Wan Yongfang, Gritsch Cristina Sanchis, Hawkesford Malcolm J, Shewry Peter R
Department of Plant Biology and Crop Science, Rothamsted Research, Harpenden, Hertfordshire AL5 2JQ, UK.
Ann Bot. 2014 Mar;113(4):607-15. doi: 10.1093/aob/mct291. Epub 2013 Dec 15.
The ω-gliadin storage proteins of wheat are of interest in relation to their impact on grain processing properties and their role in food allergy, particularly the ω-5 sub-group and wheat-dependent exercise-induced anaphylaxis. The ω-gliadins are also known to be responsive to nitrogen application. This study therefore compares the effects of cultivar and nitrogen availability on the synthesis and deposition of ω-gliadins in wheat grown under field conditions in the UK, including temporal and spatial analyses at the protein and transcript levels.
SDS-PAGE, western blotting and N-terminal amino acid sequencing were used to compare the patterns of ω-gliadin components in mature grain of six British wheat (Triticum aestivum) cultivars and their accumulation during the development of grain grown in field plots with varying nitrogen supply. Changes in gene expression during development were determined using real-time reverse transcription-PCR (RT-PCR). Spatial patterns of gene expression and protein accumulation were determined by in situ hybridization and immunofluorescence microscopy, respectively.
Two patterns of ω-gliadins were identified in the six cultivars, including both monomeric 'gliadin' proteins and subunits present in polymeric 'glutenin' fractions. Increasing the level of nitrogen fertilizer in field plots resulted in increased expression of ω-gliadin transcripts and increased proportions of ω-5 gliadins. Nitrogen supply also affected the spatial patterns of ω-gliadin synthesis and deposition, which were differentially increased in the outer layers of the starchy endosperm with high levels of nitrogen.
Wheat ω-gliadins vary in amount and composition between cultivars, and in their response to nitrogen supply. Their spatial distribution is also affected by nitrogen supply, being most highly concentrated in the sub-aleurone cells of the starchy endosperm under higher nitrogen availability.
小麦的ω-醇溶蛋白贮藏蛋白因其对谷物加工特性的影响及其在食物过敏中的作用而受到关注，尤其是ω-5亚组和小麦依赖运动诱发的过敏反应。已知ω-醇溶蛋白对施氮有响应。因此，本研究比较了品种和氮素供应对英国田间种植小麦中ω-醇溶蛋白合成与沉积的影响，包括蛋白质和转录本水平的时空分析。
采用SDS-PAGE、western印迹和N端氨基酸测序，比较了6个英国小麦（普通小麦）品种成熟籽粒中ω-醇溶蛋白组分的模式及其在不同供氮量田间小区种植的籽粒发育过程中的积累情况。利用实时逆转录PCR（RT-PCR）测定发育过程中基因表达的变化。分别通过原位杂交和免疫荧光显微镜确定基因表达和蛋白质积累的空间模式。
在6个品种中鉴定出两种ω-醇溶蛋白模式，包括单体“醇溶蛋白”和聚合物“谷蛋白”组分中的亚基。增加田间小区的氮肥水平导致ω-醇溶蛋白转录本表达增加，ω-5醇溶蛋白比例增加。氮供应也影响ω-醇溶蛋白合成和沉积的空间模式，在高氮水平下，淀粉胚乳外层的这种模式差异增加。
小麦ω-醇溶蛋白在品种间的数量和组成以及对氮供应的响应方面存在差异。它们的空间分布也受氮供应的影响，在较高氮有效性下，它们高度集中在淀粉胚乳的亚糊粉层细胞中。
