Transcriptional regulation of LMW glutenin genes were investigated in-silico, using publicly available gene sequences and expression data. Genes were grouped into different LMW glutenin types and their promoter profiles were determined using cis-acting regulatory elements databases and published results. The various cis-acting elements belong to some conserved non-coding regulatory regions (CREs) and might act in two different ways. There are elements, such as GCN4 motifs found in the long endosperm box that could serve as key factors in tissue-specific expression. Some other elements, such as the AACA/TA motifs or the individual prolamin box variants, might modulate the level of expression. Based on the promoter sequences and expression characteristic LMW glutenin genes might be transcribed following two different mechanisms. Most of the s- and i-type genes show a continuously increasing expression pattern. The m-type genes, however, demonstrate normal distribution in their expression profiles. Differences observed in their expression could be related to the differences found in their promoter sequences. Polymorphisms in the number and combination of cis-acting elements in their promoter regions can be of crucial importance in the diverse levels of production of single LMW glutenin gene types.

Analysis of gene expression data generated by high-throughput microarray transcript profiling experiments coupled with cis-regulatory elements enrichment study and cluster analysis can be used to define modular gene programs and regulatory networks. Unfortunately, the high molecular weight glutenin subunits of wheat (Triticum aestivum) are more similar than microarray data alone would allow to distinguish between the three homoeologous gene pairs. However, combining cDNA expression libraries with microarray data a co-expressional network was built that highlighted the hidden differences between these highly similar genes. Duplex clusters of cis-regulatory elements were used to focus the co-expressional network of transcription factors to the putative regulatory network of Glu-1 genes. The focused network helped to identify several modules of transcriptional gene programs in the endosperm. Many of these programs demonstrated a conserved temporal pattern across the studied genotypes, however few others showed variance. Based on this network, transient gene expression assays were performed with mutated promoters to inspect the control of tissue specificity. Results indicated that the interactions of the ABRE│CBF cluster with distal promoter regions may have a dual role in regulation by both recruiting the transcription complex as well as suppressing it in non-endosperm tissue. A putative model of regulation is discussed.

A B S T R A C T High molecular weight glutenin subunits of wheat are economically important seed storage proteins. They are coded by paralog pairs of the Glu-1 gene on each of the three genomes in the hexaploid wheat. Their expressions are under both temporal and spatial control. Many factors have been identified that influence the activity of Glu-1 genes, but the underlying regulatory mechanisms are still unclear. In order to identify motifs and motif clusters responsible for quantitative regulation of Glu-1 gene expressions, promoter profiles and transcription dynamics of the genes were analysed. It was found that promoter motif compositions of homoeolog Glu-1 genes are conserved. Our results demonstrated that while promoter profiles explain the differences of expression between homoeologs and between paralogs, it does not explain the variation of activity between alleles. Interestingly, our analyses revealed that the promoters of Glu-1 genes are divided into six cis-regulatory modules that are either locally overrepresented by binding sites belonging to unique but distinct transcription factor (TF) families or have conserved motif clusters. Moreover, our analyses demonstrated that the varying expression dynamics of TFs across genotypes is likely to be the primary contributor of the allelic variation of Glu-1 gene expressions. Thus, the six putative cis-regulatory modules in the Glu-1 gene promoters bound by the differentially expressed TFs are suggested to play a key role in the quantitative and tissue specific regulation of these genes. I N T R O D U C T I O N Wheat seed storage proteins (SSPs) are one of the primary sources of proteins in human diets and animal feed worldwide. These proteins are synthesized in the endosperm; a tissue specialized to starch and protein biosynthesis and storage. The prolamin superfamily of SSPs is a main component of wheat flour and their composition and ratio control dough properties, thus the quality of the end product. There are three main types of prolamin proteins: the sulphur (S) rich prolamins (alpha-, gamma-gliadins and low molecular weight (LMW) glutenins), sulphur poor prolamins (omega gliadins) and the high molecular weight (HMW) glutenins. Consequently, the quality of wheat dough is determined by the allele composition present in the genotypes and the proportion of the prolamin proteins thereby expressed.

<div> <p>Unprotected, primary 2-azidoamines are versatile precursors to vicinal diamines, which are among the most common motifs in biologically active compounds. Herein, we report their operationally simple synthesis through an iron-catalyzed difunctionalization of alkenes. A wide array of alkene substrates are tolerated, including complex drug-like molecules and a tripeptide. Facile derivatizations of the azidoamine group demonstrate the versatility of this masked diamine motif in chemoselective, orthogonal transformations. Applications of the methodology in the concise synthesis of RO 20-1724 and in a formal total synthesis of (±)-hamacanthin B further demonstrate the broad synthetic potential of this highly functional group tolerant reaction.</p> </div>

No abstract

Motivation: In silico enzymatic digestion tools mostly can be used for digestion of single sequence query, which means a significant limitation in their utility when a number of sequences need to be processed. The other limitation of these applications is the selection options of restriction enzymes that are usually allow only simultaneous digestion. Non-conventional proteins such as cereal prolamins require multienzyme multi step digestion, and for cereal proteomics experts this type of application is missing. Results: PDMQ, Protein Digestion Multi Query application was developed having multi query and multi enzyme options and that way can be customized for any digestion protocol. Availability and implementation: PDMQ is implemented in C# using the .NET framework and can be downloaded from

Wheat has been cultivated for 10000 years and ever since the origin of hexaploid wheat it has been exempt from natural selection. Instead, it was under the constant selective pressure of human agriculture from harvest to sowing during every year, producing a vast array of varieties. Wheat has been adopted globally, accumulating variation for genes involved in yield traits, environmental adaptation and resistance. However, one small but important part of the wheat genome has hardly changed: the regulatory regions of both the x- and y-type high molecular weight glutenin subunit (HMW-GS) genes, which are alone responsible for approximately 12% of the grain protein content. The phylogeny of the HMW-GS regulatory regions of the Triticeae demonstrates that a genetic bottleneck may have led to its decreased diversity during domestication and the subsequent cultivation. It has also highlighted the fact that the wild relatives of wheat may offer an unexploited genetic resource for the regulatory region of these genes. Significant research efforts have been made in the public sector and by international agencies, using wild crosses to exploit the available genetic variation, and as a result synthetic hexaploids are now being utilized by a number of breeding companies. However, a newly emerging tool of genome editing provides significantly improved efficiency in exploiting the natural variation in HMW-GS genes and incorporating this into elite cultivars and breeding lines. Recent advancement in the understanding of the regulation of these genes underlines the needs for an overview of the regulatory elements for genome editing purposes.

High molecular weight glutenin subunits (HMW GS) represent an important fraction of endosperm-specific seed-storage proteins that provide elasticity to bread dough. Previously, the second cis-regulatory module (CRM2) was found to be one of the most conserved part of HMW GS promoters, which indicated its pre-eminent role in their gene regulation. Here, we observed that deletion of CRM2 from the promoters of the Bx7 and By8 HMW GS genes increased the leakage of their transient expression in wheat leaf tissue. The effect of a VP1, an Myb and an antisense bZIP transcription factor (TF)-binding site, potentially involved in endosperm-specific regulation within CRM2, was then studied. The activity of a Bx7 gene promoter containing a mutant CRM2 with altered VP1 and Myb TF-binding sites, but an intact bZIP TF-binding site, was similarly low to that of the wild type in leaves. Transactivation analysis and EMSA indicated the binding of TFs TabZIP34 and TabZIP115 to the Skn-1 motif GTCAT in CRM2 and the repression of Bx7 and By8 HMW GS gene promoter activity in leaves. TabZIP34 and TabZIP115 may be involved in the downregulation of HMW GS gene expression in vegetative tissues and early-stage endosperm as well its modulation during seed maturation.