植物Ⅰ类H+-PPase基因调控花青素积累的分子机制研究

The function of plant type I H+-PPases are gradually being recognized. Up-regulation of type I H+-PPases triggers enhanced growth/biomass and photosynthetic capacity in a variety of agriculturally important crops grown under normal or stressful conditions such as nutrient limitations, water scarcity, and salinity. AVP1 plays a fundamental role in phloem loading and transport of photoassimilates in Arabidopsis, some AVP1 mutants showed Suc-dependent phenotypes. Our study demonstrated that overexpression of MtVP1 in potato and Arabidopsis resulted in increased anthocyanin biosynthesis. We speculated that these phenotypes were related to transport of sucrose, but the molecular mechanism remains unclear. Based on our previous experimental results, we will design MS medium containing all kinds of sugars to investigate the changes of anthocyanin biosynthesis in transgenic Arabidopsis overexpressing MtVP1, the expression profiles of transgenic and wild type Arabidopsis will be examined by RNA-Seq quantification, the qPCR was applied to study the changes of gene expression related to sucrose transport and anthocyanin biosynthesis, in order to verify the expression profile data. Finally, we could preliminarily figure out the molecular mechanism of modulation the anthocyanin accumulation of plant type I H+-PPase gene.

植物Ⅰ类H+-PPase的功能正逐步被人们所认识。不论是正常条件还是胁迫条件（如营养胁迫、水分胁迫和盐胁迫）下，在一系列重要的作物中上调H+-PPase基因的表达均会导致转基因株系生长加快、生物量增大以及光合能力增强。在拟南芥中，AVP1在韧皮部装载和光合产物的转运方面具有重要作用，一些AVP1突变体表现出蔗糖依赖的表型。我们的研究表明过表达MtVP1能促进马铃薯和拟南芥的花青素积累，我们推测上述表型跟蔗糖的转运相关，但其分子机制尚不清楚。本项目拟在前期工作基础上，设计各种含糖的MS培养基研究转MtVP1拟南芥花青素积累的变化，利用RNA-Seq技术分析比较转MtVP1拟南芥和野生型拟南芥的差异表达基因，并利用qPCR验证跟蔗糖转运和花青素积累相关的差异表达基因，最终初步勾画出植物Ⅰ类H+-PPase基因调控花青素积累的分子机制。

研究背景：前人研究发现I型焦磷酸酶（H+-PPase）支持糖异生和蔗糖水解。外源添加蔗糖不能挽救avp1-1突变体。然而，其他突变体可以被蔗糖挽救，葡萄糖模拟了蔗糖的作用，蔗糖可以挽救fugu5突变体的表型，但不能被果糖挽救。因此，I型H+-PPase可能有其他机制影响糖代谢。.方法与结果：我们在马铃薯和拟南芥中过表达了截形苜蓿焦磷酸酶基因（MtVP1），并观察了多种条件下地表型变化。我们用HPIC-MS测定了糖含量，用转录组测序分析了转录谱，并用实时荧光定量PCR进行了验证。转MtVP1基因马铃薯茎红色，花紫色，毛状体发达。单株块茎数减少，块茎收缩速度加快。转基因马铃薯块茎中淀粉、葡萄糖和果糖含量下降，芽中葡萄糖和果糖含量也下降。果糖降低了马铃薯和拟南芥的花青素含量，转基因拟南芥的枝条呈冻伤状。此外，还检测了果糖-1,6-二磷酸酶和果糖-2,6-二磷酸酶基因的上调。.科学意义：过表达的I型H+-PPase基因通过上调果糖-1,6-二磷酸酶和果糖-2,6-二磷酸酶基因，可有条件地显著影响糖代谢，尤其是果糖代谢。