Abstract
BackgroundCeliac disease (CD) is caused by specific sequences of gluten proteins found in cereals such as bread wheat (Triticum aestivum ssp. aestivum) and spelt (T. aestivum ssp. spelta). Among them, the α-gliadins display the highest immunogenicity, with four T-cell stimulatory epitopes. The toxicity of each epitope sequence can be reduced or even suppressed according to the allelic form of each sequence. One way to address the CD problem would be to make use of this allelic variability in breeding programs to develop safe varieties, but tools to track the presence of toxic epitopes are required. The objective of this study was to develop a tool to accurately detect and quantify the immunogenic content of expressed α-gliadins of spelt and bread wheat.ResultsFour TaqMan probes that only hybridize to the canonical—i.e. toxic—form of each of the four epitopes were developed and their specificity was demonstrated. Six TaqMan probes targeting stable reference genes were also developed and constitute a tool to normalize qPCR data. The probes were used to measure the epitope expression levels of 11 contrasted spelt accessions and three ancestral diploid accessions of bread wheat and spelt. A high expression variability was highlighted among epitopes and among accessions, especially in Asian spelts, which showed lower epitope expression levels than the other spelts. Some discrepancies were identified between the canonical epitope expression level and the global amount of expressed α-gliadins, which makes the designed TaqMan probes a useful tool to quantify the immunogenic potential independently of the global amount of expressed α-gliadins.ConclusionsThe results obtained in this study provide useful tools to study the immunogenic potential of expressed α-gliadin sequences from Triticeae accessions such as spelt and bread wheat. The application of the designed probes to contrasted spelt accessions revealed a high variability and interesting low canonical epitope expression levels in the Asian spelt accessions studied.
Highlights
Celiac disease (CD) is caused by specific sequences of gluten proteins found in cereals such as bread wheat (Triticum aestivum ssp. aestivum) and spelt (T. aestivum ssp. spelta)
The specificity of each probe was confirmed by qPCR analyses: a fluorescent signal was clearly observable with the α-gliadin clones displaying the canonical epitope, while it was insignificant or absent with clones containing allelic variants (Fig. 1)
The specificity of the probes was further confirmed on the cDNA of the three diploid species representative of the ancestral genomes (A, B and D) of spelt and bread wheat
Summary
Celiac disease (CD) is caused by specific sequences of gluten proteins found in cereals such as bread wheat (Triticum aestivum ssp. aestivum) and spelt (T. aestivum ssp. spelta). Gluten is a group of water-insoluble proteins which display useful properties for dough formation These proteins are found in the seed of cereals such as bread wheat WA involves immunoglobulins (Ig)-E that link to repeat sequences in wheat proteins whereas CD is an autoimmune disorder with a genetic predisposition of the patient, mediated by Ig-A and Ig-G antibodies Beside these two disorders, a novel pathologic entity—called non-celiac gluten sensitivity (NCGS)—has gained importance and involves neither autoimmune nor allergic mechanisms. A novel pathologic entity—called non-celiac gluten sensitivity (NCGS)—has gained importance and involves neither autoimmune nor allergic mechanisms People affected by this illness display symptoms similar to CD but with usually normal small intestinal histology. Intra- and extraintestinal symptoms are encountered like diarrhea, bowel pain, fatigue, weight loss, anemia, osteoporosis, headaches and growth retardation [5, 6, 10, 11]
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