ProtView: A Versatile Tool for Protease Evaluation and Selection in a Proteomic and Proteogenomic Context.

Many tools have been created to generate proteome digests with different protease enzymes and provide useful information for selecting optimal digest schemes for specific needs. This can save on time and resources and generate insights on the observable proteome. However, there remains a need for a tool that evaluates digest schemes beyond protein and amino acid coverages in the proteomic domain.

Hypothyroidism impairs development of the gastrointestinal tract in the ovine fetus.

Growth and maturation of the fetal gastrointestinal tract near term prepares the offspring for the onset of enteral nutrition at birth. Structural and functional changes are regulated by the prepartum rise in cortisol in the fetal circulation, although the role of the coincident rise in plasma tri-iodothyronine (T3) is unknown. This study examined the effect of hypothyroidism on the structural development of the gastrointestinal tract and the activity of brush-border digestive enzymes in the ovine fetus near term.

The proteome of developing barley anthers during meiotic prophase I.

Flowering plants reproduce sexually by combining a haploid male and female gametophyte during fertilization. Male gametophytes are localized in the anthers, each containing reproductive (meiocyte) and non-reproductive tissue necessary for anther development and maturation. Meiosis, where chromosomes pair and exchange their genetic material during a process called recombination, is one of the most important and sensitive stages in breeding, ensuring genetic diversity.

Barley Anther and Meiocyte Transcriptome Dynamics in Meiotic Prophase I.

In flowering plants, successful germinal cell development and meiotic recombination depend upon a combination of environmental and genetic factors. To gain insights into this specialized reproductive development program we used short- and long-read RNA-sequencing (RNA-seq) to study the temporal dynamics of transcript abundance in immuno-cytologically staged barley () anthers and meiocytes. We show that the most significant transcriptional changes in anthers occur at the transition from pre-meiosis to leptotene-zygotene, which is followed by increasingly stable transcript abundance throughout prophase I into metaphase I-tetrad.

Application of a Sensitive and Reproducible Label-Free Proteomic Approach to Explore the Proteome of Individual Meiotic-Phase Barley Anthers.

Meiosis is a highly dynamic and precisely regulated process of cell division, leading to the production of haploid gametes from one diploid parental cell. In the crop plant barley (), male meiosis occurs in anthers, in specialized cells called meiocytes. Barley meiotic tissue is scarce and not easily accessible, making meiosis study a challenging task.

Identification and rapid mapping of a gene conferring broad-spectrum late blight resistance in the diploid potato species Solanum verrucosum through DNA capture technologies.

A broad-spectrum late blight disease-resistance gene from Solanum verrucosum has been mapped to potato chromosome 9. The gene is distinct from previously identified-resistance genes. We have identified and characterised a broad-spectrum resistance to Phytophthora infestans from the wild Mexican species Solanum verrucosum.

A high quality Arabidopsis transcriptome for accurate transcript-level analysis of alternative splicing.

Alternative splicing generates multiple transcript and protein isoforms from the same gene and thus is important in gene expression regulation. To date, RNA-sequencing (RNA-seq) is the standard method for quantifying changes in alternative splicing on a genome-wide scale. Understanding the current limitations of RNA-seq is crucial for reliable analysis and the lack of high quality, comprehensive transcriptomes for most species, including model organisms such as Arabidopsis, is a major constraint in accurate quantification of transcript isoforms.

Arabidopsis PTB1 and PTB2 proteins negatively regulate splicing of a mini-exon splicing reporter and affect alternative splicing of endogenous genes differentially.

This paper examines the function of Arabidopsis thaliana AtPTB1 and AtPTB2 as plant splicing factors. The effect on splicing of overexpression of AtPTB1 and AtPTB2 was analysed in an in vivo protoplast transient expression system with a novel mini-exon splicing reporter. A range of mutations in pyrimidine-rich sequences were compared with and without AtPTB and NpU2AF65 overexpression.

Plant SILAC: stable-isotope labelling with amino acids of arabidopsis seedlings for quantitative proteomics.

Stable Isotope Labelling by Amino acids in Cell culture (SILAC) is a powerful technique for comparative quantitative proteomics, which has recently been applied to a number of different eukaryotic organisms. Inefficient incorporation of labelled amino acids in cell cultures of Arabidopsis thaliana has led to very limited use of SILAC in plant systems. We present a method allowing, for the first time, efficient labelling with stable isotope-containing arginine and lysine of whole Arabidopsis seedlings.

Alternative splicing and nonsense-mediated decay modulate expression of important regulatory genes in Arabidopsis.

Alternative splicing (AS) coupled to nonsense-mediated decay (NMD) is a post-transcriptional mechanism for regulating gene expression. We have used a high-resolution AS RT-PCR panel to identify endogenous AS isoforms which increase in abundance when NMD is impaired in the Arabidopsis NMD factor mutants, upf1-5 and upf3-1. Of 270 AS genes (950 transcripts) on the panel, 102 transcripts from 97 genes (32%) were identified as NMD targets.

Regulation of plant gene expression by alternative splicing.

AS (alternative splicing) is a post-transcriptional process which regulates gene expression through increasing protein complexity and modulating mRNA transcript levels. Regulation of AS depends on interactions between trans-acting protein factors and cis-acting signals in the pre-mRNA (precursor mRNA) transcripts, termed 'combinatorial' control. Dynamic changes in AS patterns reflect changes in abundance, composition and activity of splicing factors in different cell types and in response to cellular or environmental cues.

Involvement of the nuclear cap-binding protein complex in alternative splicing in Arabidopsis thaliana.

The nuclear cap-binding protein complex (CBC) participates in 5' splice site selection of introns that are proximal to the mRNA cap. However, it is not known whether CBC has a role in alternative splicing. Using an RT-PCR alternative splicing panel, we analysed 435 alternative splicing events in Arabidopsis thaliana genes, encoding mainly transcription factors, splicing factors and stress-related proteins.

Aberrant mRNA transcripts and the nonsense-mediated decay proteins UPF2 and UPF3 are enriched in the Arabidopsis nucleolus.

The eukaryotic nucleolus is multifunctional and involved in the metabolism and assembly of many different RNAs and ribonucleoprotein particles as well as in cellular functions, such as cell division and transcriptional silencing in plants. We previously showed that Arabidopsis thaliana exon junction complex proteins associate with the nucleolus, suggesting a role for the nucleolus in mRNA production. Here, we report that the plant nucleolus contains mRNAs, including fully spliced, aberrantly spliced, and single exon gene transcripts.

Alternative splicing in plants.

The impact of AS (alternative splicing) is well-recognized in animal systems as a key regulator of gene expression and proteome complexity. In plants, AS is of growing importance as more genes are found to undergo AS, but relatively little is known about the factors regulating AS or the consequences of AS on mRNA levels and protein function. We have established an accurate and reproducible RT (reverse transcription)-PCR system to analyse AS in multiple genes.

Proteomic analysis of the Arabidopsis nucleolus suggests novel nucleolar functions.

The eukaryotic nucleolus is involved in ribosome biogenesis and a wide range of other RNA metabolism and cellular functions. An important step in the functional analysis of the nucleolus is to determine the complement of proteins of this nuclear compartment. Here, we describe the first proteomic analysis of plant (Arabidopsis thaliana) nucleoli, in which we have identified 217 proteins.

Determinants of plant U12-dependent intron splicing efficiency.

Factors affecting splicing of plant U12-dependent introns have been examined by extensive mutational analyses in an in vivo tobacco (Nicotiana tabacum) protoplast system using introns from three different Arabidopsis thaliana genes: CBP20, GSH2, and LD. The results provide evidence that splicing efficiency of plant U12 introns depends on a combination of factors, including UA content, exon bridging interactions between the U12 intron and flanking U2-dependent introns, and exon splicing enhancer sequences (ESEs). Unexpectedly, all three plant U12 introns required an adenosine at the upstream purine position in the branchpoint consensus UCCUURAUY.

Cloning and characterization of two subunits of Arabidopsis thaliana nuclear cap-binding complex.

In this report we characterize two Arabidopsis thaliana proteins, named AtCBP20 and AtCBP80, that are homologues of human subunits of a nuclear cap-binding protein complex (CBC). AtCBP20 has a calculated molecular mass of 29.9 kDa, and AtCBP80 is a 96.