Protocol to measure ribosome density along mRNA transcripts of Arabidopsis thaliana tissues using Ribo-seq.

Ribosome profiling (Ribo-seq) measures ribosome density along messenger RNA (mRNA) transcripts and is used to estimate the "translational fitness" of a given mRNA in response to environmental or developmental cues with high resolution. Here, we describe a protocol for Ribo-seq in plants adapted for the model plant Arabidopsis thaliana. We describe steps for lysis and nucleolytic digestion and ribosome footprinting.

Iron sensing in plants.

The ease of accepting or donating electrons is the raison d'être for the pivotal role iron (Fe) plays in a multitude of vital processes. In the presence of oxygen, however, this very property promotes the formation of immobile Fe(III) oxyhydroxides in the soil, which limits the concentration of Fe that is available for uptake by plant roots to levels well below the plant's demand. To adequately respond to a shortage (or, in the absence of oxygen, a possible surplus) in Fe supply, plants have to perceive and decode information on both external Fe levels and the internal Fe status.

Tandem Mass Tag-Based Phosphoproteomics in Plants.

Mass spectrometry-based proteomics provide a powerful tool for plant research, allowing global detection of steady-state levels of proteins under a given experimental setup. Here, we provide an optimized protocol for proteomic profiling using tandem mass tag (TMT) labeling followed by liquid chromatography-mass spectrometry (LC-MS/MS) to quantitate phosphopeptides and non-phosphopeptides from the same samples. The outlined protocol comprises a series of successive steps, namely, SDS (sodium dodecyl sulfate) protein extraction, protein precipitation, digestion, TMT labeling, phosphopeptide enrichment, high pH reversed-phase fractionation, LC-MS/MS analysis, protein identification, and data analysis.

Chromatin Enrichment for Proteomics in Plants (ChEP-P).

Chromatin enrichment for proteomics (ChEP) is a technique that allows for unbiased proteomic profiling of the chromatin landscape using mass spectrometry. While the method has been successfully employed to survey chromatin-associated proteins in various organisms and cell types, ChEP has not yet been applied to plant materials. Here, we describe a detailed ChEP protocol which has been modified for plants and designated ChEP-P (ChEP in plants).

How Plants Recalibrate Cellular Iron Homeostasis.

Insufficient iron supply poses severe constraints on plants, restricting species with inefficient iron uptake mechanisms from habitats with low iron availability and causing yield losses in agricultural ecosystems. Iron deficiency also poses a severe threat on human health. Anemia resulting from insufficient iron intake is affecting one of four people in the world.

Chromatin enrichment for proteomics in plants (ChEP-P) implicates the histone reader ALFIN-LIKE 6 in jasmonate signalling.

Background: Covalent modifications of core histones govern downstream DNA-templated processes such as transcription by altering chromatin structure and function. Previously, we reported that the plant homeodomain protein ALFIN-LIKE 6 (AL6), a bona fide histone reader that preferentially binds trimethylated lysin 4 on histone 3 (H3K4me3), is critical for recalibration of cellular phosphate (Pi) homeostasis and root hair elongation under Pi-deficient conditions.

Results: Here, we demonstrate that AL6 is also involved in the response of Arabidopsis seedlings to jasmonic acid (JA) during skotomorphogenesis, possibly by modulating chromatin dynamics that affect the transcriptional regulation of JA-responsive genes.

Scopoletin 8-Hydroxylase-Mediated Fraxetin Production Is Crucial for Iron Mobilization.

Iron (Fe) is an essential mineral nutrient and an important factor for the composition of natural plant communities. Low Fe availability in aerated soils with neutral or alkaline pH has led to the evolution of elaborate mechanisms that extract Fe from the soil solution. In Arabidopsis (), Fe is acquired by an orchestrated strategy that comprises mobilization, chelation, and reduction of Fe prior to its uptake.

Isobaric Tag for Relative and Absolute Quantitation (iTRAQ)-Based Protein Profiling in Plants.

Isobaric tags for relative and absolute quantitation (iTRAQ) is a technology that utilizes isobaric reagents to label the primary amines of peptides and proteins and is used in proteomics to study quantitative changes in the proteome by tandem mass spectrometry . Here, we present an adaptation of the iTRAQ experimental protocol for plants that allows the identification and quantitation of more than 12,000 plant proteins in Arabidopsis with a false discovery rate of less than 5 %.

The regulation and plasticity of root hair patterning and morphogenesis.

Root hairs are highly specialized cells found in the epidermis of plant roots that play a key role in providing the plant with water and mineral nutrients. Root hairs have been used as a model system for understanding both cell fate determination and the morphogenetic plasticity of cell differentiation. Indeed, many studies have shown that the fate of root epidermal cells, which differentiate into either root hair or non-hair cells, is determined by a complex interplay of intrinsic and extrinsic cues that results in a predictable but highly plastic pattern of epidermal cells that can vary in shape, size and function.

Role of plant-specific N-terminal domain of maize CK2β1 subunit in CK2β functions and holoenzyme regulation.

Protein kinase CK2 is a highly pleiotropic Ser/Thr kinase ubiquituous in eukaryotic organisms. CK2 is organized as a heterotetrameric enzyme composed of two types of subunits: the catalytic (CK2α) and the regulatory (CK2β). The CK2β subunits enhance the stability, activity and specificity of the holoenzyme, but they can also perform functions independently of the CK2 tetramer.

Specific characteristics of CK2β regulatory subunits in plants.

In all eukaryotes, the typical CK2 holoenzyme is an heterotetramer composed of two catalytic (CK2α and CK2α') and two regulatory (CK2β) subunits. One of the distinctive traits of plant CK2 is that they present a greater number of genes encoding for CK2α/β subunits than animals or yeasts, for instance, in Arabidopsis and maize both CK2α/β subunits belong to multigenic families composed by up to four genes. Here, we conducted a genome-wide survey examining 34 different plant genomes in order to investigate if the multigenic property of CK2β genes is a common feature through the entire plant kingdom.