普通玉米籽粒中是胺基酸組成不均衡，尤其是必需胺基酸——賴氨酸含量低，蛋白質品質較差。玉米胚乳中重要的轉錄調控因子Opaque 2 基因發生突變會使玉米中的賴氨酸含量大幅度上升，可極大地改善玉米籽粒的蛋白質品質，利用 opaque2 ( o2) 突變體是目前獲得高賴氨酸優質蛋白玉米的主要手段。但 o2玉米常會產生一些不良的農藝性狀，尤其會形成粉質胚乳。作為胚乳中重要的轉錄調控因子， o2 蛋白通過調控醇溶蛋白(Zeins)、澱粉合成（SSIII）以及丙酮酸磷酸雙激酶( PPDK)的表達，進而影響胚乳品質特性。明確o2蛋白的調控網絡對玉米品質改良具有重要意義。目前， Zeins和SSIII突變體的作用機制業已明確，但PPDK的調控機理還不完全清晰。
近日，愛荷華州立大學的科研人員首次報導了PPDK參與玉米胚乳品質性狀的調控模式。玉米中有兩個PPDK基因，分別為pdk1 和pdk2。研究人員通過構建pdk1 和 pdk2基因的單突變和雙突變，系統解析了PPDK基因在胚乳發育中的作用，結果顯示，PPDK 基因通過調控糖酵解通路，間接影響胚乳品質，而不影響籽粒生物量的積累。該研究表明，與之前通過調控轉錄因子進行玉米品質遺傳改良相比，通過精準調控具體的某個代謝路徑更為有效。
PNAS; published ahead of print December 18, 2017
Functions of maize genes encoding pyruvate phosphate dikinase in developing endosperm
Authors
Ryan R. Lappea, John W. Baierb, Susan K. Boehleinb, Ryan Huffmanc, Qiaohui Lina, Fabrice Wattebledd, A. Mark Settlesb, L. Curtis Hannahb, Ljudmilla Borisjuke, Hardy Rolletscheke, Jon D. Stewartf, M. Paul Scottc,g, Tracie A. Hennen-Bierwagena, and Alan M. Myersa,
Significance
Mutations affecting the transcription factor encoded by the gene o2 are important in maize agriculture because they result in improved grain nutritional quality. The mutations also cause detrimental effects by reducing kernel hardness and diminishing agronomic quality and food applications. The undesirable characteristics are not fully understood because the o2 product regulates multiple targets that could contribute to the phenotype. This study investigated one target that had not been previously mutated, pyruvate phosphate dikinase (PPDK), and showed that PPDK deficiency in isolation causes the negative phenotype associated with reduced kernel hardness. Thus, maize improvement may be better accomplished by targeting individual metabolic pathways determining protein and amino acid balance rather than pleiotropic regulators such as the o2 transcription factor.
Abstract
Maize opaque2 (o2) mutations are beneficial for endosperm nutritional quality but cause negative pleiotropic effects for reasons that are not fully understood. Direct targets of the bZIP transcriptional regulator encoded by o2 include pdk1 and pdk2 that specify pyruvate phosphate dikinase (PPDK). This enzyme reversibly converts AMP, pyrophosphate, and phosphoenolpyruvate to ATP, orthophosphate, and pyruvate and provides diverse functions in plants. This study addressed PPDK function in maize starchy endosperm where it is highly abundant during grain fill. pdk1 and pdk2 were inactivated individually by transposon insertions, and both genes were simultaneously targeted by endosperm-specific RNAi. pdk2 accounts for the large majority of endosperm PPDK, whereas pdk1 specifies the abundant mesophyll form. The pdk1- mutation is seedling-lethal, indicating that C4 photosynthesis is essential in maize. RNAi expression in transgenic endosperm eliminated detectable PPDK protein and enzyme activity. Transgenic kernels weighed the same on average as nontransgenic siblings, with normal endosperm starch and total N contents, indicating that PPDK is not required for net storage compound synthesis. An opaque phenotype resulted from complete PPDK knockout, including loss of vitreous endosperm character similar to the phenotype conditioned by o2-. Concentrations of multiple glycolytic intermediates were elevated in transgenic endosperm, energy charge was altered, and starch granules were more numerous but smaller on average than normal. The data indicate that PPDK modulates endosperm metabolism, potentially through reversible adjustments to energy charge, and reveal that o2- mutations can affect the opaque phenotype through regulation of PPDK in addition to their previously demonstrated effects on storage protein gene expression.
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