Wheat proteomics: relationship between fine chromosome deletion and protein expression.
To explore the relationship between fine chromosome deletion and protein expression in common wheat, the changes in protein composition of wheat seed proteome were investigated by using chromosome 1B. A momosomic alien chromosome addition line of common wheat was used to produce the fine deletion lines. Endosperm and embryo proteins were separated by two-dimensional gel electrophoresis (2-DE) and visualized by staining with Commassie Brilliant Blue, and gel images were analyzed with a computer assisted image analyzer. For the first time, fine gene locations of a few endosperm and embryo proteins were identified on the chromosome 1B. These proteins with their specific gene location on the chromosome can be used as protein markers in breeding programs for quality of wheat proteins. To identify wheat seed proteins and to understand their expression in relation to chromosome deletion, the feasibility of a new analytical approach based on isotope coded affinity tag labeling (ICAT) of peptides in tryptic digests followed by electrospray ionization mass spectrometry has been described. Simplification of the complex tryptic digest prior to mass spectral analysis was performed by treating the samples with light and heavy ICAT labeling reagents. A clear separation of peptide fragment containing the light and heavy reagents was achieved in mass spectral analysis. Out of the 14 peptides detected by mass fragment analysis of the euploid, four were down-regulated, nine up-regulated and one did not show any change due to the terminal deletion of chromosome 1B. Selected peptide fragments were subjected to tandem mass spectrometry analysis for sequence information and the resulting sequence information was submitted to databases for protein identification. Of the five proteins submitted, four were identified as alpha-amylase inhibitor, alpha-amylase/subtilisin inhibitor precursor, proteasome subunit alpha-type 7 and 1,4 alpha-glucan-D-maltohydrolase. With this approach it is possible to identify wheat seed proteins and to understand their expression, which have been reported to be difficult by 2-DE due to cosynthesis of proteins by genes from three genomes, A, B and D.
Islam
Nazrul
N
Kihara Institute for Biological Research, Yokohama City University, Japan.
Tsujimoto
Hisashi
H
Hirano
Hisashi
H
D002872:Chromosome Deletion; D032461:Chromosomes, Plant; D015536:Down-Regulation; D015180:Electrophoresis, Gel, Two-Dimensional; D018745:Genome, Plant; D013058:Mass Spectrometry; D010455:Peptides / Q000737:chemistry; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics*; D020543:Proteome; D012394:Rosaniline Dyes / Q000494:pharmacology; D021241:Spectrometry, Mass, Electrospray Ionization; D014908:Triticum / Q000378:metabolism*; D014357:Trypsin / Q000494:pharmacology; D015854:Up-Regulation
D010455:Peptides; D010940:Plant Proteins; D020543:Proteome; D012394:Rosaniline Dyes; D014357:Trypsin; C004692:coomassie Brilliant Blue