Waksman Institute of Microbiology, Rutgers University, Piscataway, NJ 08854, USA.
Proc Natl Acad Sci U S A. 2010 Jul 20;107(29):12810-5. doi: 10.1073/pnas.1004721107. Epub 2010 Jul 6.
Essential amino acids like lysine and tryptophan are deficient in corn meal because of the abundance of zein storage proteins that lack these amino acids. A natural mutant, opaque 2 (o2) causes reduction of zeins, an increase of nonzein proteins, and as a consequence, a doubling of lysine levels. However, o2's soft inferior kernels precluded its commercial use. Breeders subsequently overcame kernel softness, selecting several quantitative loci (QTLs), called o2 modifiers, without losing the high-lysine trait. These maize lines are known as "quality protein maize" (QPM). One of the QTLs is linked to the 27-kDa gamma-zein locus on chromosome 7S. Moreover, QPM lines have 2- to 3-fold higher levels of the 27-kDa gamma-zein, but the physiological significance of this increase is not known. Because the 27- and 16-kDa gamma-zein genes are highly conserved in DNA sequence, we introduced a dominant RNAi transgene into a QPM line (CM105Mo2) to eliminate expression of them both. Elimination of gamma-zeins disrupts endosperm modification by o2 modifiers, indicating their hypostatic action to gamma-zeins. Abnormalities in protein body structure and their interaction with starch granules in the F1 with Mo2/+; o2/o2; gammaRNAi/+ genotype suggests that gamma-zeins are essential for restoring protein body density and starch grain interaction in QPM. To eliminate pleiotropic effects caused by o2, the 22-kDa alpha-zein, gamma-zein, and beta-zein RNAis were stacked, resulting in protein bodies forming as honeycomb-like structures. We are unique in presenting clear demonstration that gamma-zeins play a mechanistic role in QPM, providing a previously unexplored rationale for molecular breeding.
由于富含缺乏这些氨基酸的醇溶蛋白贮藏蛋白，玉米粉中的必需氨基酸（如赖氨酸和色氨酸）含量不足。一个天然的突变体，不透明 2（o2）导致醇溶蛋白减少，非醇溶蛋白增加，结果，赖氨酸水平增加了一倍。然而，o2 的软质胚乳使其无法商业化。随后，育种家克服了胚乳柔软的问题，选择了几个数量性状基因座（QTL），称为 o2 修饰基因，而不失去高赖氨酸特性。这些玉米品系被称为“优质蛋白玉米”（QPM）。其中一个 QTL 与染色体 7S 上的 27kDa γ-醇溶蛋白基因座相连。此外，QPM 品系的 27kDa γ-醇溶蛋白水平提高了 2-3 倍，但这种增加的生理意义尚不清楚。由于 27kDa 和 16kDa γ-醇溶蛋白基因在 DNA 序列上高度保守，我们将一个显性 RNAi 转基因引入到 QPM 品系（CM105Mo2）中，以消除它们的表达。γ-醇溶蛋白的消除扰乱了 o2 修饰物对胚乳的修饰，表明它们对 γ-醇溶蛋白具有基础作用。在 Mo2/+；o2/o2；γRNAi/+基因型的 F1 中，蛋白质体结构异常及其与淀粉粒的相互作用表明，γ-醇溶蛋白对于恢复 QPM 中蛋白质体密度和淀粉粒相互作用是必不可少的。为了消除 o2 引起的多效性影响，我们将 22kDa α-醇溶蛋白、γ-醇溶蛋白和β-醇溶蛋白 RNAis 进行了叠加，结果形成了类似蜂窝状的蛋白质体结构。我们是唯一明确证明γ-醇溶蛋白在 QPM 中起机械作用的，为分子育种提供了一个以前未探索的合理依据。
