Isolation and characterization of wheat triticin cDNA revealing a unique lysine-rich repetitive domain

Abstract

Polyclonal antibodies were raised against a purified 22 kDa triticin polypeptide (delta) and were used to screen a wheat seed cDNA library in the Escherichia coli expression vector lambda gt11. The isolated cDNA clones were grouped into three families based on their cross-hybridization reactions in DNA dot-blot studies. Southern blots of genomic DNAs extracted from ditelocentric and nullisomic-tetrasomic lines of Chinese Spring wheat, probes with the excised cDNA inserts, indicated that one of the three families (9 clones) had triticin clones. This was finally confirmed by comparing the predicted amino acid sequences of two of these clones (lambda Tri-12, lambda Tri-25) with the published tryptic peptide sequences of triticin. The Southern blots also showed that there is at least one triticin gene located on the short arm of each of the homoeologous group 1 chromosomes (1A, 1B, 1D), although till now no triticin protein product has been identified for the chromosome 1B. The nucleotide sequence of the largest triticin cDNA clone lambda Tri-25 (1567 bp) is presented here, and its predicted amino acid sequence shows strong homology with the legumin-like proteins of oats (12S globulin), rice (glutelin) and legume seeds. A unique feature of the triticin sequence is that it contains a lysine-rich repetitive domain, inserted in the hypervariable region of the typical legumin-like genes. Northern blotting of total RNA extracted from different stages of the developing wheat seed revealed that the triticin gene expression is switched on 5-10 days after anthesis (DAA). There was a steady increase in the level of triticin mRNA until 20 DAA, after which it started decreasing. The maximum mRNA accumulation occurred between 17 and 20 DAA. These observations conform closely with the published data on triticin protein accumulation during grain development.

Seed-storage-protein loci in RFLP maps of diploid, tetraploid, and hexaploid wheat

“…We report here the linkage relationships among genes encoding low-M glutenin and gliadin proteins in diploid, tetraploid and hexaploid wheats, using DNA hybridization. DNA markers for other storage proteins, such as high-M glutenins (Anderson et al 1989) and triticin (Singh et al 1993) were also included in this study.…”

Effect of Allelic Variation in Triticin on Bread- and Chapati-Making Qualities of Wheat (Triticum aestivum)

“…Triticin is a minor seed storage protein which accounts for only about 5 % of the total endosperm protein in wheat. It is leguminlike protein and has a lysine-rich repetitive domain in its hyper variable region which offers new opportunities to genetic engineers for increasing lysine content of wheat [36]. Our study found two alleles of triticin showing significant positive effect on bread-making quality parameters.…”

“…[3,25,28,29,31,32,36]. Triticin genes are located on the short arm of wheat chromosomes 1A and 1D, near the centromere far away from the major Gli-1/Glu-3 loci [28,31].…”

“…Triticin is synthesized specifically during the wheat seed development and is deposited in the electron dense inclusion bodies within the main storage protein bodies of the endosperm [3,29]. Unlike the glutenin and gliadin which are prolamin type proteins (rich in proline and glutamine amino acids), triticin shows homology to the 11-12S legumin-like storage globulins of leguminous species [32,36]. However, the role of triticin in determining wheat dough properties and bread-making quality has not yet been investigated.…”

Comparative proteomic analysis of wheat developing grains between Chinese Spring and 1Sl/1B substitution line

“…Wheat triticin polypeptides are synthesized specifically during seed development 8 to 21 days after anthesis, and are deposited in the endosperm protein bodies. It may play an important role in determining the functionality of wheat flour dough by participating in thioldisulphide inter-change reactions (Singh et al 1993). Our results from 2-DE also indicated that this protein accumulated at the early stages and reduced gradually at the later stages (Fig.…”