Barley guard cell and subsidiary cell specific transcripts
Ontology highlight
ABSTRACT: We prepared RNA from total leaves and epidermal peels, containing living guard cells (GC) and subsidiary cells (SC) only. In the third sample we blended epidermal peels to mechanically destroy the SC, leading to a pure guard cells fraction. By RNAseq and substraction of GC from SC transcripts, we were able to reliably assign transcripts to the barley SCs and GCs.
Project description:We intend to identify transporters and their regulators involved in barley stomatal movement as well as components of the ABA signaling pathway. We isolated epidermal peels where only stomatal guard cells and their subsidiary cells survived, while other epidermal cell were mechanically disrupted and sequenced the resulting RNAs. Thus we analyzed transcripts differentially expressed between the barley stomatal complex and total leaves. These data served as the first overview of genes expressed in the stomatal complex and for cloning of relevant transporters.
Project description:Soil salinity is a major environmental constraint affecting crop growth and threatening global food security. Plants adapt to salinity by optimizing performance of stomata, the microscopic sphincters inserted into the wax-covered epidermis of the shoot, which balance CO2 intake for photosynthetic carbon gain and concomitant water loss. Stomata are formed by two guard cells (GCs) that are morphologically and functionally distinct from the other leaf cells. In order to better understand the molecular mechanisms underlying stomatal function under saline conditions we used proteomics approach to study isolated GCs from the salt-tolerant sugar beet species.
Project description:To identify genes of the guard cell transcriptome of Arabidopsis thaliana enriched guard cell samples were compared with total leaf tissue. Genes of the abscisic acid and humidity response of Arabidopsis thaliana guard cells were identified by treatment with ABA-Spray and low humidity. total samples analysed are 24: 4 biological independent replicates of: total leaf (COL-0) vs. enriched guard cells (COL-0); ABA-sprayed enriched guard cells (gl1-1) vs. control-sprayed enriched guard cells (gl1-1); low humidity (20%rh) treated enriched guard cells (COL-0) vs. high humidity (80%) treated enriched guard cells (COL-0)
Project description:To identify genes of the guard cell transkriptome of Arabidopsis thaliana enriched guard cell samples were compared with total leaf tissue. Genes of the abscisic acid and humidity response of Arabidopsis thaliana guard cells were identified by treatment with ABA-Spray and low humidity. Ost1-2 and slac1-3 mutants were compared to their wildtype. total samples analysed are 35: 4 biolocigal independent replicates of: total leaf (COL-0) vs. enriched guard cells (COL-0); ABA-sprayed enriched guard cells (gl1-1) vs. control-sprayed enriched guard cells (gl1-1); enriched guard cells (slac1-3) vs. enriched guard cells (gl1-1);guard cells (ost1-2) vs. guard cells (ler);low humidity(20%rh) treated enriched guard cells (COL-0) vs. high humidity(80%) treated enriched guard cells (COL0)
Project description:Stomata are highly specialized organs which consist of pairs of guard cells and regulate gas and water vapor exchange in plants. While early stages of guard cell differentiation have been described and were interpreted in analogy to processes of cell type differentiation in animals, the downstream development of functional stomatal guard cells remains poorly understood. We have isolated an Arabidopsis mutant, scap1 (stomatal carpenter 1), that develops irregularly shaped guard cells and lacks the ability to control stomatal aperture, including CO2-induced stomatal closing and light-induced stomatal opening. SCAP1 was identified as a plant-specific Dof-type transcription factor expressed in maturing guard cells but not in guard mother cells. SCAP1 regulates the expression of genes encoding key elements of stomatal functioning and morphogenesis, such as a K+ channel protein, MYB60 transcription factor, and pectin methyl esterase. Consequently, ion homeostasis was disturbed in scap1 guard cells, and esterification of extracellular pectins was impaired so that the cell walls lining the pores did not mature normally. We conclude that SCAP1 regulates essential processes of stomatal guard cell maturation and functions as a key transcription factor regulating the final stages of guard cell differentiation. We isolated guard cell protoplasts from 4-week-old WT(Col-0) and scap1 mutant plants and extracted RNA independently. Four biological replicates were performed for each experiment.
Project description:Plant salt glands are nature’s desalination devices that harbour potentially useful information pertaining to salt and water transport during secretion. As part of the program toward deciphering secretion mechanisms in salt glands, we adopted a shotgun approach to look into the proteome of salt-gland enriched tissues of the mangrove tree species Avicennia officinalis. To achieve this, we isolated the adaxial epidermal peels (which harbour the salt glands), and separated them from the mesophyll tissues of the leaves. Three biological replicates were prepared and total proteins were extracted from these tissues. Proteins that were found to be unique in salt gland-enriched tissues were obtained by eliminating proteins found in the mesophyll tissues. The proteins that are uniquely present in salt gland-enriched tissues were then selected and categorized by GO analysis.
Project description:Plants are often challenged by numerous biotic and abiotic environmental stresses. They have evolved efficient signaling networks in response to the environmental challenges, such as pathogen invasion, drought, CO2 changes. Guard cells are specialized epidermal cells enclosing pores on leaf surface, aka, stomata. Stomatal guard cells are at the frontline of biotic and abiotic stress responses. The opening and closing of stomata are regulated by a sophisticated intracellular signaling networks. Mitogen-activated protein kinase 4 (MPK4) was first identified as negative regulator in systemic acquired resistance (SAR). It was also play important roles in cytokinesis, reproduction, and photosynthesis. MPK4 has higher expression levels in plant guard cells than in mesophyll cells. Arabidopsis mpk4 mutant has higher levels of salicylic acid (SA) and reactive oxygen species (ROS), which are related with stomatal movement. The specific function of MPK4 in guard cells is unknown. In order to understand MPK4 functions in guard cells, it is essential to investigate MPK4-associated protein complex. Here we focus on identification of guard cell specific MPK4 complex using affinity purification following by mass spectrometry (AP-MS). Potential substrates were selected and validated with yeast two hybrid (Y2H).
Project description:Quinoa, like a large fraction of halophytes, use so-called bladder cells to detoxify excess salt. These leaf exterior structures are formed by specialized trichomes and consist of a leaf epidermal cell, a stalk cell and the bladder cell itself. Under salt stress, Na+ and Cl- as well as K+ and metabolites are imported from leaf sources and stored in the bladder. During this process the stalk cell simultaneously operates as both a selectivity filter and a flux controller. We submerged detached leaves in buffer, then lightly brushed the abaxial surface with a fine paintbrush to remove only the EBCs. The abaxial epidermis was then removed using tweezers and sampled in buffer for RNA extraction and RNA sequencing to separate the gene expression profiles of stalk cells from bladders and leaves.
Project description:Epithelial cells (EC) lining the most inner layers of secretory glands in Citrus peel are hypothesized to be the specialized cells that synthesize citrus essential oil. The major biosynthetic pathway(s) for essential oil are therefore postulated to be specifically and highly active in EC; transcripts that are involved in the pathway(s) are expected to be highly up-regulated in the cells as well. We performed cell-specific transcriptional analysis using GeneChip Citrus Genome Arrays to probe the global gene expression in EC during initial stage of essential oil biosynthesis and to identify groups of highly expressed genes in the EC. Grapefruits of two different sizes (diameters at 28 mm and 41 mm), at which points where biosynthesis of essential oil entering its active phase, were selected for the purpose of this experiment. For comparison, we selected parenchyma cells (PC), the non-oil-biosynthesizing cells, as the control cell type. Grapefruit peels were fixed, embedded in paraffin, sectioned and mounted on slides prior to cells isolation using laser microdissection and pressure catapulting. RNA was extracted from the isolated cells and then used for microarray hybridization. Transcriptional analysis were carried out by comparing transcript profiles between different cell types of the same time points, and same cell type at the different time points.
Project description:Heterotrimeric G proteins mediate crucial and diverse signaling pathways in eukaryotes. To gain insights into the regulatory modes of the G protein and the co-regulatory modes of the G protein and the stress hormone abscisic acid (ABA), we generated and analyzed gene expression in G protein subunit single and double mutants of the model plant Arabidopsis thaliana. Through a Boolean modeling approach, our analysis reveals novel modes of heterotrimeric G protein action. Keywords: transcriptome analysis; G protein subunit mutants; abscisic acid (ABA) Microarray data were generated from four genotypes (wild type, gpa1-4 mutant, agb1-2 mutant, agb1-2 gpa1-4 double mutant) with or without ABA treatment. Arabidopsis plants were grown in growth chambers with an 8 hr light/16hr dark. Three hundred Arabidopsis leaves excised from 60-70 five-week-old plants were used as the starting material for each guard cell microarray. Ten mature leaves taken from 3-4 plants grown side-by side with the plants for guard cell isolation were used for each leaf sample. Excised leaf and isolated guard cell samples were treated with ABA (50 μM) or EtOH (solvent control) for 3 hrs. For each type of sample (guard cells or leaves), three independent biological replicates were performed, resulting in a total of 48 microarray hybridizations (2 sample types ´ 4 genotypes ´ two treatments ´3 replicates).