We further observed that the above induction triggered by SNP and 8-Br-cGMP was partially prevented by zinc protoporphyrin IX (ZnPPIX), an inhibitor of HO-1. These blocking effects were clearly reversed by CO, confirming that the above response was HO-1-specific. Further analyses showed that both SNP and 8-Br-cGMP rapidly up-regulated HO-1 gene expression and increased HO activity, and SNP responses were sensitive to cPTIO and the guanylate cyclase inhibitor 6-anilino-5,8-quinolinedione (LY83583). Molecular evidence confirmed that GA-induced GAMYB and ABA-triggered PKABA1 transcripts were up-regulated or down-regulated by SNP, 8-Br-cGMP or CO cotreated with GA. Contrasting changes were observed when cPTIO, LY83583, or ZnPPIX was added. Together, our results suggested that HO-1 is involved in NO- and cGMP-induced α-Amy2/54 gene expression in GA-treated aleurone layers.

Topics: alpha-Amylases; Aminoquinolines; Benzoates; Carbon Monoxide; Cyclic GMP; Enzyme Inhibitors; Gene Expression; Genes, Plant; Gibberellins; Guanylate Cyclase; Heme Oxygenase-1; Imidazoles; Nitric Oxide; Nitric Oxide Donors; Nitroprusside; Plant Growth Regulators; Plant Proteins; Protoporphyrins; Signal Transduction; Spermine; Triticum

The cyclic nucleotide cGMP has been shown to play important roles in plant development and responses to abiotic and biotic stress. To date, the techniques that are available to measure cGMP in plants are limited by low spatial and temporal resolution. In addition, tissue destruction is necessary. To circumvent these drawbacks we have used the δ-FlincG fluorescent protein to create an endogenous cGMP sensor that can report cellular cGMP levels with high resolution in time and space in living plant cells. δ-FlincG in transient and stably expressing cells shows a dissociation constant for cGMP of around 200 nm giving it a dynamic range of around 20-2000 nm. Stimuli that were previously shown to alter cGMP in plant cells (nitric oxide and gibberrellic acid) evoked pronounced fluorescence signals in single cells and in root tissues, providing evidence that δ-FlincG reports changes in cellular cGMP in a physiologically relevant context.

Topics: Arabidopsis; Cyclic GMP; Enzyme-Linked Immunosorbent Assay; Fluorescent Dyes; Gibberellins; Green Fluorescent Proteins; Microscopy, Confocal; Nitric Oxide; Oryza; Plant Growth Regulators; Protoplasts

Cyclic guanosine 3',5'-monophosphate (cGMP) is an important second messenger in animals, and is emerging as a player in regulatory functions in plants. In this study, we investigated the role of cGMP in seed germination in Arabidopsis thaliana (Col-0). We demonstrated that both, a membrane-permeant analogue of cGMP (8-Br-cGMP) and the cyclic nucleotide phosphodiesterase (PDE) inhibitor Tadalafil promoted A. thaliana seed germination, whereas the guanylate cyclase inhibitor LY 83583 (6-anilino-5,8-quinolinedione; LY) inhibited it. LY blocked gibberellic acid (GA)-induced seed germination, whereas GA and 8-Br-cGMP co-treatment increased the germination rate and more effectively overcame LY-inhibition than 8-Br-cGMP alone. The gibberellin biosynthesis inhibitor paclobutrazol (PAC) also blocked 8-Br-cGMP and Tadalafil promotion of seed germination. Furthermore, 8-Br-cGMP and Tadalafil decreased abscisic acid (ABA) sensitivity during seed germination. These findings highlight that cGMP is a positive regulator and plays a crucial role in Arabidopsis seed germination. Furthermore, both GA and cGMP are required for seed germination.

Topics: Abscisic Acid; Aminoquinolines; Arabidopsis; Carbolines; Cyclic GMP; Enzyme Inhibitors; Germination; Gibberellins; Guanylate Cyclase; Phosphodiesterase Inhibitors; Seeds; Tadalafil; Triazoles